KR20210023842A - 시알릴화 사카라이드의 발효 생산 - Google Patents
시알릴화 사카라이드의 발효 생산 Download PDFInfo
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Abstract
시알릴화 사카라이드의 발효 생산 방법 및 상기 방법에서 사용하기 위한 유전자 조작된 미생물 세포 (여기서, 유전자 조작된 미생물 세포는 시알릴화 사카라이드를 생산하기 위한, (i) 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함하는 시알산 생합성 경로, (ii) 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제; 및 (iii) 시알릴트랜스퍼라제를 포함한다), 뿐만 아니라 영양 조성물을 제공하기 위한 상기 시알릴화 올리고사카라이드의 용도가 개시된다.
Description
배경
본 발명은 시알릴화 사카라이드의 발효 생산 방법, 뿐만 아니라 그에 사용된 재조합 또는 유전자 조작된 미생물 세포(microbial cell)에 관한 것이다.
현재까지 150개 초과의 구조적으로 구별되는 모유 올리고사카라이드 (HMO)가 확인되었다. 비록 HMO는 모유의 총 영양소 중 단지 적은 양을 나타내긴 하지만, 모유 수유 유아의 발달에 대한 그의 유익한 효과는 지난 수십년에 걸쳐 분명해졌다.
HMO 중에서, 시알릴화 HMO (SHMO)는 장병원성(enteropathogenic) 박테리아 및 바이러스에 대한 내성을 뒷받침하는 것으로 관찰되었다. 흥미롭게도, 최근 연구는 미숙아에서 가장 흔하고 치명적인 질환 중 하나인 괴사성 장염(necrotizing enterocolitis)에 대한 장쇄 SHMO의 보호 효과를 추가로 입증하였다. 게다가, SHMO는 유아의 두뇌 발달 및 그의 인지 능력을 뒷받침하는 것으로 여겨진다. 또한, 시알릴화 올리고사카라이드는 에스케리치아 콜라이(Escherichia coli), 비브리오 콜레라(Vibrio cholerae) 및 살모넬라(Salmonella)를 포함한 다양한 병원성 미생물의 장독소를 중화시키는 것으로 나타났다. 추가로, 시알릴화 올리고사카라이드는 헬리코박터 파일로리(Helicobacter pylori)에 의한 장(gut)의 군집화를 방해하여 위 및 십이지장 궤양을 예방 또는 억제하는 것으로 밝혀졌다.
시알릴화 올리고사카라이드 중에, 3'-시알릴락토스, 6'-시알릴락토스, 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c 및 디시알릴락토-N-테트라오스가 모유에서 가장 널리 사용되는 구성원이다.
시알릴화 올리고사카라이드는 복잡한 구조를 가지고 있기 때문에, 그의 화학 또는 (화학-)효소 합성은 도전적이고 광범위한 어려움, 예를 들어 입체화학의 제어, 특이적 연결의 형성, 공급원료의 가용성 등과 연관된다. 결과적으로, 시판되는 시알릴화 올리고사카라이드는 천연 공급원의 그의 적은 양으로 인해 매우 고가였다.
따라서, 시알릴화 올리고사카라이드를 생산하기 위한 미생물의 대사 공학에 대한 노력이 이루어져 왔는데, 그 이유는 이러한 접근법이 산업적 규모로 HMO를 생산하기 위한 가장 유망한 방법이기 때문이다. 미생물 발효에 의한 SHMO의 생산을 위해, 미생물은 전형적으로 외인성 시알산의 존재 하에 배양된다.
국제 공개 WO 2007/101862 A1은 배양 배지에서 미생물을 배양함으로써 세포내 UDP-GlcNAc 풀(pool)에 의존하는 시알릴화 올리고사카라이드의 대규모 생체내 합성 방법을 개시하며, 여기서 상기 미생물은 CMP-Neu5Ac 신테타제(synthetase), 시알산 신타제, GlcNAc-6-포스페이트 2 에피머라제 및 시알릴트랜스퍼라제를 코딩하는 이종유래(heterologous) 유전자를 포함한다. 게다가, 시알산 알돌라제 (NanA) 및 ManNac 키나제 (NanK)를 코딩하는 내인성 유전자는 결실되었다.
국제 공개 WO 2014/153253 A1은 박테리아를 조작하여 시알릴화 올리고사카라이드를 생산하는 방법 및 조성물뿐만 아니라 박테리아에서 시알릴화 올리고사카라이드를 생산하는 방법을 개시하며, 상기 박테리아는 외인성 시알릴트랜스퍼라제, 결핍된 시알산 이화 경로, 시알산 합성 능력, 및 기능성 락토스 퍼미아제 유전자를 포함하고, 여기서 상기 박테리아는 락토스의 존재 하에 배양된다. 시알산 합성 능력은 외인성 CMP-Neu5Ac 신테타제, 외인성 시알산 신타제, 및 외인성 UDP-GlcNAc-2-에피머라제를 발현하는 것을 포함한다.
그러나, 발효 동안 외인성 시알산의 존재 및/또는 첨가를 필요로 하지 않는 미생물 발효에 의해 시알릴화 올리고사카라이드를 생산하는 것이 바람직하다. 또한, UDP-N-아세틸글루코사민 (UDP-GlcNAc)의 세포내 풀에 접근할 필요가 없는 미생물에 의해 시알릴화 올리고사카라이드를 생산하는 것이 바람직하며, 그 이유는 이것이 세포에 에너지적으로 유익하다고 여겨지기 때문이다.
요약
상기 목적은, 특히, 외인성 시알산을 첨가할 필요가 없는, 시알릴화 사카라이드의 전체 세포 발효 생산 방법을 제공함으로써, 그리고 외인성 시알산의 부재 하에 시알릴화 사카라이드를 합성할 수 있는 유전자 조작된 미생물 세포에 의해 해결된다.
한 측면에 따르면, 시알릴화 사카라이드의 생산 방법이 제공되며, 이 방법은 a) (i) N-아세틸뉴라민산 (Neu5Ac, NeuNAc)의 세포내 생합성을 위한 시알산 생합성 경로 (여기서 상기 시알산 생합성 경로는 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함한다) (ii) 시티딘 5'-모노포스포-(CMP)-시알산 신테타제, 및 (iii) 이종유래 시알릴트랜스퍼라제를 포함하는 적어도 하나의 유전자 조작된 미생물 세포를 제공하는 단계; b) 적어도 하나의 유전자 조작된 미생물 세포를 발효 브로쓰에서 그리고 상기 시알릴화 사카라이드의 생산을 위해 허용되는 조건 하에 배양하는 단계; 및 임의로 c) 상기 시알릴화 사카라이드를 회수하는 단계를 포함한다.
또 다른 측면에 따르면, 시알릴화 사카라이드를 생산하기 위한 유전자 조작된 미생물 세포가 제공되며, 여기서 미생물 세포는 (i) N-아세틸뉴라민산의 세포내 생합성을 위한 시알산 생합성 경로 (여기서 상기 시알산 생합성 경로는 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함한다); (ii) N-아세틸뉴라민산을 시티딘 5'-모노포스페이트로 전달하여 CMP-활성화된 N-아세틸뉴라민산을 생성하기 위한 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제; 및 (iii) 이종유래 시알릴트랜스퍼라제를 포함한다.
또 다른 측면에 따르면, 본 발명에 따른 방법 또는 유전자 조작된 미생물 세포에 의해 생산될 수 있는 시알릴화 사카라이드가 제공된다.
또 다른 측면에 따르면, 영양 조성물, 바람직하게는 유아용 조성물을 제조하기 위해 본 발명에 따른 방법에 의해 생산된 시알릴화 사카라이드 또는 유전자 조작된 미생물 세포의 용도가 제공된다.
또 다른 측면에 따르면, 본 발명에 따른 방법에 의해 생산된 적어도 하나의 시알릴화 사카라이드 또는 본 발명에 따른 유전자 조작된 미생물 세포를 함유하는 영양 조성물이 제공된다.
도 1은 시알릴화 사카라이드의 발효 생산을 위해 유전자 조작된 미생물 세포에 의해 사용될 수 있는 시알산 생합성 경로의 개략도이며, 여기서 상기 시알산 생합성 경로는 UDP-GlcNAc를 이용한다.
도 2는 시알릴화 사카라이드의 발효 생산을 위해 본 발명의 유전자 조작된 미생물 세포에 의해 사용될 수 있는 시알산 생합성 경로의 개략도이다.
도 3은 시알릴화 사카라이드의 발효 생산을 위해 본 발명의 유전자 조작된 미생물 세포에 의해 사용될 수 있는 또 다른 시알산 생합성 경로의 개략도이다.
도 2는 시알릴화 사카라이드의 발효 생산을 위해 본 발명의 유전자 조작된 미생물 세포에 의해 사용될 수 있는 시알산 생합성 경로의 개략도이다.
도 3은 시알릴화 사카라이드의 발효 생산을 위해 본 발명의 유전자 조작된 미생물 세포에 의해 사용될 수 있는 또 다른 시알산 생합성 경로의 개략도이다.
제1 측면에 따르면, 시알릴화 사카라이드의 발효 생산 방법이 제공된다. 상기 방법은 a) 시알릴화 사카라이드를 합성할 수 있는 적어도 하나의 유전자 조작된 미생물 세포를 제공하는 단계이며, 상기 적어도 하나의 유전자 조작된 미생물 세포가 (i) 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함하는 시알산 생합성 경로; (ii) 시티딘 5'-모모포포-(CMP)-N-아세틸뉴라민산 신테타제; 및 (iii) 이종유래 시알릴트랜스퍼라제를 포함하는 것인 단계; b) 적어도 하나의 유전자 조작된 미생물 세포를 발효 브로쓰에서 그리고 상기 시알릴화 사카라이드의 생산을 위해 허용되는 조건 하에 배양하는 단계, 및 임의로 c) 상기 시알릴화 사카라이드를 회수하는 단계를 포함한다.
따라서, 제2 측면에서, 본 발명은 또한 시알릴화 사카라이드의 발효 생산을 위한 유전자 조작된 미생물 세포에 관한 것이며, 여기서 미생물 세포는 (i) N-아세틸뉴라민산의 세포내 생합성을 위한 시알산 생합성 경로 (여기서 상기 시알산 생합성 경로는 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함한다); (ii) N-아세틸뉴라민산을 시티딘 5'-모노포스페이트로 전달하여 CMP-활성화된 N-아세틸뉴라민산을 생성하기 위한 시티딘 5'-모노포스포-(CMP)-시알산 신테타제; 및 (iii) 시알릴화 사카라이드의 세포내 생합성을 결과하는, N-아세틸뉴라민산 모이어티를 공여자 기질로서 CMP-활성화된 시알산으로부터 수용체(acceptor) 분자 (이 수용체 분자는 사카라이드 분자이다)로 전달하기 위한 시알릴 트랜스퍼라제를 포함한다.
유전자 조작된 미생물 세포는 UDP-GlcNAc를 이용하지 않는 N-아세틸뉴라민산의 세포내 생합성을 위한 시알산 생합성 경로를 포함한다. 유전자 조작된 미생물 세포는 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제인 N-아세틸뉴라민산의 세포내 생합성을 위한 시알산 생합성 경로를 포함한다. N-아세틸뉴라민산의 세포내 생합성을 위해 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 사용하는 시알산 생합성 경로는 시알산의 생합성을 위해 UDP-GlcNAc를 이용하지 않는다 (도 2 및 도 3).
시알산 생합성 경로는 글루타민: 프럭토스-6-포스페이트 아미노트랜스퍼라제 및 N-아세틸뉴라민산 신타제의 효소 활성을 포함한다. 시알산 생합성 경로는 a) 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제, N-아세틸글루코사민-6-포스페이트 포스파타제 및 N-아세틸글루코사민 2-에피머라제의 효소 활성 (도 2); 및/또는 b) 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제, N-아세틸글루코사민-6-포스페이트 에피머라제 및 N-아세틸만노사민-6-포스페이트 포스파타제의 효소 활성 (도 3)을 추가로 포함한다. 따라서, 유전자 조작된 미생물 세포가 세포내 시알산 생합성을 위해 UDP의 수반되는 방출과 함께 포스포글루코사민 뮤타제, N-아세틸글루코사민-1-포스페이트 우리딜트랜스퍼라제 및 UDP N-아세틸글루코사민 2-에피머라제의 효소 활성을 포함할 필요는 없다 (도 1). 따라서, 추가 및/또는 대안적 실시양태에서, 시알산을 합성 가능한 유전자 조작된 미생물 세포는 UDP의 수반되는 방출과 함께 포스포글루코사민 뮤타제, N-아세틸글루코사민-1-포스페이트 우리딜트랜스퍼라제 및 UDP N-아세틸글루코사민 2-에피머라제로 이루어진 군으로부터 선택된 하나 이상의 효소 활성을 포함하지 않는다.
효소 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제 (EC 2.6.1.16)는 글루타민을 사용하여 프럭토스-6-포스페이트 (Frc-6P)의 글루코사민-6-포스페이트 (GlcN-6P)로의 전환을 촉매한다. 이 효소 반응은 전형적으로 헥소사민 생합성 경로의 제1 단계로 간주된다. 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제의 대안적 명칭은 D-프럭토스-6-포스페이트 아미노-트랜스퍼라제, GFAT, 글루코사민-6-포스페이트 신타제, 헥소스포스페이트 아미노트랜스퍼라제, 및 L-글루타민-D-프럭토스-6-포스페이트 아미노트랜스퍼라제이다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제, 바람직하게는 이종유래 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제, 보다 바람직하게는 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제 (이는 이. 콜라이(E. coli) (이. 콜라이 GlmS (UniProtKB - P17169; 서열번호: 67)로부터 유래된다), 또는 이. 콜라이 GlmS의 기능적 변이체를 보유한다. 가장 바람직하게는, 상기 기능성 변이제는 야생형 효소가 하는 바와 같이 글루코사민-6-포스페이트 억제에 대해 상당히 감소된 민감도를 나타내는 이. 콜라이 GlmS의 버전이다. 글루코사민-6-포스페이트 억제에 대해 상당히 감소된 민감도를 나타내는 이. 콜라이 GlmS의 기능적 변이체의 예는 서열번호: 68)으로 표시된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제, 바람직하게는 이. 콜라이 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제 GlmS (서열번호: 69)를 코딩하는 뉴클레오티드 서열을 포함하는 핵산 분자를 함유하거나, 기능적 변이체를 코딩하는 뉴클레오티드 서열은 야생형 효소와 비교하여 글루코사민-6-포스페이트 억제에 대해 상당히 감소된 민감도를 나타내는 이. 콜라이 GlmS의 버전이다 (glmS*54 또는 glmS* (서열번호: 70으로 표시된 바와 같음)).
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
i) 서열번호: 67 및 서열번호: 68 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii) 서열번호: 69 및 서열번호: 70 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 67 및 서열번호: 68 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 69 및 서열번호: 70 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 세포내 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제 활성을 보유한다. 상기 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제 활성은 GlcN-6P를 N-아세틸글루코사민-6-포스페이트 (GlcNAc-6P)로 전환시킨다. 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제의 예는 사카로마이세스 세레비지애(Saccharomyces cerevisiae) Gna1 (UniProtKB - P43577; 서열번호: 77)이다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제, 바람직하게는 이종유래 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제, 보다 바람직하게는 에스. 세레비지애(S. cerevisiae) Gna1 (서열번호: 78로 표시된 바와 같은 뉴클레오티드 서열에 의해 코딩됨) 또는 그의 기능적 변이체를 함유한다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는
i) 서열번호: 77로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii) 서열번호: 78로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 77로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열과 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 78로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 세포내 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민-6-포스페이트 포스파타제 활성을 보유한다. 상기 N-아세틸글루코사민-6-포스페이트 포스파타제 활성은 GlcNAc-6P를 N-아세틸글루코사민 (GlcNAc)으로 전환시킨다. N-아세틸글루코사민-6-포스페이트 포스파타제의 예는 GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제이다. 효소의 HAD-유사 효소 슈퍼패밀리는 박테리아 효소 할로산 데히드로게나제의 이름을 따서 명명되었으며 포스파타제를 포함한다. GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 적합한 포스파타제는 프럭토스-1-포스페이트 포스파타제 (YqaB, UniProtKB - P77475; 서열번호: 79) 및 알파-D-글루코스 1-포스페이트 포스파타제 (YihX, UniProtKB - P0A8Y3; 서열번호: 80)로 이루어진 군으로부터 선택될 수 있다. 이. 콜라이 YqaB 및 이. 콜라이 YihX 효소가 또한 GlcNAc6P에 대해 작용하는 것으로 여겨진다 (Lee, S.-W. and Oh, M.-K. (2015) Metabolic Engineering 28: 143-150).
추가 및/또는 대안적 실시양태에서, GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제는 유전자 조작된 미생물 세포에서의 이종유래 효소이다. 추가 및/또는 대안적 실시양태에서, GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제는 이. 콜라이 YqaB, 이. 콜라이 YihX, 및 그의 기능적 변이체로 이루어진 군으로부터 선택된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제를 코딩하는 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유한다. 추가 및/또는 대안적 실시양태에서, GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제를 코딩하는 뉴클레오티드 서열은 이종유래 뉴클레오티드 서열이다. 추가 및/또는 대안적 실시양태에서, GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제를 코딩하는 뉴클레오티드 서열은 이. 콜라이 프럭토스-1-포스페이트 포스파타제 또는 이. 콜라이 알파-D-글루코스 1-포스페이트 포스파타제 또는 이들 두 효소 중 하나의 기능적 단편을 코딩한다.
이. 콜라이 YqaB는 서열번호: 81로 표시된 바와 같은 뉴클레오티드 서열에 의해 코딩되며, 한편 이. 콜라이 YihX는 서열번호: 82로 표시된 바와 같은 뉴클레오티드 서열에 의해 코딩된다. 따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
i) 서열번호: 79 및 서열번호: 80 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii) 서열번호: 81 및 서열번호: 82 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 79 및 서열번호: 80 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 81 및 서열번호: 82 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 GlcNAc6P의 GlcNAc로의 전환을 촉매하는 세포내 당 포스파타제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, 자연적으로 발생하지 않는 미생물은 GlcNAc6P의 GlcNAc로의 전환을 촉매하는 HAD-유사 슈퍼패밀리의 당 포스파타제 또는 상기 HAD 포스파타제의 기능적 단편을 코딩하는 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하고/하거나 HAD-유사의 당 포스파타제를 포함하도록 유전자 조작되었다
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민 2-에피머라제 활성을 보유한다. N-아세틸글루코사민 2-에피머라제 (EC 5.1.3.8)는 N-아세틸글루코사민 (GlcNAc)의 N-아세틸만노사민 (ManNAc)으로의 전환을 촉매하는 효소이다. 효소는 탄수화물 및 그의 유도체에 작용하는 라세마제이다. 이 효소 클래스의 계통명은 N-아실-D-글루코사민 2-에피머라제이다. 이 효소는 아미노-당 대사 및 뉴클레오티드-당 대사, 바람직하게는 이종유래 N-아세틸글루코사민 2-에피머라제에 참여한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민 2-에피머라제, 바람직하게는 이종유래 N-아세틸글루코사민 2-에피머라제를 포함한다. N-아세틸글루코사민 2-에피머라제의 예는 아나베나 배리어빌리스(Anabena variabilis), 아카리오클로리스(Acaryochloris) 종, 노스톡(Nostoc) 종, 노스톡 펑크티포르메(Nostoc punctiforme), 박테로이데스 오바투스(Bacteroides ovatus) 또는 시네초시스티스(Synechocystis) 종으로부터 기재되었다. 적합한 N-아세틸글루코사민 2-에피머라제의 예는 유전자 BACOVAβ01816 (서열번호: 85)에 의해 코딩된 바와 같은 비. 오바투스(B. ovatus) ATCC 8483 (UniProtKB - A7LVG6, 서열번호: 83)의 N-아세틸글루코사민 2-에피머라제이다. 또 다른 예는 시네초시스티스 종 (균주 PCC 6803) (UniProtKB - P74124; 서열번호: 84)의 N-아세틸글루코사민 2-에피머라제이며 이는 또한 레닌-결합 단백질로도 공지되어 있으며 slr1975 유전자 (서열번호: 86)에 의해 코딩된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민 2-에피머라제, 바람직하게는 비. 오바투스 ATCC 8483의 N-아세틸글루코사민 2-에피머라제 또는 시네초시스티스 종 (균주 PCC 6803) 또는 그의 기능적 변이체를 코딩하는 뉴클레오티드 서열을 포함하는 핵산 분자를 함유한다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
i) 서열번호: 83 및 서열번호: 84 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii) 서열번호: 85 및 서열번호: 86 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 83 및 서열번호: 84 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 85 및 서열번호: 86 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 세포내 N-아세틸글루코사민 2-에피머라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민-6-포스페이트 에피머라제 활성 및 N-아세틸만노사민-6-포스페이트 포스파타제 활성을 보유한다. N-아세틸글루코사민-6-포스파타제 에피머라제는 N-아세틸글루코사민-6-포스페이트 (GlcNAc-6P)를 N-아세틸만노사민-6-포스페이트 (ManNAc-6P)로 전환시키며, 한편 N-아세틸만노사민-6-포스페이트 포스파타제는 ManNAc-6P를 탈인산화시켜 N-아세틸만노사민 (ManNAc)을 제공한다. N-아세틸글루코사민-6-포스페이트 에피머라제 활성 및 N-아세틸만노사민-6-포스페이트 포스파타제 활성을 보유하는 것은 Neu5Ac 생산을 위해 ManNAc을 제공하는 추가 또는 대안적 방법을 제공한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민-6-포스페이트 에피머라제를 함유한다. 적합한 N-아세틸글루코사민-6-포스페이트 에피머라제의 예는 이. 콜라이 nanE 유전자 (서열번호: 88)에 의해 코딩된 바와 같은 이. 콜라이 NanE (UniprotKB P0A761, 서열번호: 87)이다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민-6-포스페이트 에피머라제를 코딩하는 뉴클레오티드 서열, 바람직하게는 이. 콜라이 NanE을 코딩하는 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유한다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
i) 서열번호: 87로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii) 서열번호: 88로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 87로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열과 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 88로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 세포내 N-아세틸글루코사민-6-포스페이트 에피머라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸만노사민-6-포스페이트 포스파타제를 함유한다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸만노사민-6-포스페이트 포스파타제를 코딩하는 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 시알산 신타제 활성을 포함한다. 시알산 신타제는 ManNAc와 포스포에놀피루베이트 (PEP)의 N-아세틸뉴라민산 (NeuNAc)으로의 축합을 촉매한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 시알산 신타제 또는 그의 기능적 변이체, 바람직하게는 이종유래 시알산 신타제를 포함한다. 시알산 신타제의 예는 여러 가지의 박테리아 종 예컨대 캄필로박터 제주니(Campylobacter jejuni), 스트렙토코커스 아갈락티애(Streptococcus agalactiae), 부티리비브리오 프로테오클라스티쿠스(Butyrivibrio proteoclasticus), 메타노브레비박터 루미나티움(Methanobrevibacter ruminatium), 아세토박테리움 우디이(Acetobacterium woodii), 데술포바쿨라 톨루올리카(Desulfobacula toluolica), 에스케리치아 콜라이, 프레보텔라 니게센스(Prevotella nigescens), 할로르햅두스 티아마테아(Halorhabdus tiamatea), 데술포티그넘 포스피톡시단즈(Desulfotignum phosphitoxidans), 또는 캔디다투스 스칼린두아(Candidatus Scalindua) 종, 이도마리나 로이히엔시스(Idomarina loihiensis), 푸소박테리움 뉴클레아툼(Fusobacterium nucleatum) 또는 나이세리아 메닝기티디스(Neisseria meningitidis)로부터 공지되어 있다. 바람직하게는, 시알산 신타제는 씨. 제주니 neuB 유전자 (서열번호: 90)에 의해 코딩된 바와 같은 씨. 제주니(C. jejuni) (서열번호: 89)의 N-아세틸뉴라민산 신타제 NeuB이다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
i) 서열번호: 89로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드;
ii) 서열번호: 90으로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 89로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열과 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 90으로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 세포내 N-아세틸뉴라민산 신타제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
유전자 조작된 미생물 세포는 시티딘 5'-모노포스페이트를 N-아세틸뉴라민산에 전달하여 CMP-활성화된 N-아세틸뉴라민산 (CMP-NeuNAc)을 생성하기 위한 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제 활성을 보유한다. 몇몇 5'-모노포스포-(CMP)-시알산 신테타제, 예를 들어 이. 콜라이, 나이세리아 메닝기티디스, 캄필로박터 제주니, 스트렙토코커스 종 등으로부터의 5'-모노포스포-(CMP)-시알산 신테타제가 관련 기술분야에 공지되어 있고 기재되어 있다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제, 바람직하게는 이종유래 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제, 보다 바람직하게는 이. 콜라이로부터의 N-아세틸뉴라미네이트 시티딜트랜스퍼라제를 함유한다. 이. 콜라이 NeuA (UnitProtKB - P13266; 서열번호: 91)는 이. 콜라이 neuA 유전자 (서열번호: 92)에 의해 코딩된다.
따라서, 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
i) 서열번호: 91로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii) 서열번호: 92로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 91로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열과 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv) 서열번호: 92로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하며;
여기서 상기 뉴클레오티드 서열은 N-아세틸뉴라미네이트 시티딜트랜스퍼라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
유전자 조작된 미생물 세포는 시알릴트랜스퍼라제 활성, 바람직하게는 이종유래 시알릴트랜스퍼라제 활성, 보다 바람직하게는 α-2,3-시알릴트랜스퍼라제 활성, α-2,6-시알릴트랜스퍼라제 활성 및/또는 α-2,8-시알릴트랜스퍼라제 활성으로 이루어진 군으로부터 선택된 시알릴트랜스퍼라제 활성을 보유한다. 시알릴트랜스퍼라제 활성은 CMP-NeuNAc로부터의 N-아세틸뉴라민산 모이어티를 수용체 분자에 전달 가능하여 (여기서 상기 수용체 분자는 사카라이드 분자이다), 시알릴화 사카라이드를 제공한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 적어도 1종의 시알릴트랜스퍼라제, 바람직하게는 적어도 1종의 이종유래 시알릴트랜스퍼라제를 함유하며, 여기서 상기 시알릴트랜스퍼라제는 공여자 기질로서 CMP-NeuNAc로부터의 NeuNAc 모이어티를 수용체 사카라이드에 전달하기 위한 α-2,3-시알릴트랜스퍼라제 활성 및/또는 α-2,6-시알릴트랜스퍼라제 활성 및/또는 α-2,8-시알릴트랜스퍼라제 활성을 보유한다.
본원에 사용된 바와 같은 용어 "시알릴트랜스퍼라제(sialyltransferase)"는 시알릴트랜스퍼라제 활성을 보유 가능한 폴리펩티드를 지칭한다. "시알릴트랜스퍼라제 활성"은 공여자 기질로부터 수용체 분자로의 시알산 잔기, 바람직하게는 N-아세틸뉴라민산 (Neu5Ac) 잔기의 전달을 지칭한다. 용어 "시알릴트랜스퍼라제"는 본원에 기재된 시알릴트랜스퍼라제의 기능성 단편, 본원에 기재된 시알릴트랜스퍼라제의 기능적 변이체, 및 기능적 변이체의 기능적 단편을 포함한다. 이와 관련하여 "기능적"은 단편 및/또는 변이체가 시알릴트랜스퍼라제 활성을 보유 가능함을 의미한다. 시알릴트랜스퍼라제의 기능적 단편은 그것 자연 발생 유전자에 의해 코딩된 시알릴트랜스퍼라제의 말단절단된 버전을 포함하며, 이 말단절단된 버전은 시알릴트랜스퍼라제 활성을 보유 가능하다. 말단절단된 버전의 예는 전형적으로 폴리펩티드를 특이적 세포하 국소화로 지향하는 소위 리더 서열을 포함하지 않는 시알릴트랜스퍼라제이다. 전형적으로, 이러한 리더 서열은 그의 세포하 수송 동안 폴리펩티드로부터 제거되고, 자연 발생 성숙 시알릴트랜스퍼라제에 또한 부재한다.
이종유래 시알릴트랜스퍼라제는 시알산 잔기를 공여자 기질로부터 수용체 분자로 전달 가능하다. 이종유래 시알릴트랜스퍼라제와 관련하여 용어 "가능하다"는 이종유래 시알릴트랜스퍼라제의 시알릴트랜스퍼라제 활성 및 이종유래 시알릴트랜스퍼라제가 그의 효소 활성을 보유하기 위해 적합한 반응 조건이 요구된다는 조항을 지칭한다. 적절한 반응 조건의 부재 하에, 이종유래 시알릴트랜스퍼라제는 그의 효소 활성을 보유하지 않으나, 적합한 반응 조건이 회복되는 경우 그의 효소 활성을 유지하고 그의 효소 활성을 보유한다. 적합한 반응 조건은 적합한 공여자 기질의 존재, 적합한 수용체 분자의 존재, 필수 보조인자 예컨대 - 예를 들어 1가 또는 2가 이온의 존재, 적절한 범위의 pH 값, 적합한 온도 등을 포함한다. 이종유래 시알릴트랜스퍼라제의 효소 반응을 수행하는 각각의 그리고 모든 인자에 대한 최적 값이 충족될 필요는 없으나, 반응 조건은 이종유래 시알릴트랜스퍼라제가 그의 효소 활성을 수행하도록 되어야 한다. 따라서, 용어 "가능하다"는 이종유래 시알릴트랜스퍼라제의 효소 활성이 비가역적으로 손상되는 임의의 조건을 배제하고 또한 이러한 임의의 조건에 대한 이종유래 시알릴트랜스퍼라제의 노출을 배제하였다. 대신에, "가능하다"는 허용 반응 조건 (시알릴트랜스퍼라제가 그의 효소 활성을 수행하는 데 필요한 모든 요건)이 시알릴트랜스퍼라제에 제공되는 경우, 시알릴트랜스퍼라제가 효소적으로 활성이라는 것, 즉 그의 시알릴트랜스퍼라제 활성을 보유한다는 것을 의미한다.
시알릴트랜스퍼라제는 그들이 형성하는 당 연결의 유형에 따라 구별될 수 있다. 본원에 사용된 바와 같이 용어 "α-2,3-시알릴트랜스퍼라제" 및 "α-2,3-시알릴트랜스퍼라제 활성"은 α-2,3 연결을 가진 시알산 잔기를 갈락토스, N-아세틸갈락토사민 또는 수용체 분자의 갈락토스 또는 N-아세틸갈락토사민 잔기에 부가하는 폴리펩티드 및 그의 효소 활성을 지칭한다. 마찬가지로, 용어 "α-2,6-시알릴트랜스퍼라제" 및 "α-2,6-시알릴트랜스퍼라제 활성"은 α-2,6 연결을 가진 시알산 잔기를 갈락토스, N-아세틸갈락토사민 또는 수용체 분자의 갈락토스 또는 N-아세틸갈락토사민 잔기에 부가하는 폴리펩티드 및 그의 효소 활성을 지칭한다. 마찬가지로, 용어 "α-2,8-시알릴트랜스퍼라제" 및 "α-2,8-시알릴트랜스퍼라제 활성"은 α-2,8 연결을 가진 시알산 잔기를 갈락토스, N-아세틸갈락토사민 또는 수용체 분자의 갈락토스 또는 N-아세틸갈락토사민 잔기에 부가하는 폴리펩티드 및 그의 효소 활성을 지칭한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는,
I. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드;
II. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 아미노산 서열 중 어느 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드; 및
III. I. 및 II.의 폴리펩티드 중 어느 하나의 단편
으로 이루어진 군으로부터 바람직하게 선택된 이종유래 시알릴트랜스퍼라제를 함유한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 이종유래 시알릴트랜스퍼라제를 코딩하는 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하도록 형질전환되었다. 바람직하게는, 뉴클레오티드 서열은 표 1로부터 추론될 수 있는 바와 같다. 추가 및/또는 대안적 실시양태에서, 뉴클레오티드 서열은,
i. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii. 서열번호: 34 내지 66 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv. 서열번호: 34 내지 66으로 표시된 뉴클레오티드 서열 중 어느 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v. i., ii., iii. 및 iv의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi. i., ii., iii., iv. 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택되며;
여기서 상기 뉴클레오티드 서열은 시알릴트랜스퍼라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
표 1: 시알릴트랜스퍼라제-코딩 뉴클레오티드 서열의 목록. 시알릴트랜스퍼라제-코딩 뉴클레오티드 서열은 그의 야생형 단백질 코딩 영역과 비교하여 전장 구축물 (FL)로서 또는 예측된 신호 펩티드 (Δ) 없이 클로닝되었다. Δ 뒤에 있는 숫자는 상응하는 서열로부터 결실된 N-말단 아미노산을 나타낸다.
표현 "서열번호: 1 내지 33 중 어느 하나"는 서열번호: 1, 서열번호: 2, 서열번호: 3, 서열번호: 4, 서열번호: 5, 서열번호: 6, 서열번호: 7, 서열번호: 8, 서열번호: 9, 서열번호: 10, 서열번호: 11, 서열번호: 12, 서열번호: 13. 서열번호: 14. 서열번호: 15, 서열번호: 16, 서열번호: 17, 서열번호: 18, 서열번호: 19, 서열번호: 20, 서열번호: 21, 서열번호: 22, 서열번호: 23, 서열번호: 24, 서열번호: 25, 서열번호: 26, 서열번호: 27, 서열번호: 28, 서열번호: 29, 서열번호: 30, 서열번호: 31, 서열번호: 32, 및 서열번호: 33으로 이루어진 군 중 어느 하나를 지칭한다. 동일한 원칙이 표현 "서열번호: 34 내지 66 중 어느 하나"에 적용된다. 일반적으로 말해서, 표현 "서열번호: X 내지 Z 중 어느 하나" (여기서 "X" 및 "Z"는 자연수를 나타낸다)는, X에서 Z까지의 식별 번호를 포함하는 "서열번호" 중 어느 하나로 표시된 모든 서열 (뉴클레오티드 서열 또는 아미노산 서열)을 지칭한다.
게다가, 유전자 조작된 미생물 세포는 이종유래 시알릴트랜스퍼라제를 코딩하는 뉴클레오티드 서열을 발현하도록 유전자 조작되었다. 이를 위해, 이종유래 시알릴트랜스퍼라제를 코딩하는 뉴클레오티드 서열은 유전자 조작된 세포에서 이종유래 시알릴트랜스퍼라제를 코딩하는 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 발현 제어에 작동 가능하게 연결된다.
본원에 사용된 바와 같은 용어 "작동 가능하게 연결된"은, 이종유래 시알릴트랜스퍼라제를 코딩하는 뉴클레오티드 서열과 제2 뉴클레오티드 서열, 핵산 발현 제어 서열 (예컨대 프로모터, 오퍼레이터, 인핸서, 조절제, 전사 인자 결합 부위의 어레이, 전사 종결자, 리보솜 결합 부위) 사이의 기능적 연결을 지칭하며, 여기서 발현 제어 서열은 이종유래 시알릴트랜스퍼라제를 코딩하는 뉴클레오티드 서열에 상응하는 핵산의 전사 및/또는 번역에 영향을 미친다. 따라서, 용어 "프로모터"는 대개 DNA 중합체에서 유전자에 "선행하고" mRNA로의 전사 개시 부위를 제공하는 DNA 서열을 지정한다. "조절자" DNA 서열, 또한 대개 주어진 DNA 중합체에서 유전자의 "상류" (즉, 선행)는, 전사 개시의 빈도 (또는 속도)를 결정하는 단백질에 결합한다. "프로모터/조절자" 또는 "제어" DNA 서열로 총칭되어, 기능적 DNA 중합체에서 선택된 유전자 (또는 일련의 유전자)를 선행하는 이들 서열이 협력하여 유전자의 전사 (및 최종적인 발현)가 발생할 지 여부를 결정한다. DNA 중합체에서 유전자를 "따르고" mRNA로의 전사 종결 신호를 제공하는 DNA 서열은 전사 "종결자" 서열로 지칭된다.
추가 및/또는 대안적 실시양태에서, α-2,3-시알릴트랜스퍼라제 활성을 보유 가능한 이종유래 시알릴트랜스퍼라제는,
I 서열번호: 1 내지 27 중 어느 하나로 표시된 바와 같은 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드;
II. 서열번호: 1 내지 27 중 어느 하나로 표시된 바와 같은 아미노산 서열 중 어느 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드; 및
III. I. 및 II.의 폴리펩티드 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 α-2,3-시알릴트랜스퍼라제 활성을 보유 가능한 상기 이종유래 시알릴트랜스퍼라제를 코딩하는 적어도 하나의 뉴클레오티드 서열을 포함하는 핵산 분자를 함유하며, 여기서 상기 적어도 하나의 뉴클레오티드 서열은,
i. 서열번호: 1 내지 27 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii. 서열번호: 34 내지 60 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii.
서열번호: 1 내지 27 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv. 서열번호: 34 내지 60으로 표시되는 뉴클레오티드 서열 중 어느 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v. i., ii., iii. 및 iv의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi. i., ii., iii., iv. 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택되며;
여기서 상기 뉴클레오티드 서열은 α-2,3-시알릴트랜스퍼라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, α-2,3-시알릴트랜스퍼라제 활성을 보유 가능한 이종유래 시알릴트랜스퍼라제는 LC-MS/MS를 사용한 LNT 시알릴화의 정량적 분석에 의한 서열번호: 27로 표시된 바와 같은 시알릴트랜스퍼라제의 상대적 효능과 비교하여, 적어도 100배, 적어도 200배, 적어도 300배, 적어도 1000배, 적어도 10,000배의 상대적 효능을 갖는다.
또 다른 실시양태에서, 이종유래 시알릴트랜스퍼라제는 α-2,6-시알릴트랜스퍼라제 활성을 보유 가능하다.
추가 실시양태에서, α-2,6-시알릴트랜스퍼라제 활성을 보유 가능한 이종유래 시알릴트랜스퍼라제는,
I. 서열번호: 28 내지 33 중 어느 하나로 표시된 바와 같은 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드;
II. 서열번호: 28 내지 33 중 어느 하나로 표시된 바와 같은 아미노산 서열 중 어느 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 동일성을 갖는 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드; 및
III. I. 및 II.의 폴리펩티드 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 α-2,6-시알릴트랜스퍼라제 활성을 보유 가능한 상기 이종유래 시알릴트랜스퍼라제를 코딩하는 적어도 하나의 뉴클레오티드 서열을 포함하는 핵산 분자를 함유하며, 여기서 상기 적어도 하나의 뉴클레오티드 서열은,
i. 서열번호: 28 내지 33 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii. 서열번호: 61 내지 66 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii. 서열번호: 28 내지 33 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
iv. 서열번호: 61 내지 66으로 표시된 뉴클레오티드 서열 중 어느 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 동일성을 갖는 뉴클레오티드 서열;
v. i., ii., iii. 및 iv의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi. i., ii., iii., iv. 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
로 이루어진 군으로부터 선택되며;
여기서 상기 뉴클레오티드 서열은 α-2,6-시알릴트랜스퍼라제 활성을 제공하기 위해 유전자 조작된 미생물 세포에서 상기 뉴클레오티드 서열의 전사 및/또는 번역을 수행하는 적어도 하나의 핵산 발현 제어 서열에 작동 가능하게 연결된다.
추가 및/또는 대안적 실시양태에서, α-2,6-시알릴트랜스퍼라제 활성을 보유 가능한 이종유래 시알릴트랜스퍼라제는 LNT 시알릴화의 정량적 분석에 의한 서열번호: 33으로 표시된 바와 같은 시알릴트랜스퍼라제의 상대적 효능과 비교하여, 적어도 100배, 적어도 200배, 적어도 300배의 상대적 효능을 갖는다.
추가 및/또는 대안적 실시양태에서, 이종유래 시알릴트랜스퍼라제는 α-2,8-시알릴트랜스퍼라제 활성을 보유 가능하다. α-2,8-시알릴트랜스퍼라제 활성을 보유 가능한 이종유래 시알릴트랜스퍼라제의 예는 캄필로박터 제주니 OH4384의 시알릴트랜스퍼라제 CstII이다.
시알릴트랜스퍼라제는 시알산 잔기, 예를 들어 N-아세틸뉴라민산 (Neu5Ac) 잔기를, 공여자 기질, 예를 들어 CMP-Neu5Ac로부터, 수용체 분자로 전달 가능하다. 수용체 분자는 사카라이드 분자, 바람직하게는 표 2에 제시된 사카라이드 분자이다.
표 2: 시알릴화 사카라이드 생산을 위한 수용체 기질(acceptor substrate)로서 사용될 수 있는 사카라이드의 목록. 시알릴화 사카라이드 자체가 추가 시알릴화 사카라이드의 생산을 위한 수용체 기질로서 또한 사용될 수 있다.
추가 및/또는 대안적 실시양태에서, 수용체 분자는 모노사카라이드, 바람직하게는 N-아세틸글루코사민, 갈락토스 및 N-아세틸갈락토사민으로 이루어진 군으로부터 선택된 모노사카라이드이다.
추가 및/또는 대안적 실시양태에서, 수용체 분자는 디사카라이드, 바람직하게는 락토스, 락툴로스, N-아세틸락토사민, 락토-N-비오스, 락툴로스 및 멜리비오스로 이루어진 군으로부터 선택된 디사카라이드이다.
추가 및/또는 대안적 실시양태에서, 수용체 분자는 트리사카라이드, 바람직하게는 라피노스, 락토-N-트리오스 II, 2'-푸코실락토스, 3-푸코실락토스, 3'-시알릴락토스, 6'-시알릴락토스, 3'-시알릴-N-아세틸락토사민, 6'-시알릴-N-아세틸락토사민, 3'-갈락토실락토스 및 6'-갈락토실락토스로 이루어진 군으로부터 선택된 트리사카라이드이다.
추가 및/또는 대안적 실시양태에서, 수용체 분자는 테트라사카라이드, 바람직하게는 락토-N-테트라오스, 락토-N-네오테트라오스, 2'3-디푸코실락토스, 3-푸코실-3'-시알릴락토스 및 3-푸코실-6'-시알릴락토스로 이루어진 군으로부터 선택된 테트라사카라이드이다.
추가 및/또는 대안적 실시양태에서, 수용체 분자는 펜타사카라이드, 바람직하게는 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c, 락토-N-푸코펜타오스 I, 락토-N-푸코펜타오스 II, 락토-N-푸코펜타오스 III, 락토-N-푸코펜타오스 V, 락토-N-네오푸코펜타오스 I 및 락토-N-네오푸코펜타오스 V로 이루어진 군으로부터 선택된 펜타사카라이드이다.
본원에 사용된 바와 같은 용어 "기능적 변이체"는, 본원에 언급된 바와 같은 효소와 관련하여, 활성 손실 없이 지정된 효소의 폴리펩티드 변이체를 지칭하며, 이는 지정된 효소의 아미노산 서열과 적어도 70%, 바람직하게는 적어도 80%, 적어도 85%, 적어도 90%, 적어도 95%, 적어도 98% 또는 최소 99% 동일성을 공유한다. 이는 이들 폴리펩티드가 유래된 게놈 서열 데이터의 일부 가변성의 가능성, 및 또한 이들 폴리펩티드에 존재하는 일부 아미노산이 효소의 촉매 활성에 상당히 영향을 미치지 않고 대체될 수 있는 가능성을 고려한다.
용어 "기능적 변이체"는 또한 촉매 활성의 상당한 손실 없이 효소의 말단절단된 변이체를 나타내는 지정된 효소의 폴리펩티드 변이체를 포함한다. 따라서, 말단절단된 변이체의 아미노산 서열은 1개, 2개 또는 2개 초과의 연속된 아미노산의 스트레치가 부재한다는 점에서 지정된 효소의 아미노산 서열과 상이할 수 있다. 말단절단은 아미노 말단 (N-말단)에서, 카르복실 말단 (C-말단)에서 및/또는 지정된 효소의 아미노산 서열 내에 있을 수 있다.
용어 "작동 가능하게 연결된"은 핵산 발현 제어 서열 (예컨대 프로모터, 신호 서열, 또는 전사 인자 결합 부위의 어레이)과 제2 핵산 서열 사이의 기능적 연결을 지칭하며, 여기서 발현 제어 서열은 제2 서열에 상응하는 핵산의 전사 및/또는 번역에 영향을 미친다.
이미 상기 효소를 코딩하는 하나 이상의 유전자를 보유하고, NeuNAc, CMP-NeuNAc 및/또는 시알릴화 사카라이드를 생산하기에 충분한 방식으로 상기 유전자를 발현하는 미생물 세포는 시알산 생합성을 완료하고 시알산 모이어티를 사카라이드 수용체로 전달하기 위해 유전자 조작될 필요가 없음을 이해하여야 하나, 그럼에도 불구하고 유전자 조작되어 상기 유전자 중 하나 이상의 발현 수준을 변경하여 상기 하나 이상의 유전자 생성물의 세포내 수준 예컨대 - 예를 들어 글루타민:프럭토스-6-포스페이트 아미노트랜스퍼라제, 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제, N-아세틸글루코사민-6-포스페이트 포스파타제, N-아세틸글루코사민 2-에피머라제 및/또는 N-아세틸뉴라민산 신타제의 양을 증가시켜, 따라서 유전자 조작된 세포에서, Neu5Ac 생합성의 속도를 증가시키고, 결과적으로 시알릴화 사카라이드의 속도를 증가시킬 수 있음을 이해하여야 한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 세포의 야생형보다 더 많은 PEP를 합성한다. 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 향상된 PEP 생합성 경로를 갖도록 유전적으로 조작되었다. 바람직하게는, 유전자 조작된 미생물 세포는 증가된 포스포에놀피루베이트 신타제 활성을 보유하도록 유전자 조작되었는데, 예를 들어 포스포에놀피루베이트 신타제 유전자를 코딩하는 ppsA 유전자가 과발현된다는 점에서 및/또는 자연적으로 발생하지 않는 미생물이 포스포에놀피루베이트 신타제 또는 그의 기능적 변이체의 발현을 허용하는 뉴클레오티드 서열의 적어도 하나의 추가 카피를 함유한다는 점에서이다. ppsA의 과발현은 세포내 PEP 합성을 향상시켜 시알산 생산에 더 많은 PEP를 이용 가능하게 할 수 있게 된다. 예를 들어, 적합한 포스포에놀피루베이트 신타제는 이. 콜라이의 PpsA이다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 이. 콜라이 PpsA 또는 그의 기능적 변이체를 코딩하는 뉴클레오티드 서열를 포함하는 핵산 분자를 함유한다. 이. 콜라이 PpsA 또는 그의 기능적 변이체를 코딩하는 상기 뉴클레오티드 서열은 이. 콜라이 ppsA 유전자와 적어도 80%, 적어도 85%, 적어도 90%, 적어도 95%, 적어도 98% 또는 적어도 99%의 서열 동일성을 갖는다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 수크로스 퍼미아제, 수크로스 히드롤라제, 프럭토키나제, L-글루타민:D-프럭토스-6-포스페이트 아미노트랜스퍼라제, 글루코사민-6-포스페이트-N-아세틸트랜스퍼라제, N-아세틸글루코사민-2-에피머라제, 시알산 신타제, 포스포에놀피루베이트 신타제로 이루어진 군으로부터 선택된 효소 활성을 보유 가능한 폴리펩티드를 코딩하는 하나 이상의 유전자를 추가로 포함하며, 여기서 바람직하게는 이들 유전자 중 적어도 하나, 바람직하게는 모두는 야생형 미생물 세포와 비교하여 유전자 조작된 미생물 세포에서 과발현된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포의 전구 세포주에서 자연적으로 발생하는 시알산 이화 경로는 유전적으로 조작된 미생물 세포에서는 불능상태로 되었다.
상기 방법 및 유전자 조작된 미생물 세포의 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 유전자 조작된 미생물 세포의 전구 세포와 비교하여, α-N-아세틸갈락토사미니다제 (예를 들어 NagA), N-아세틸글루코사민키나제 (예를 들어 NagK), N-아세틸뉴라미네이트 리아제 (= N-아세틸뉴라민산 알돌라제, 예를 들어 NanA), β-갈락토시다제, 글루코사민-6-포스페이트 데아미나제, N-아세틸글루코사민-6-포스페이트 데아세틸라제, N-아세틸만노사민 키나제 및/또는 N-아세틸만노사민-6-포스페이트 에피머라제로 이루어진 군으로부터 선택된 하나 이상의 효소 활성이 결핍되거나 감소된 활성을 보유한다.
상기 방법 및 유전자 조작된 미생물 세포의 추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸글루코사민-1-포스페이트 우리딜트랜스퍼라제, 글루코사민-1-포스페이트 아세틸 트랜스퍼라제, 포스포글루코사민 뮤타제, UDP-N-아세틸글루코사민-2-에피머라제, UDP-갈락토스-4-에피머라제, 갈락토스-1-포스페이트 우리딜릴트랜스퍼라제, 포스포글루코뮤타제, 글루코스-1-포스페이트 우리딜릴트랜스퍼라제, 포스포만노뮤타제, 만노스-1-포스페이트 구아노실트랜스퍼라제, GDP-만노스-4,6-데히드라타제, GDP-L-푸코스 신타제 및 푸코스키나제/L-푸코스-1-포스페이트-구아닐트랜스퍼라제로 이루어진 군으로부터 선택된 효소 활성을 보유 가능한 폴리펩티드를 코딩하는 하나 이상의 유전자를 추가로 포함한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 기능적 락토스 퍼미아제, 기능적 시알산 수송체 (외수송체(exporter))로 이루어진 군으로부터 선택된 적어도 하나를 포함하며, 여기서 바람직하게는 기능적 락토스 퍼미아제, 기능적 수크로스 퍼미아제, 기능적 시알산 수송체 (외수송체)로 이루어진 군으로부터 선택된 하나를 코딩하는 적어도 하나의 뉴클레오티드 서열을 포함하고 발현하며, 여기서 바람직하게는 이들 뉴클레오티드 서열 중 적어도 하나는 세포에서 과발현된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 PEP를 소비하지 않는 메커니즘을 통해 상기 유일한 탄소 공급원을 세포로 전달 가능하도록 추가로 변형된다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 기능적 수크로스 이용 시스템을 보유한다. 상기 기능성 수크로스 이용 시스템은 외인성으로 공급된 수크로스 및 그 가수분해의 세포 내수송(import)을 가능하게 하여 생성된 모노사카라이드 글루코스 및 프럭토스가 유전자 조작된 세포의 대사 및 원하는 시알릴화 올리고사카라이드 생산에 의해 대사적으로 이용될 수 있도록 한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 유전자 변형되어 기능적 수크로스 이용 시스템을 보유한다. 추가 및/또는 대안적 실시양태에서 자연적으로 발생하지 않는 미생물의 수크로스 이용 시스템은 수크로스 양성자 공수송(proton symport) 수송 시스템, 프럭토키나제, 인버타제 및 수크로스 오페론 리프레서(repressor)를 포함한다.
적합한 수크로스 양성자 공수송 수송 시스템은 cscB 유전자에 의해 코딩된 CscB, 예를 들어 이. 콜라이의 cscB 유전자에 의해 코딩된 바와 같은 이. 콜라이의 CscB (UniProtKB - P30000)이다.
적합한 프럭토키나제 (EC 2.7.1.4)는 cscK 유전자에 의해 코딩된 CscK, 예를 들어 이. 콜라이의 cscK 유전자에 의해 코딩된 바와 같은 이. 콜라이의 CscK (UniProtKB - P40713)이다.
β-D-프럭토푸라노시드에서 말단 비-환원 β-D-프럭토푸라노시드 잔기를 가수분해하는 적합한 인버타제 (EC 3.2.1.26)는 CscA, 예를 들어 이. 콜라이의 cscA 유전자에 의해 코딩된 바와 같은 이. 콜라이의 CscA (UniProtKB - O86076)이다.
적합한 수크로스 오페론 리프레서는 cscR 유전자에 의해 코딩된 바와 같은 CscR, 예를 들어 이. 콜라이의 cscR 유전자에 의해 코딩된 바와 같은 이. 콜라이의 CscR (UniProtKB - P62604)이다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 세포는 수크로스 양성자 공수송 수송 시스템, 프럭토키나제, 인버타제 및 수크로스 오페론 리프레서 또는 이들 단백질 중 어느 하나의 기능적 변이체를 보유하도록 유전자 조작되었다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 세포는 상기 수크로스 양성자 공수송 수송 시스템, 프럭토키나제, 인버타제 및 수크로스 오페론 리프레서의 발현을 위한 수크로스 양성자 공수송 수송 시스템, 프럭토키나제, 인버타제 및 수크로스 오페론 리프레서를 코딩하는 뉴클레오티드 서열을 포함하는 핵산 분자를 보유하도록 유전자 조작되었다. 추가 및/또는 대안적 실시양태에서, 유전자 조작된 세포는 유전자 cscB, cscK, cscA, 바람직하게는 이. 콜라이 유전자 cscB, cscK, cscA 및 cscR를 발현하도록 유전자 조작되었다.
추가 및/또는 대안적 실시양태에서, CscB, CscK, CscA 또는 CscR의 기능적 변이체를 코딩하는 뉴클레오티드 서열은 이. 콜라이 cscB, cscK, cscA 또는 cscR 각각과 적어도 80%, 적어도 85%, 적어도 90%, 적어도 95%, 적어도 98% 또는 적어도 99%의 서열 동일성을 갖는다.
추가 및/또는 대안적 실시양태에서, 자연적으로 발생하지 않는 미생물은 β-갈락토시드 퍼미아제 및 β-갈락톡시다제를 발현한다.
추가 및/또는 대안적 실시양태에서, 자연적으로 발생하지 않는 미생물은 β-갈락토시드 퍼미아제, 바람직하게는 이. 콜라이 락토스 퍼미아제 LacY (서열번호: 93) 또는 그의 기능적 변이체 및 β-갈락톡시다제, 바람직하게는 이. 콜라이 LacZ (서열번호: 95) 또는 그의 기능적 변이체를 발현하도록 유전자 조작되었다. 추가 및/또는 대안적 실시양태에서, 자연적으로 발생하지 않는 미생물은 β-갈락토시드 퍼미아제를 코딩하는 뉴클레오티드 서열, 바람직하게는 이. 콜라이 LacY (서열번호: 94) 또는 그의 기능적 변이체를 코딩하는 뉴클레오티드 서열, 및/또는 β-갈락톡시다제를 코딩하는 뉴클레오티드 서열, 바람직하게는 이. 콜라이 LacZ (서열번호: 96) 또는 그의 기능적 변이체를 코딩하는 뉴클레오티드 서열을 포함하는 핵산 분자를 수반하도록 유전자 조작되었다.
추가 및/또는 대안적 실시양태에서, 이. 콜라이 LacY 또는 그의 기능적 변이체를 코딩하는 뉴클레오티드 서열은 이. 콜라이 lacY와 적어도 80%, 적어도 85%, 적어도 90%, 적어도 95%, 적어도 98% 또는 적어도 99%의 서열 동일성을 갖는다.
추가 및/또는 대안적 실시양태에서, 이. 콜라이 LacZ 또는 그의 기능적 변이체를 코딩하는 뉴클레오티드 서열은 이. 콜라이 lacZ와 적어도 80%, 적어도 85%, 적어도 90%, 적어도 95%, 적어도 98% 또는 적어도 99%의 서열 동일성을 갖는다.
CMP-Neu5Ac를 생산할 수 있으며, 기능적 β-갈락토시드 퍼미아제 및 기능적 β-갈락톡시다제를 발현하는 자연적으로 발생하지 않는 미생물은 유일한 탄소 공급원으로서 락토스 상에서 상기 자연적으로 발생하지 않는 미생물의 배양을 가능하게 한다.
시알릴화 사카라이드를 생산할 수 있는 유전자 조작된 미생물 세포는 - 임의로 - 추가 특색을 포함할 수 있으며, 이들 추가 특색을 보유하도록 유전적으로 조작될 수 있다. 이들 추가 특색은 자연적으로 발생하지 않는 미생물의 생산성을 개선시켜 더 높은 시알릴화 사카라이드 수율을 야기하는 것으로 간주된다.
추가 및/또는 대안적 실시양태에서 유전자 조작된 미생물 세포는, 바람직하게는 wcaJ 유전자 또는 그의 기능적 변이체를 결실시킴으로써, wcaJ 유전자 또는 그의 기능적 변이체의 발현을 손상시킴으로써, 또는 변경된 뉴클레오티드 서열에 의해 코딩되는 폴리펩티드가 WcaJ의 효소 활성을 보유하지 않도록 돌연변이를 단백질-코딩 영역에 도입함으로써 WcaJ 효소의 활성을 폐지함으로써, UDP-글루코스:운데카프레닐포스페이트 글루코스-1-포스페이트 트랜스퍼라제 활성을 폐지하도록 유전자 조작되었다. WcaJ는 UDP-글루코스:운데카프레닐포스페이트 글루코스-1-포스페이트 트랜스퍼라제를 코딩한다. 상기 UDP-글루코스:운데카프레닐포스페이트 글루코스-1-포스페이트 트랜스퍼라제는 콜란산 생합성에서의 제1 효소이다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 β-갈락톡시다제 유전자 (lacZ)가 결실되었거나, β-갈락톡시다제 유전자의 발현이 손상되거나, β-갈락톡시다제 유전자의 단백질 코딩 영역의 뉴클레오티드 서열이 수정되어 상기 변경된 뉴클레오티드 서열(들)에 의해 코딩되는 폴리펩티드가 β-갈락토시다제의 효소 활성을 보유하지 않도록 한다는 점에서 유전자 조작되었다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 갈락토스 키나제 (예를 들어 galK 유전자)를 코딩하는 유전자가 결실되었거나, galK 유전자의 발현이 손상되거나, galK 유전자의 단백질 코딩 영역의 뉴클레오티드 서열이 수정되어 상기 변경된 뉴클레오티드 서열(들)에 의해 코딩되는 폴리펩티드가 갈락토스 키나제의 효소 활성을 보유하지 않도록 한다는 점에서 유전자 조작되었다. galK 유전자 / GalK의 결실 또는 불활성화는 유전자 조작된 미생물 세포가 시알화 반응만을 위한 수용체 기질로서 갈락토스를 이용할 수 있다는 점에서 유리하다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 N-아세틸갈락토사미니다제 (nagA)를 코딩하는 유전자가 결실되었거나, 그의 발현이 손상되었거나, 단백질 코딩 영역의 뉴클레오티드 서열이 수정되어 상기 변경된 뉴클레오티드 서열(들)에 의해 코딩되는 폴리펩티드가 N-아세틸갈락토사미니다제의 효소 활성을 보유하지 않도록 한다는 점에서 유전자 조작되었다. nagA / NagA의 결실 또는 불활성화는 유전자 조작된 미생물 세포가 시알화 반응만을 위한 수용체체 기질로서 GlcNAc 또는 GlcNAc-6-포스페이트를 이용할 수 있다는 점에서 유리하다.
추가 및/또는 대안적 실시양태에서 유전자 조작된 미생물 세포는, 바람직하게는 fucI 유전자를 결실시킴으로써, fucI 유전자의 발현을 손상시킴으로써, 또는 상기 변경된 뉴클레오티드 서열에 의해 코딩되는 폴리펩티드가 푸코스 이소머라제 활성을 보유하지 않도록 fucI 유전자의 단백질-코딩 영역을 변형함으로써 푸코스 이소머라제 활성을 폐지하도록 유전자 조작되었다. 예를 들어, 이. 콜라이 L-푸코스 이소머라제 FucI (UniProtKB - P69922)는 이. 콜라이 fucI 유전자에 의해 코딩된다.
푸쿨로키나제는 푸코스의 인산화를 촉매한다. 푸쿨로키나제는 L-푸코스로부터 글리세론 포스페이트와 L-락트알데히드를 합성하는 하위경로에서의 제2 효소이다. 이. 콜라이 푸쿨로키나제 FucK (UniProtKB - P11553)는 이. 콜라이 fucK 유전자에 의해 코딩된다. 이. 콜라이 푸쿨로키나제는 더 낮은 효율로, D-리불로스, D-크실룰로스 및 D-프럭토스를 또한 인산화할 수 있다.
추가 및/또는 대안적 실시양태에서 유전자 조작된 세포는, 바람직하게는 fucK 유전자를 결실시킴으로써 또는, fucK 유전자의 발현을 손상시킴으로써, 또는 상기 변경된 뉴클레오티드 서열에 의해 코딩되는 폴리펩티드가 푸코스 이소머라제 활성을 보유하지 않도록 fucK 유전자의 단백질-코딩 영역에 돌연변이를 도입함으로써 푸코스 이소머라제 활성을 폐지하도록 유전자 조작되었다.
N-아세틸갈락토사민-6-포스페이트 데아세틸라제는 하기 반응을 촉매한다: N-아세틸-D-갈락토사민 6-포스페이트 + H2O → 갈락토사민 6-포스페이트 + 아세테이트. N-아세틸갈락토사민-6-포스페이트 데아세틸라제는 agaA 유전자에 의해 코딩된다. 이. 콜라이에서 N-아세틸갈락토사민-6-포스페이트 데아세틸라제 AgaA (UniProtKB - P42906)는 이. 콜라이 agaA 유전자에 의해 코딩된다.
추가 및/또는 대안적 실시양태에서 유전자 조작된 미생물 세포는, 바람직하게는 agaA 유전자를 결실시킴으로써, agaA 유전자의 발현을 손상시킴으로써, 또는 상기 변경된 뉴클레오티드 서열에 의해 코딩되는 폴리펩티드가 N-아세틸갈락토사민-6-포스페이트 데아세틸라제 활성을 보유하지 않도록 agaA 유전자의 단백질-코딩 영역에 돌연변이를 도입함으로써 N-아세틸갈락토사민-6-포스페이트 데아세틸라제 활성을 폐지하도록 유전자 조작되었다.
추가 및/또는 대안적 실시양태에서, 적어도 하나의 유전자 조작된 미생물 세포는 UDP-N-아세틸글루코사민, UDP-갈락토스 및 GDP-푸코스로 이루어진 군으로부터 선택된 하나 이상의 뉴클레오티드-활성화된 당의 증가된 생산을 보유한다. 바람직하게는, 적어도 하나의 유전자 조작된 미생물 세포는 하나 이상의 상기 뉴클레오티드-활성화된 당의 증가된 생산을 보유하도록 추가로 유전자 조작되었다. 상기 뉴클레오타이드 활성화 당 중 적어도 하나의 생산은, 상기 뉴클레오티드-활성화된 당 중 적어도 하나의 증가된 생산을 보유하도록 추가로 유전자 조작되기 전에 추가로 유전자 조작된 미생물 세포의 전구 세포에서 동일한 뉴클레오티드-활성화된 당(들)의 생산과 비교하여 추가로 유전자 조작된 세포에서 증가된다.
추가 및/또는 대안적 실시양태에서, 적어도 하나의 미생물 세포는 L-글루타민:D-프럭토스-6-포스페이트 아미노트랜스퍼라제, N-아세틸글루코사민-1-포스페이트 우리딜트랜스퍼라제, 글루코사민-1-포스페이트 아세틸 트랜스퍼라제, 포스포글루코사민 뮤타제, UDP-갈락토스-4-에피머라제, 갈락토스-1-포스페이트 우리딜릴-트랜스퍼라제, 포스포글루코뮤타제, 글루코스-1-포스페이트 우리딜릴트랜스퍼라제, 포스포만노-뮤타제, 만노스-1-포스페이트 구아노실트랜스퍼라제, GDP-만노스-4,6-데히드라타제, GDP-L-푸코스 신타제 및 푸코스 키나제/L-푸코스-1-포스페이트-구아닐트랜스퍼라제로 이루어진 군으로부터 선택된 효소 활성을 보유 가능한 폴리펩티드를 코딩하는 하나 이상의 유전자를 과발현하도록 추가로 유전자 조작되었다.
현재, 그리고 일반 분야에서 이해되는 바와 같이, 여기에서 각각 본원에 논의된 모든 폴리뉴클레오티드 또는 핵산과 관련하여, 하나 이상의 유전자 또는 폴리펩티드의 상기 과발현은 상기 하나 이상의 유전자 또는 폴리펩티드의 과발현을 보유하도록 추가로 유전자 조작되지 전에 추가로 유전자 조작된 미생물 세포의 전구 세포와 비교하여 과발현이다.
하나 이상의 상기 유전자의 과발현은 유전자 조작된 미생물 세포에서 상응하는 폴리펩티드, 즉 효소(들)의 양을 증가시키고, 따라서 세포에서 상응하는 효소 활성을 증가시켜 시알릴화 사카라이드의 세포내 생산을 향상시킨다.
추가 및/또는 대안적 실시양태에서, 적어도 하나의 유전자 조작된 세포는 유전자 조작되기 전의 세포와 비교하여 β-갈락톡시다제 활성, 글루코사민-6-포스페이트 데아미나제, N-아세틸글루코사민-6-포스페이트 데아세틸라제, N-아세틸만노사민 키나제, N-아세틸만노사민-6-포스페이트 에피머라제 및 N-아세틸뉴라민산 알돌라제로 이루어진 군으로부터 선택된 하나 이상의 효소 활성이 결핍되거나 감소된 활성을 보유한다.
추가 및/또는 대안적 실시양태에서, β-갈락톡시다제, 글루코사민-6-포스페이트 데아미나제, N-아세틸글루코사민-6-포스페이트 데아세틸라제, N-아세틸만노사민 키나제, N-아세틸만노사민-6-포스페이트 에피머라제 및 N-아세틸뉴라민산 알돌라제를 코딩하는 유전자의 하나 이상은 유전자 조작된 세포의 게놈으로부터 결실되었거나 β-갈락톡시다제, 글루코사민-6-포스페이트 데아미나제, N-아세틸글루코사민-6-포스페이트 데아세틸라제, N-아세틸만노사민 키나제, N-아세틸만노사민-6-포스페이트 에피머라제 및 N-아세틸뉴라민산 알돌라제를 코딩하는 유전자의 하나 이상의 발현이 불활성화되었거나 세포의 추가 유전자 조작에 의해 유전자 조작된 세포에서 적어도 감소된다. 상기 유전자의 발현은 상기 유전자의 감소된 발현을 보유하도록 추가로 유전자 조작되기 전에 추가로 유전자 조작된 세포의 전구 세포와 비교하여 추가로 유전자 조작된 세포에서 감소된다.
유전자 조작된 미생물 세포, 바람직하게는 원핵 세포. 적절한 미생물 세포는 효모 세포, 박테리아 세포, 아르케박테리아(archaebacterial) 세포, 조류 세포, 및 진균 세포를 포함한다.
추가 및/또는 대안적 실시양태에서, 유전자 조작된 미생물 세포는 박테리아 세포, 바람직하게는 바실러스(Bacillus), 락토바실러스(Lactobacillus), 락토코쿠스(Lactococcus), 엔테로코쿠스(Enterococcus), 비피도박테리움( Bifidobacterium), 스포로락토바실러스(Sporolactobacillus) 종, 마이크로모모스포라(Micromomospora) 종, 마이크로코쿠스(Micrococcus) 종, 로도코쿠스(Rhodococcus) 종, 및 슈도모나스(Pseudomonas)로 이루어진 군으로부터 선택된 박테리아 세포이다. 적합한 박테리아 종은 바실러스 서브틸리스(Bacillus subtilis), 바실러스 리체니포르미스(Bacillus licheniformis), 바실러스 코아룰란즈(Bacillus coagulans), 바실러스 서모필루스(Bacillus thermophilus), 바실러스 라테로스포러스(Bacillus laterosporus), 바실러스 메가테리움(Bacillus megaterium), 바실러스 미코이데스(Bacillus mycoides), 바실러스 푸밀러스(Bacillus pumilus), 바실러스 렌터스( Bacillus lentus), 바실러스 세레루스(Bacillus cereus), 바실러스 서르쿨란즈(Bacillus circulans), 비피도박테리움 롱굼(Bifidobacterium longum), 비피도박테리움 인펀티스(Bifidobacterium infantis), 비피도박테리움 비피둠(Bifidobacterium bifidum), 시트로박터 프룬디이(Citrobacter freundii), 클로스트리디움 셀룰로리티컴(Clostridium cellulolyticum), 클로스트리디움 융달리(Clostridium ljungdahlii), 클로스트리디움 아우토에타노게눔(Clostridium autoethanogenum), 클로스트리디움 아세토부틸리쿰(Clostridium acetobutylicum), 코리네박테리움 글루타미쿰(Corynebacterium glutamicum), 엔테로코쿠스 패시움(Enterococcus faecium), 엔테로코쿠스 써모필레스(Enterococcus thermophiles), 에스케리치아 콜라이, 에르이니아 헤르비콜라(Erwinia herbicola) (판토에아 아글로메란즈(Pantoea agglomerans)), 락토바실러스 악시도필루스(Lactobacillus acidophilus), 락토바실러스 살리바리우스(Lactobacillus salivarius), 락토바실러스 플란타룸(Lactobacillus plantarum), 락토바실러스 헬베티쿠스(Lactobacillus helveticus), 락토바실러스 델브루엑키이(Lactobacillus delbrueckii), 락토바실러스 램노수스(Lactobacillus rhamnosus), 락토바실러스 불가리쿠스(Lactobacillus bulgaricus), 락토바실러스 크리스파투스(Lactobacillus crispatus), 락토바실러스 가세리(Lactobacillus gasseri), 락토바실러스 카세이(Lactobacillus casei), 락토바실러스 류테리(Lactobacillus reuteri), 락토바실러스 젠세니이(Lactobacillus jensenii), 락토코커스 락티스(Lactococcus lactis), 판토에아 시트레아(Pantoea citrea), 펙토박테리움 카로토보룸(Pectobacterium carotovorum), 프로프리오니박테리움 프류덴라이치이(Proprionibacterium freudenreichii), 슈도모나스 플루오레센스(Pseudomonas fluorescens), 슈도모나스 아에루기노사(Pseudomonas aeruginosa), 스트렙토코커스 써모필레스(Streptococcus thermophiles) 및 크산토모나스 캄페스트리스(Xanthomonas campestris)이다.
대안적 실시양태에서, 유전자 조작된 세포는, 바람직하게는 사카로마이세스 종, 특히 사카로마이세스 세레비지애, 사카로마이콥시스(Saccharomycopsis) 종, 피티아(Pichia) 종, 특히 피치아(Pichia pastoris), 한세눌라(Hansenula) 종, 클루이베로마이세스(Kluyveromyces) 종, 요로위아(Yarrowia) 종, 로도톨루라(Rhodotorula) 종, 및 스키조사카로마이세스(Schizosaccharomyces) 종으로 이루어진 군으로부터 선택된 효모 세포이다.
유전자 조작된 세포는 NeuNAc 생합성 경로, 시티딘 5'-모노포스포- (CMP)-시알산 신테타제 활성, 및 시알릴트랜스퍼라제 활성을 포함하도록 유전자 조작되었다.
본원에 사용된 바와 같은 용어 "유전자 조작된"은 분자 생물학적 방법을 사용하여 미생물 세포의 유전적 구성(make-up)의 변형을 지칭한다. 미생물 세포의 유전적 구성의 변형은 종 경계 이내 및/또는 종 경계를 가로질러 유전자의 전달, 뉴클레오티드, 트리플렛, 유전자, 오픈 리딩 프레임, 프로모터, 인핸서, 종결자 및 유전자 발현을 매개 및/또는 제어하는 뉴클레오티드 서열의 삽입, 결실, 대체 및/또는 변형을 포함한다. 미생물 세포의 유전적 구성의 변형은 특정한, 원하는 특성을 보유한 유전자 변형된 유기체를 생성하는 것을 목표로 한다. 유전자 조작된 미생물 세포는 세포의 천연 (그러나 유전자 조작되지 않은) 형태에 존재하지 않는 하나 이상의 유전자를 함유할 수 있다. 세포의 유전 정보의 뉴클레오티드 서열을 삽입, 결실 또는 변경하기 위해 외인성 핵산 분자 (재조합, 이종유래)를 세포의 유전 정보에 삽입하고/하거나 외인성 핵산 분자를 도입하는 기술은 통상의 기술자에게 공지되어 있다. 유전자 조작된 미생물 세포는 세포의 천연 형태에 존재하는 하나 이상의 유전자를 함유할 수 있으며, 여기서 상기 유전자는 인공적 수단에 의해 미생물 세포로 변형되고 재-도입된다. 용어 "유전자 조작된"은 또한 세포에 내인성인 핵산 분자를 함유하고, 세포로부터 핵산 분자를 제거하지 않고 변형된 미생물 세포를 포함한다. 이러한 변형은 유전자 대체, 부위-특이적 돌연변이, 및 관련 기술에 의해 수득된 것들을 포함한다.
본원에 사용된 바와 같은 용어 "이종유래(heterologous)"은 세포 또는 유기체, 즉 폴리펩티드, 아미노산 서열, 핵산 분자 또는 뉴클레오티드 서열에 대해 이질적이며 상기 세포 또는 유기체에서 자연적으로 발생하지 않는 폴리펩티드, 아미노산 서열, 핵산 분자 또는 뉴클레오티드 서열을 지칭한다. 본원에 사용된 바와 같은, "이종유래 서열" 또는 "이종유래 핵산" 또는 "이종유래 폴리펩티드"는 특정한 숙주 세포 (예를 들어, 상이한 종으로부터)에 이질적인 공급원으로부터 유래하거나, 동일한 공급원으로부터 유래하는 경우는, 그의 원래 형태로부터 변형된 것이다. 따라서, 프로모터에 작동 가능하게 연결된 이종유래 핵산은 프로모터가 유래된 것과는 상이한 공급원으로부터 유래하거나, 동일한 공급원으로부터인 경우는, 그의 원래 형태로부터 변형된다. 이종유래 서열은, 예를 들어 형질감염, 형질전환, 접합 또는 형질도입에 의해 숙주 미생물 숙주 세포의 게놈에 안정적으로 도입될 수 있으며, 따라서 유전자 변형된 숙주 세포를 나타낸다. 서열이 도입될 숙주 세포에 따라 달라지는 기술이 적용될 수 있다. 다양한 기술이 관련 기술분야의 통상의 기술자에게 공지되어 있고, 예를 들어, 문헌 [Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Ed., Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y. (1989)]에 개시되어 있다. 따라서, "이종유래 폴리펩티드"는 세포에서 자연적으로 발생하지 않는 폴리펩티드이고, "이종유래 시알릴트랜스퍼라제"는 미생물 세포에서 자연적으로 발생하지 않는 시알릴트랜스퍼라제이다.
한 측면에서, 시알릴화 사카라이드가 발효에 의해, 즉, 본원에 앞서 제시된 바와 같이 유전자 조작된 미생물 세포를 사용하여, 전체 세포 생체촉매작용(biocatalysis)에 의해 생산될 수 있는 방법이 제공된다. 상기 시알릴화 사카라이드의 생산은 N-아세틸글루코사민, N-아세틸만노사민 및/또는 N-아세틸뉴라민산을 발효 브로쓰에 첨가하는 것 및/또는 시알릴화 사카라이드의 세포내 생합성을 위한 N-아세틸글루코사민, N-아세틸만노사민 및/또는 N-아세틸뉴라민산의 존재 하에 유전자 조작된 미생물 세포를 배양하는 것을 필요로 하지 않는다.
상기 방법에서, 적어도 하나의 유전자 조작된 미생물 세포는 발효 브로쓰에서 그리고 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 사카라이드의 생산에 허용되는 조건 하에 배양된다.
추가 및/또는 대안적 실시양태에서, 발효 브로쓰는 적어도 하나의 탄소 공급원을 함유하고, 적어도 하나의 탄소 공급원은 바람직하게는 글루코스, 프럭토스, 수크로스, 글리세롤 및 그의 조합으로 이루어진 군으로부터 선택된다.
상기 공정 및 유전자 변형/조작된 미생물 세포가 발효 브로쓰에서 탄소 공급원을 사용하지만, 글루코사민 및/또는 N-아세틸뉴라민산 및/또는 N-아세틸글루코사민 및/또는 N-아세틸만노사민을 발효 브로쓰에 첨가할 필요가 없는데, 그 이유는 N-아세틸뉴라민산이 유전자 조작된 미생물 세포에 의해 세포내에서 생산되기 때문이다. 따라서, 추가 및 또는 대안적 실시양태에서, 적어도 하나의 유전자 조작된 미생물 세포는 글루코사민, N-아세틸글루코사민, N-아세틸만노사민 및 N-아세틸뉴라민산으로 이루어진 군으로부터 선택된 하나 이상의 부재 하에 및/또는 그의 첨가 없이 배양된다. 유전자 조작된 미생물 세포는 갈락토스가 시알릴트랜스퍼라제 반응을 위한 수용체 기질로서 공급되지 않는 한, 갈락토스의 부재 하에 및/또는 첨가 없이 배양될 수 있다. 추가 및/또는 대안적 실시양태에서, 적어도 하나의 유전자 조작된 미생물 세포는 하나 이상의 모노사카라이드 (예를 들어 갈락토스), 디사카라이드 (예를 들어 락토스), 트리사카라이드 (예를 들어 락토-N-트리오스 II), 테트라사카라이드 (예를 들어 락토-N-테트라오스) 및/또는 펜타사카라이드 (예를 들어 시알릴락토-N-테트라오스 a)의 존재 하에 배양된다.
추가 및/또는 대안적 실시양태에 따르면, 적어도 하나의 유전자 조작된 미생물 세포는 갈락토스, N-아세틸갈락토사민, N-아세틸글루코사민, 락토스, 락툴로스, N-아세틸락토사민, 락토-N-비오스, 락토-N-트리오스, 2'-푸코실-락토스, 3-푸코실락토스, 3'-시알릴락토스, 6'-시알릴락토스, 3'-시알릴-N-아세틸락토사민, 6'-시알릴-N-아세틸락토사민, 3'-갈락토실락토스, 6'-갈락토실락토스, 락토-N-트리오스 II, 락토-N-테트라오스, 락토-N-네오테트라오스, 2'3-디푸코실-락토스, 3-푸코실-3'-시알릴락토스 및 3-푸코실-6'-시알릴락토스로 이루어진 군으로부터 선택된 적어도 하나의 수용체 기질의 존재 하에 배양된다. 이들 기질은 세포로 내수송되어 세포에서 수용체 분자로 사용된다.
유전자 조작된 세포는 시알릴화 올리고사카라이드의 성장, 증식 및 생산을 위한 탄소 공급원을 필요로 한다. 추가 및/또는 대안적 실시양태에서, 유전자 조작된 세포는 저렴한 유일한 탄소 공급원, 예컨대 - 예를 들어 - 글리세롤, 글루코스 또는 수크로스 상에서 성장할 수 있다. 상기 유일한 탄소 공급원은 유전자 조작된 세포에서 CMP-시알산 생합성을 위한 유리체(educt)를 제공한다. 따라서, 시알릴화 올리고사카라이드의 생산을 위해, Neu5Ac, ManNAc, GlcNAc 또는 글루코사민 (GlcN)의 존재 하에 유전자 조작된 세포를 배양할 필요는 없다.
상기 방법은 발효 브로쓰에서 그의 배양 동안 적어도 하나의 유전자 조작된 미생물 세포에 의해 생산된 시알릴화 사카라이드를 회수하는 임의적 단계를 포함한다. 시알릴화 사카라이드는 유전자 조작된 미생물 세포가 제거된 후, 예를 들어 원심분리에 의해 발효 브로쓰로부터 회수될 수 있고/있거나, 예를 들어 세포가 원심분리에 의해 발효 브로쓰로부터 수확되고 세포 용해 단계에 적용된다는 점에서 세포로부터 회수될 수 있다. 후속적으로, 시알릴화 사카라이드는 통상의 기술자에게 공지된 적합한 기술에 의해 발효 브로쓰 및/또는 세포 용해물로부터 추가로 정제될 수 있다. 적합한 기술은 미세여과, 한외여과, 투석여과, 모의 이동층 유형 크로마토그래피, 전기 투석, 역삼투, 겔 여과, 음이온 교환 크로마토그래피, 양이온 교환 크로마토그래피 등을 포함한다.
상기 방법 및 상기 방법에 사용된 유전자 조작된 미생물 세포는 시알릴화 사카라이드의 생산에 사용된다. 용어 "시알릴화 사카라이드"는 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 사카라이드 분자를 지칭한다.
추가 및/또는 대안적 실시양태에서, 시알릴화 사카라이드는 올리고사카라이드이다. 본원에 사용된 바와 같은 용어 "올리고사카라이드(oligosaccharide)"는 모노사카라이드 잔기의 중합체를 지칭하며, 여기서 상기 중합체는 적어도 2개의 모노사카라이드 잔기, 그러나 10개 이하의 모노사카라이드 잔기, 바람직하게는 7개 이하의 모노사카라이드 잔기를 포함한다. 올리고사카라이드는 모노사카라이드의 선형 쇄이거나 분 지형이다. 게다가, 올리고사카라이드의 모노사카라이드 잔기는 다수의 화학적 변형을 특색으로 할 수 있다. 따라서, 올리고사카라이드는 하나 이상의 비-사카라이드 모이어티를 포함할 수 있다. 본원에 사용된 바와 같은 용어 "시알릴화 올리고사카라이드"는 하나 이상의 N-아세틸뉴라민산 모이어티를 포함하는 올리고사카라이드를 지칭한다.
추가 및/또는 대안적 실시양태에 따르면, 시알릴화 올리고사카라이드는 3'-시알릴락토스, 6'-시알릴락토스, 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c, 푸코실-시알릴락토-N-테트라오스 a, 푸코실-시알릴락토-N-테트라오스 b, 푸코실-시알릴락토-N-테트라오스 c, 디시알릴락토-N-테트라오스, 푸코실디시알릴락토-N-테트라오스 I, 푸코실디시알릴락토-N-테트라오스 II, 3'-시알릴갈락토스, 6'-시알릴갈락토스, 3'-시알릴-N-아세틸락토사민 및 6'-시알릴-N-아세틸락토사민으로 이루어진 군으로부터 선택된다.
본 발명의 또 다른 측면에서, 전체 세포 발효 공정에서 시알릴화 사카라이드의 생산을 위한 본원에 앞서 기재된 바와 같은 유전자 조작된 미생물 세포의 용도가 제공되며, 즉 시알릴화 사카라이드는 유전자 조작된 미생물 세포에 의해 합성된다.
본 발명의 또 다른 측면에서, 상기 방법에 의해 및/또는 본원에 앞서 기재된 바와 같은 유전자 조작된 미생물 세포를 사용함으로써 생산된 시알릴화 사카라이드가 제공된다. 추가 및/또는 대안적 실시양태에서, 시알릴화 사카라이드는 시알릴화 올리고사카라이드, 바람직하게는 3'-시알릴락토스, 6'-시알릴락토스, 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c, 푸코실-시알릴락토-N-테트라오스 a, 푸코실-시알릴락토-N-테트라오스 b, 푸코실-시알릴락토-N-테트라오스 c, 디시알릴락토-N-테트라오스, 푸코실디시알릴락토-N-테트라오스 I, 푸코실디시알릴락토-N-테트라오스 II, 3'-시알릴-갈락토스, 6'-시알릴갈락토스, 3'-시알릴-N-아세틸락토사민 및 6'-시알릴-N-아세틸-락토사민으로 이루어진 군으로부터 선택된 시알릴화 올리고사카라이드이다.
본 발명의 또 다른 측면에서, 앞서 본원에 기재된 바와 같은 방법에 의해 및/또는 영양 조성물의 제조를 위한 앞서 본원에 기재된 바와 같은 유전자 조작된 미생물 세포를 사용함으로써 생산된 시알릴화 사카라이드의 용도가 제공된다.
따라서, 본 발명의 또 다른 측면에 따르면, 앞서 본원에 기재된 바와 같은 방법에 의해 및/또는 유전자 조작된 미생물 세포에 의해 생산된, 적어도 하나의 시알릴화 사카라이드, 바람직하게는 적어도 하나의 시알릴화 올리고사카라이드를 함유하는 영양 조성물이 제공된다. 추가 및/또는 대안적 실시양태에서, 시알릴화 올리고사카라이드는 3'-시알릴락토스, 6'-시알릴락토스, 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c, 푸코실-시알릴락토-N-테트라오스 a, 푸코실-시알릴락토-N-테트라오스 b, 푸코실-시알릴락토-N-테트라오스 c, 디시알릴락토-N-테트라오스, 푸코실디시알릴락토-N-테트라오스 I, 푸코실디시알릴락토-N-테트라오스 II로 이루어진 군으로부터 선택된다.
추가 및/또는 대안적 실시양태에서, 영양 조성물은 적어도 하나의 중성 HMO, 바람직하게는 2'-FL을 추가로 함유한다.
추가 및/또는 대안적 실시양태에서, 영양 조성물은 3-SL, 6-SL 및 2'-FL을 함유한다.
추가 실시양태에서, 영양 조성물은 의약, 제약, 제제, 유아용 유동식(Infant formula) 및 식이 보충제로 이루어진 군으로부터 선택된다.
영양 조성물은 분말, 과립, 플레이크 및 펠렛을 포함하나, 이에 제한되지는 않는 액체 형태 또는 고체 형태로 존재할 수 있다.
본 발명은 특정한 실시양태와 관련하여 더 기재될 것이나, 본 발명은 이에 제한되는 것이 아니라 청구범위에 의해서만 제한된다. 더욱이, 설명 및 청구범위에서 용어 제1, 제2 등은 유사한 요소를 구별하기 위해 사용되며 시간적으로, 공간적으로, 순위로 또는 임의의 다른 방식으로 순서를 기재하기 위해 반드시 필요한 것은 아니다. 이렇게 사용된 용어는 적절한 상황에서 상호교환 가능하고, 본원에 기재된 본 발명의 실시양태가 본원에 기재되거나 예시된 것과 다른 순서로 작동 가능하다는 것을 이해하여야 한다.
청구범위에서 사용된 용어 "포함하는"은, 이후에 열거된 수단에 제한되는 것으로 해석되어서는 안되며; 상기 용어는 다른 요소 또는 단계를 제외하지 않는다는 점에 유의하여야 한다. 따라서 언급된 바와 같이 서술된 특색, 정수, 단계 또는 구성요소의 존재를 구체화하는 것으로 해석되어야 하나, 하나 이상의 다른 특색, 정수, 단계 또는 구성요소, 또는 그의 군의 존재 또는 첨가를 배제하지 않는다. 따라서, 표현 "수단 A 및 B를 포함하는 장치"의 범위는 구성요소 A 및 B로만 이루어진 장치로 제한되지 않아야 한다. 이는 본 발명과 관련하여, 장치의 유일한 관련 구성요소가 A 및 B임을 의미한다.
본 명세서의 전체에 걸쳐 "한 실시양태" 또는 "일 실시양태"에 대한 언급은 실시양태와 관련하여 기재된 특정한 특색, 구조 또는 특성이 본 발명의 적어도 하나의 실시양태에 포함된다는 것을 의미한다. 따라서, 본 명세서 전체에 걸쳐 다양한 곳에서 어구 "한 실시양태에서" 또는 "일 실시양태에서"의 출현은 반드시 모두 동일한 실시양태를 지칭하는 것은 아니나, 지칭할 수도 있다. 더욱이, 특정한 특색, 구조 또는 특성은 하나 이상의 실시양태에서, 본 개시내용으로부터 관련 기술분야의 통상의 기술자에게 명백한 바와 같이, 임의의 적합한 방식으로 조합될 수 있다.
유사하게, 본 발명의 예시적인 실시양태의 설명에서, 본 발명의 다양한 특색은 때때로 개시내용을 간소화하고 다양한 발명적 측면 중 하나 이상의 이해를 지원하기 위해 단일 실시양태에서, 도면, 또는 그의 설명과 함께 그룹화된다는 것을 인식하여야 한다. 그러나, 이 개시 방법은, 청구된 발명이 각각의 청구항에 명시적으로 언급된 것보다 더 많은 특색을 필요로 한다는 의도를 반영하는 것으로서 해석되어서는 안된다. 오히려, 하기 청구범위가 반영하는 바와 같이, 발명적 측면은 단일의 전술한 개시된 실시양태의 모든 특색보다 적게 있다. 따라서, 상세한 설명 후의 청구범위는 이 상세한 설명에 명시적으로 혼입되고, 각각의 청구항은 본 발명의 개별 실시양태로서 그 자체로 존재한다.
더욱이, 본원에 기재된 일부 실시양태는 다른 실시양태에 포함된 다른 특색은 아니나 일부를 포함하지만, 상이한 실시양태의 특색의 조합은 본 발명의 범위 내에 있고, 관련 기술분야의 통상의 기술자에 의해 이해되는 바와 같이 상이한 실시양태를 형성하는 것을 의미한다. 예를 들어, 하기 청구항에서, 청구된 실시양태 중 임의의 것이 임의의 조합으로 사용될 수 있다.
더욱이, 실시양태 중 일부는 컴퓨터 시스템의 프로세서에 의해 또는 기능을 수행하는 다른 수단에 의해 구현될 수 있는 방법의 요소의 조합 또는 방법으로서 본원에 기재된다. 따라서, 이러한 방법 또는 방법의 요소를 수행하는 데 필요한 지침을 가진 프로세서는 방법 또는 방법의 요소를 수행하기 위한 수단을 형성한다. 더욱이, 장비 실시양태의 본원에 기재된 요소는 본 발명을 수행하기 위해 요소에 의해 수행되는 기능을 수행하기 위한 수단의 예이다.
본원에 제공된 설명 및 도면에서, 다수의 구체적 세부사항이 제시된다. 그러나, 본 발명의 실시양태는 이들 구체적 세부사항 없이도 실시될 수 있음이 이해된다. 다른 경우에, 널리 공지된 방법, 구조 및 기술은 이 설명의 이해를 모호하게 하지 않기 위해 상세히 나타내지 않았다.
본 발명은 이제 본 발명의 몇몇 실시양태의 상세한 설명에 의해 설명될 것이다. 본 발명의 다른 실시양태는 본 발명의 진정한 사상 또는 기술적 교시내용을 벗어나지 않고 관련 기술분야의 통상의 기술자의 지식에 따라 구성될 수 있으며, 본 발명은 첨부된 청구범위의 조건에 의해서만 제한된다는 것이 분명하다.
실시예
도 1 내지 도 3은 NeuNAc, CMP-NeuNAc 및 시알릴화 사카라이드의 세포내 생합성을 위한 대안적 경로를 나타내는 도식을 나타낸다.
본원에 기재된 바와 같이 유전자 변형된 세포로, 시알릴화 사카라이드의 발효 생산이 달성될 수 있다. 제공된 유일한 탄소 공급원 (예를 들어 수크로스)은 미생물 세포로 내수송되어 대사되어 프럭토스-6-포스페이트를 산출한다 (도 1 내지 도 3). 다음으로, L-글루타민: D-프럭토스-6-포스페이트 아미노트랜스퍼라제 (GlmS)는 프럭토스-6-포스페이트의 글루코사민-6-포스페이트로의 전환을 수행하며 (도 1 내지 도 3), 이는 결국 글루코사민-6-포스페이트 N-아세틸-트랜스퍼라제 (Gna1)에 의해 N-아세틸글루코사민-6-포스페이트로 대사된다 (도 2 및 도 3). N-아세틸글루코사민-6-포스페이트는 i) N-아세틸글루코사민-6-포스페이트 에피머라제 (NanE)에 의해 N-아세틸만노사민-6-포스페이트로 전환되고 추가로 N-아세틸만노사민-6-포스페이트 포스파타제에 의해 N-아세틸만노사민으로 전환될 수 있거나 (도 3) ii) N-아세틸글루코사민-6-포스페이트 포스파타제 (YihX/YqaB)에 의해 N-아세틸글루코사민로 전환되고 추가로 N-아세틸글루코사민 2-에피머라제 (Slr1975)에 의해 N-아세틸만소사민으로 대사될 수 있다 (도 2). 시알산 신타제 (NanA)는 N-아세틸만노사민을 N-아세틸 뉴라민산으로 전환하고, 이는 CMP-시알산 신테타제에 의해 CMP-N-아세틸뉴라민산으로 전환된다 (도 1 내지 도 3). 수용체 기질은 배양 브로쓰에 공급되고 세포로 내수송되어 재조합 숙주 세포에 의해 변형되거나 네노보(de novo) 합성될 수 있다. 수용체 기질은 시알릴트랜스퍼라제 (SiaT)에 의해 촉매되는 반응에서 N-아세틸뉴라민산과 라이게이션되어 시알릴화 사카라이드를 산출하며, 이는 배양 브로쓰로 외수송될 수 있다.
실시예 1: 다양한 시알릴화 올리고사카라이드의 생산
특성화되거나 추정되는 시알릴트랜스퍼라제의 유전자 서열은 문헌 및 공개 데이터베이스로부터 수령하였다. 시알릴트랜스퍼라제는 그의 신호 펩티드가 결실되는 경우 더 높은 활성을 나타내는 것으로 종종 기재되기 때문에, 우리는 온라인 예측 도구 시그널(Signal)P (Petersen et al., Nature Methods, 2011 Sep 29;8(10):785-6)에 의해 상응하는 단백질 서열을 분석하였다. 유전자는 주석이 달린 바와 같이, 전장 형태로 또는, 신호 펩티드가 예측되는 경우, N-말단 신호 펩티드가 결핍된 말단절단된 변이체로서 진스크립트 코포레이션(GenScript cooperation)에 의해 합성적으로 합성되었다.
시알릴트랜스퍼라제 1 내지 26을 각각 유전자 특이적 프라이머를 사용하여 SLIC에 의해 pDEST14에 neuA와 함께 오페론으로 서브클로닝하여, 일반적인 종류의 플라스미드: pDEST14-siaT-neuA를 산출하였다. 나머지 시알릴트랜스퍼라제 27 내지 100은 제한 부위 NdeI 및 BamHI를 사용하여 진스크립트 코포레이션에 의해 플라스미드 pET11a로 직접 서브클로닝하였다. 두 발현 시스템 모두 IPTG-유도성 유전자 발현을 가능하게 한다. 시험관내 활성 스크리닝을 위해, 플라스미드를 LacZ 활성이 결핍된 이. 콜라이 BL21 (DE3) 균주로 형질전환시켰다.
siaT9 (α-2,3-시알릴트랜스퍼라제) 및 siaT18 (α-2,6-시알릴트랜스퍼라제) 발현을 위한 플라스미드를 보유한 이. 콜라이 균주를 암피실린 100 μg ml-1이 보충된 20 ml의 2YT 배지가 채워진 100 ml 진탕 플라스크에서 30℃에서 성장시켰다. 배양이 0.1 내지 0.3의 OD600에 도달한 경우, 0.3 mM IPTG를 첨가하여 유전자 발현을 유도하고 12 내지 16 시간 동안 배양을 계속하였다. 세포를 원심분리에 의해 수확하고 유리 비드를 사용하여 규정된 부피의 50 mM Tris-HCl pH 7.5에서 기계적으로 파괴하였다. 단백질 추출물은 검정이 시작될 때까지 얼음 위에 유지시켰다. 시험관 검정은 50 mM Tris-HCl pH7.5, 5 mM MgCl2, 10 mM CMP-Neu5Ac 및 5 내지 20 mM의 적절한 수용체 기질을 포함한 25 μl의 총 부피에서 수행하였다. 검정은 3 μl 단백질 추출물의 첨가로 시작하여 16시간 동안 계속하였다. 시알릴트랜스퍼라제의 활성으로부터 생긴 시알릴화 올리고사카라이드의 형성은 박층 크로마토그래피에 의해 결정하였다.
따라서, 샘플은 실리카겔 60 F254 (머크 카게아아(Merck KGaA), 독일 다름슈타트)-플레이트에 적용하였다. 부탄올:아세톤:아세트산:H2O (35/35/7/23 (v/v/v/v))의 혼합물을 이동상으로서 사용하였다. 분리된 물질의 검출을 위해, TLC 플레이트를 티몰 시약 (95 ml 에탄올에 용해된 0.5 g 티몰, 5 ml 황산 첨가)에 침지 가열하였다. 시알릴화 반응 생성물은 그의 수용체 기질보다 더 느리게 시행되었다.
표 3: 공급된 수용체 기질에 따라 2개의 예시적인 시알릴트랜스퍼라제의 시알릴트랜스퍼라제 활성을 결정하는 시험관 내 검정. 시알릴화 사카라이드의 형성은 박층 크로마토그래피에 의해 결정하였다. (+) 시알릴화된 반응 생성물이 검출 가능하였다. (-) 시알릴화 반응 생성물이 검출되지 않았다.
시알릴트랜스퍼라제 둘 다 적어도 하나의 갈락토스 잔기를 함유하는 갈락토스 또는 다양한 올리고사카라이드를 시알릴화하는 것이 가능하였다. 수크로스를 반응에 적용하였을 때 어떤 시알릴화 올리고사카라이드도 검출되지 않았다 (표 3).
실시예 2:
N
-아세틸뉴라민산 생산을 위한 이. 콜라이 BL21(DE3) 균주의 대사 공학
대사 공학(metabolic engineering)은 특이적 내생 유전자의 돌연변이유발 및 결실 및 이종유래 유전자의 게놈 통합에 의해 달성되었다. 유전자 lacZ 및 araA는 문헌 [Ellis et al., (Proc. Natl. Acad. Sci. USA 98: 6742-6746 (2001))]에 의해 기재된 바와 같이 미스매치-올리고뉴클레오티드를 사용한 돌연변이유발에 의해 불활성화되었다.
문헌 [Datsenko and Wanner (Proc. Natl. Acad. Sci. USA 97:6640-6645 (2000))]의 방법에 따라 게놈 결실이 생성되었다. N-아세틸글루코사민의 분해를 방지하기 위해 이. 콜라이 균주 BL21 (DE3)의 게놈으로부터 하기 유전자가 결실되었다: N-아세틸글루코사민 특이적 PTS 효소 II (nagE), N-아세틸글루코사민-6-포스페이트 데아세틸라제 (nagA), 및 글루코사민-6-포스페이트 데아미나제 (nagB). N-아세틸만노사민 키나제 (nanK), N-아세틸만노사민-6-포스페이트 에피머라제 (nanE), N-아세틸뉴라민산 알돌라제 (nanA) 및 시알산 퍼미아제 (nanT)를 코딩하는 전체 N-아세틸뉴라민산 이화 유전자 클러스터가 또한 결실되었다. 글루코사민의 내수송을 용이하게 하는 포스포에놀피루베이트-의존성 포스포트랜스퍼라제 시스템을 코딩하는, 유전자 manX, manY 및 manZ가 또한 결실되었다. wzxC-wcaJ 유전자가 또한 결실되었다. wcaJ 유전자는 콜란산 합성의 제1 단계를 촉매하는 UDP-글루코스:운데카프레닐 포스페이트 글루코스-1-포스페이트 트랜스퍼라제를 코딩한다 (Stevenson et al., J. Bacteriol. 1996, 178:4885-4893). 게다가, 유전자 fucI 및 fucK 및 agaA가 결실되어, 각각 L-푸코스 이소머라제, L-푸쿨로스 키나제, 및 N-아세틸갈락토사민-6-포스페이트 데아세틸라제를 코딩한다.
이종유래 유전자의 게놈 통합은 EZ-Tn5™ 트랜스포사제 (에피센트레(Epicentre), 미국) 또는 마리너(mariner) 트랜스포사제 Himar1의 과활성 C9-돌연변이를 사용하여, 전위에 의해 달성되었다 (Proc. Natl. Acad. Sci. 1999, USA 96:11428-11433). EZ-Tn5 트랜스포솜을 생산하기 위해 FRT-부위 플랭킹된 항생제 내성 마커 (대안적으로 저항성 마커 유전자가 lox66-lox71 부위에 의해 플랭킹되었다)와 함께 관심 유전자를 증폭시켰다. 생성된 PCR-생성물은 양쪽 말단에서 EZ-Tn5 트랜스포사제에 대한 19-bp 모자이크 말단(Mosaic End) 인식 부위를 보유하였다. Himar1 트랜스포사제를 사용한 통합을 위해 관심 발현 구축물 (오페론)을 항생제 내성 마커에 의해 플랭킹된 FRT-부위/lox66-lox71-부위와 함께 유사하게 클로닝하고 아라비노스-유도성 프로모터 ParaB의 제어 하에 마리너 트랜스포사제 Himar1의 과활성 C9-돌연변이를 코딩하는 pEcomar 벡터로 옮겼다. 모든 유전자는 이. 콜라이에서의 발현을 위해 코돈-최적화되었고 진스크립트 코포레이션(GenScript Corp)에 의해 합성적으로 제조되었다.
발현 단편 <Ptet-lacY-FRT-aadA-FRT>은 EZ-Tn5 트랜스포사제를 사용하여 통합시켰다. 이. 콜라이 K12 TG1 (진뱅크(GenBank): ABN72583)로부터 락토스 내수송체(importer) LacY에 대한 유전자의 성공적인 통합 후, 내성 유전자는 플라스미드 pCP20 상에 코딩된 FLP 레콤비나제에 의해 스트렙토마이신 내성 클론으로부터 제거되었다 (Proc. Natl. Acad. Sci. 2000, USA 97:6640-6645). 수크로스 퍼미아제, 프럭토키나제, 수크로스 히드롤라제, 및 전사 리프레서에 대한 유전자 (각각 유전자 cscB, cscK, cscA, 및 cscR)를 포함하고, 균주를 유일한 탄소 공급원으로서 수크로스 상에서 성장하게 하는 것을 가능하게 하는, 이. 콜라이 W로부터의 csc-유전자 (진뱅크: CP002185.1)는, 또한 게놈에 삽입되었다. 이 클러스터는 플라스미드 pEcomar-cscABKR을 사용한 전위에 의해 이. 콜라이 BL21(DE3) 균주의 게놈에 통합시켰다.
생성된 균주는 하기 발현 카세트의 게놈 통합에 의해 NeuNAc의 생산을 위해 추가로 변형되었다: <Ptet-slr1975-gna1-lox66-aacC1-lox71> (서열번호: 97), <Ptet-neuB-lox66-kanR-lox71> (서열번호: 98), <Ptet-slr1975-Pt5-neuB-FRT-dhfr-FRT> (서열번호: 99), <Ptet-glmS*-gna1-lox66-aacC1-lox71> (서열번호: 100) 및 <Ptet-ppsA-lox66-aacC1-lox71> (서열번호: 101). dhfr 발현 카세트를 제외하고, 모든 내성 마커 유전자를 플라스미드 pKD-Cre (서열번호: 102)를 도입한 후 30℃에서 100 μg·mL-1 암피실린 및 100 mM L-아라비노스를 함유하는 2YT 한천 플레이트 상에서 선택하여 게놈 (다음 라운드의 유전자 통합 전)으로부터 단계적으로 제거하였다. 후속적으로 내성 클론을 암피실린뿐만 아니라 게놈 통합에 사용되는 선택적 항생제가 결핍된 2YT 한천 플레이트에 옮겼다. 플라스미드 세포를 경화시키기 위해 플레이트를 42℃에서 인큐베이션하였다. 암피실린 및 선택적 항생제에 민감한 클론을 추가 실험 및 변형에 사용하였다.
유전자 slr1975 (진뱅크: BAL35720)는 시네초시스티스 종 PCC6803 N-아세틸글루코사민 2-에피머라제를 코딩한다. 유전자 gna1 (진뱅크: NPβ116637)은 사카로마이세스 세레비지애로부터의 글루코사민-6-포스페이트 아세틸트랜스퍼라제를 코딩한다. 유전자 neuB (진뱅크: AF305571)는 캄필로박터 제주니로부터의 시알산 신타제를 코딩한다. 유전자 glmS*는 이. 콜라이 L-글루타민:D-프럭토스-6-포스페이트 아미노트랜스퍼라제 유전자의 돌연변이된 버전이다 (Metab Eng. 2005 May;7(3):201-14). 유전자 ppsA (진뱅크: ACT43527)는 이. 콜라이 BL21(DE3)의 포스포에놀피루베이트 신타제를 코딩한다.
<Ptet-slr1975-gna1-lox66-aacC1-lox71>의 생성을 위해, 유전자 slr1975 및 gna1을 구성적 프로모터 P tet 뒤에 오페론으로서 서브클로닝하고 겐타마이신 내성 유전자 (lox66/lox71 부위에 의해 플랭킹됨)에 융합시키고 무딘(blunt)-말단 라이게이션에 의해 pEcomar 벡터에 삽입하였다. 생성된 발현 카세트를 벡터 pEcomar-slr195-gna1-aacC1 및 아라비노스-유도성 프로모터 ParaB의 제어 하에 마리너 트랜스포사제 Himar1의 과활성 C9-돌연변이체를 사용함으로써 게놈에 통합시켰다.
<Ptet-neuB-lox66-kanR-lox71>의 생성을 위해, neuB를 구성적 프로모터 P tet 뒤에 클로닝하고 카나마이신 내성 유전자 (lox66/lox71 부위에 의해 플랭킹됨)에 융합시켰다. 생성된 발현 카세트는 EZ-Tn5 트랜스포사제를 사용하여 게놈에 통합시켰다. <Ptet-slr1975-Pt5-neuB-FRT-dhfr-FRT>의 생성을 위해, 유전자 slr1975 및와 neuB를 각각 구성적 프로모터 P tet 및 P t5 뒤에 개별적으로 서브클로닝하고, 트리메토프림 내성 유전자 (FRT 부위에 의해 플랭킹됨)에 융합시켰다. 생성된 발현 카세트는 EZ-Tn5 트랜스포사제를 사용함으로써 게놈에 통합시켰다.
발현 카세트 <Ptet-glmS*-gna1-lox66-aacC1-lox71>는 구성적 프로모터 P tet 뒤에 오페론으로서 glmS* 및 gna1을 클로닝함으로써 생성시켰다. 이 구축물은 추가로 겐타마이신 내성 유전자 (lox66/lox71 부위에 의해 플랭킹됨)에 융합시켰다. 생성된 발현 카세트는 EZ-Tn5 트랜스포사제를 사용하여 게놈에 통합시켰다.
<Ptet-ppsA-lox66-aacC1-lox71>의 생성을 위해, ppsA 유전자를 구성적 프로모터 P tet 뒤에 클로닝하고 겐타마이신 내성 유전자 (ox66/lox71 부위에 의해 플랭킹됨)에 융합시켰다. 생성된 발현 카세트는 EZ-Tn5 트랜스포사제를 사용함으로써 게놈에 통합시켰다.
전체적으로, 누적 게놈 변형은 Neu5Ac-생산 균주 이. 콜라이 #NANA1을 생성시켰다.
실시예 3: 3'-시알릴락토스의 생산을 위한 미생물 세포주의 생성 및 배양
EZ-Tn5 트랜스포사제를 사용함으로써 <Ptet-siaT9-Pt5-neuA-lox66-aacC1-lox71> (서열번호: 103)의 게놈 통합에 의해 균주 이. 콜라이 #NANA1을 추가로 변형하여 3'-SL 생산 균주를 산출하였다. 이. 콜라이에서의 발현에 코돈-최적화되고 진스크립트에 의해 합성적으로 제조된 유전자 siaT9 (진뱅크: BAF91160)는, 비브리오 종 JT-FAJ-16으로부터의 α-2,3-시알릴트랜스퍼라제를 코딩한다. 유전자 neuA (진뱅크: AF305571)는 캄필로박터 제주니로부터의 CMP-시알산 신테타제를 코딩한다.
균주의 배양은 96-웰 플레이트에서 수행하였다. 따라서, 균주의 단일 콜로니를 한천 플레이트로부터 7 g l-1 NH4H2PO4, 7 g l-1 K2HPO4, 2 g l-1 KOH, 0.3g l-1 시트르산, 5 g l-1 NH4Cl, 1 ml l-1 소포제, 0.1 mM CaCl2, 8 mM MgSO4, 미량 원소 및 2% 수크로스를 탄소 공급원으로서 함유하는 200 μL의 최소 배지를 함유하는 미세역가 플레이트로 옮겼다. 미량 원소는 0.101 g l-1 니트릴로트리아세트산, pH 6.5, 0.056 g l-1 시트르산제이철암모늄, 0.01 g l-1 MnCl2 x 4 H2O, 0.002 g l-1 CoCl2 x 6 H2O, 0.001g l-1 CuCl2 x 2 H2O, 0.002 g l-1 붕산, 0.009 g l-1 ZnSO4 x 7 H2O, 0.001 g l-1 Na2MoO4 x 2 H2O, 0.002 g l-1 Na2SeO3, 0.002 g l-1 NiSO4 x 6 H2O로 이루어졌다. 격렬하게 진탕하면서 30℃에서 대략 20시간 동안 배양을 수행하였다. 후속적으로, 50 μL의 배양 브로쓰를 웰당 400 μL의 최소 배지를 함유하는 딥웰 96 웰 플레이트 (2.0 mL)로 옮겼다.
48시간 더 인큐베이션 후, 배양을 중단하고 질량 분석법에 의해 상청액 중 3'-시알릴락토스 수준을 결정하였다. LC 삼중-사중극자(Triple-Quadrupole) MS 검출 시스템을 사용하여 MRM (다중 반응 모니터링)에 의해 질량 분석법 분석을 수행하였다. 전구체 이온을 사중극자 1에서 선택 및 분석하고, CID 가스로서 아르곤을 사용하여 충돌 셀에서 단편화(fragmentation)를 실시하고, 단편 이온의 선택을 사중극자 3에서 수행하였다. 배양 상청액을 H2O (LC/MS 등급)로 1:100으로 희석한 후 락토스, 3'-시알릴락토스 및 6'-시알릴락토스의 크로마토그래피 분리를, 엑스브릿지(XBridge) 아미드 HPLC 컬럼 (3.5 μm, 2.1 50 mm (워터스(Waters), 미국)과 엑스브릿지 아미드 가드(guard) 카트리지 (3.5 μm, 2.1 10 mm) (워터스, 미국) 상에서 수득하였다. HPLC 시스템의 컬럼 오븐 온도는 50℃였다. 이동상은 10 mM 암모늄 아세테이트와 함께 아세토니트릴:H2O로 구성되었다. 1 μl 샘플을 상기 기기에 주입하고; 400 μl/min의 유량으로 3.60분 동안 시행을 수행하였다. 3'-시알릴락토스 및 6'-시알릴락토스는 ESI 양이온화 모드에서 MRM에 의해 분석하였다. 질량 분석기는 단위 분해능(unit resolution)으로 작동되었다. 시알릴락토스는 m/z 656.2 [M+Na]의 이온을 형성한다. 시알릴락토스의 전구체 이온은 충돌 셀에서 단편 이온 m/z 612.15, m/z 365.15 및 m/z 314.15로 추가로 단편화되었다. 충돌 에너지, Q1 및 Q3 프리 바이어스(Pre Bias)는 각각의 분석물에 대해 개별적으로 최적화되었다. 정량화 방법은 시판되는 표준 (카르보신쓰(Carbosynth), 영국 콤프턴)을 사용하여 확립하였다. 배양의 종료시 대략 0.6 g L-1의 배양 상청액 중 3'-SL 역가에 도달하였다.
실시예 4: 6'-시알릴락토스 생산을 위한 미생물 세포주의 생성 및 배양
EZ-Tn5 트랜스포사제를 사용함으로써 <Ptet-siaT18-Pt5-neuA-lox66-aacC1-lox71> (서열번호: 104)의 게놈 통합에 의해 균주 이. 콜라이 #NANA1을 추가로 변형하여 6'-SL 생산 균주를 산출하였다. 이. 콜라이에서의 발현에 코돈-최적화되고 진스크립트에 의해 합성적으로 제조된 유전자 siaT18 (진뱅크: AB500947)는, 포토박테리움 레이오그나티 JT-SHIZ-119로부터의 α-2,6-시알릴트랜스퍼라제를 코딩한다. 유전자 neuA (진뱅크: AF305571)는 캄필로박터 제주니로부터의 CMP-시알산 신테타제를 코딩한다.
실시예 2에 기재된 바와 같이, 이 6'-SL 생산 균주를 사용하여 96-웰 플레이트에서의 배양을 수행하였다. 배양의 종료시 대략 0.9 g L-1의 배양 상청액 중 6'-SL 역가에 도달하였다.
실시예 5: 시알릴락토스를 함유하는 유아용 유동식의 조성물
SEQUENCE LISTING
<110> Jennewein Biotechnologie GmbH
<120> Production of sialylated saccharides
<130> P 1802 WO
<160> 104
<170> PatentIn version 3.5
<210> 1
<211> 1410
<212> DNA
<213> Campylobacter coli
<400> 1
atgcaaaacg tcattatcgc tggtaacggt ccgagcctgc aatcaatcaa ctatcaacgc 60
ctgccgaaag aatacgacat cttccgctgc aaccagttct acttcgaaga taaatactac 120
ctgggcaaaa acatcaaagc ggcctttttc aatccgtatc cgttcctgca gcaataccat 180
accgcgaaac agctggtgtt caacaacgaa tacaaaatcg aaaacatctt ttgtagcacg 240
ttcaatctgc cgttcatcga aaaagataac ttcatcaaca aattttacga tttctttccg 300
gacgctaaac tgggtcacaa aatcatcgaa aacctgaaag aattttacgc gtacatcaaa 360
tacaacgaaa tctacctgaa caaacgtatt accagcggca tctatatgtg cgcaattgct 420
atcgcgctgg gttataaaaa catttacctg tgtggcatcg atttctatga aggtgaaacg 480
atctacccgt tcaaagccat gtctaaaaac attaagaaaa tttttccgtg gatcaaagat 540
ttcaacccga gtaacttcca ttccaaagaa tacgacatcg aaatcctgaa actgctggaa 600
tcaatctaca aagttaacat ctacgcactg tgcgataact cggccctggc aaattacttc 660
ccgctgctgg tgaacaccga caattcattt gttctggaaa acaaatcgga tgactgtatc 720
aacgatatcc tgctgaccaa caatacgccg ggcattaact tctataaaag ccagatccaa 780
gtcaacaata ccgaaattct gctgctgaac tttcagaata tgatcagcgc caaagaaaac 840
gaaatttcta acctgaacaa aatcctgcaa gactcataca aaaccatcaa cacgaaagaa 900
aacgaaatta gtaatctgaa taaaatcctg caggattcct ataaaacgat taataccaaa 960
gaaaatgaaa tttcgaatct gaacaaaatc ctgcaggata aagacaaact gctgatcgtt 1020
aaagaaaacc tgctgaattt caaaagccgt catggtaaag ccaaatttcg cattcagaac 1080
caactgtctt ataaactggg ccaggcaatg atggtcaata gcaaatctct gctgggttat 1140
atccgtatgc cgtttgtgct gagttacatc aaagacaaac acaaacagga acaaaaaatc 1200
tatcaggaaa aaattaagaa agatccgagc ctgaccctgc cgccgctgga agattatccg 1260
gactacaaag aagctctgaa agaaaaagaa tgcctgacct atcgcctggg ccagacgctg 1320
attaaagcgg atcaagaatg gtacaaaggt ggctatgtga aaatgtggtt cgaaatcaaa 1380
aaactgaaga aagaatacaa aaagaaataa 1410
<210> 2
<211> 1146
<212> DNA
<213> Vibrio sp.
<400> 2
atgaacaacg acaactccac gaccaccaac aataacgcta ttgaaatcta tgtggatcgt 60
gcgaccctgc cgacgatcca gcaaatgacc aaaattgtta gccagaaaac gtctaacaaa 120
aaactgatct catggtcgcg ctacccgatt accgataaaa gcctgctgaa gaaaattaac 180
gcggaatttt tcaaagaaca atttgaactg acggaaagcc tgaaaaacat catcctgtct 240
gaaaacatcg ataacctgat cattcatggc aataccctgt ggagtattga tgtggttgac 300
attatcaaag aagtcaacct gctgggcaaa aatattccga tcgaactgca cttttatgat 360
gacggttccg ccgaatacgt tcgtatctac gaatttagta aactgccgga atccgaacag 420
aaatacaaaa ccagcctgtc taaaaacaac atcaaattct caatcgatgg caccgactcg 480
ttcaaaaaca cgatcgaaaa catctacggt ttcagccaac tgtatccgac cacgtaccac 540
atgctgcgtg cagatatctt cgacaccacg ctgaaaatta acccgctgcg cgaactgctg 600
tcaaacaaca tcaaacagat gaaatgggat tacttcaaag acttcaacta caaacaaaaa 660
gatatctttt actcactgac caacttcaac ccgaaagaaa tccaggaaga cttcaacaaa 720
aactcgaaca aaaacttcat cttcatcggc agtaactccg cgaccgccac ggcagaagaa 780
caaatcaata ttatcagcga agcgaagaaa gaaaacagca gcattatcac caattcaatt 840
tcggattatg acctgttttt caaaggtcat ccgtctgcca cgtttaacga acagattatc 900
aatgcacacg atatgatcga aatcaacaac aaaatcccgt tcgaagctct gatcatgacc 960
ggcattctgc cggatgccgt tggcggtatg ggtagttccg tctttttcag tatcccgaaa 1020
gaagtcaaaa acaaattcgt gttctataaa agtggtacgg atatcgaaaa taactccctg 1080
attcaggtga tgctgaaact gaatctgatt aaccgcgata atattaaact gatctctgac 1140
atttaa 1146
<210> 3
<211> 1173
<212> DNA
<213> Photobacterium sp.
<400> 3
atgggctgta atagcgactc caaccacaac aactccgacg gcaacatcac caaaaacaaa 60
acgatcgaag tttatgtcga tcgtgcaacc ctgccgacga ttcagcaaat gacccagatc 120
atcaacgaaa atagcaacaa caaaaaactg atttcatggt cgcgctaccc gatcaatgat 180
gaagaactgc tggaatcaat taacggctcg tttttcaaaa acaactctga actgatcaaa 240
agtctggatt ccatgattct gaccaatgac attaagaaag tgatcatcaa cggtaacacg 300
ctgtgggcgg ccgatgtggt taacatcatc aaatcaatcg aagcgttcgg caagaaaacc 360
gaaatcgaac tgaactttta tgatgacggt tcggccgaat atgtgcgtct gtacgacttt 420
agcaaactgc cggaatctga acaggaatac aaaattagcc tgtctaaaga taacattctg 480
agcagcatca acggcaccca gccgttcgaa aacgtcgtgg aaaacatcta cggtttcagt 540
caactgtacc cgaccacgta ccacatgctg cgtgccgata tctttgaaac caatctgccg 600
ctgcgcagtc tgaaaggcgt tctgtccaac aacatcaaac agatgaaatg ggattacttc 660
aaaaccttca acagccagca aaaagacaaa ttctacaact tcacgggttt taacccggat 720
gaaattatgg aacaatacaa agcaagcccg aacaaaaatt ttatcttcgt cggcaccaat 780
tctggcaccg caacggctga acagcaaatt gatatcctga ccgaagctaa aaacccgaac 840
agcccgatta tcacgaaatc gatccagggc ttcgacctgt ttttcaaagg tcatccgtct 900
gcaacctaca acaaacaaat catcgatgct cacaacatga tcgaaatcta caacaaaatc 960
ccgttcgaag cgctgatcat gaccgatgcc ctgccggatg cggtgggcgg tatgggcagc 1020
agcgtgtttt tcagcctgcc gaataccgtg gaaaacaaat tcattttcta taaatccgat 1080
acggacattg aaaacaatgc cctgatccag gttatgattg aactgaatat cgtgaaccgt 1140
aatgatgtga aactgatctc ggacctgcaa taa 1173
<210> 4
<211> 1167
<212> DNA
<213> Pasteurella multocida
<400> 4
atgaaaacga ttaccctgta tctggacccg gcgtccctgc cggcactgaa ccaactgatg 60
gattttacgc agaacaatga agacaaaacc catccgcgta tctttggcct gtctcgcttc 120
aaaattccgg ataacattat cacccaatat cagaatatcc actttgttga actgaaagac 180
aatcgtccga cggaagccct gttcaccatt ctggatcagt acccgggtaa cattgaactg 240
gacatccatc tgaatattgc tcacagcgtc cagctgattc gtccgatcct ggcgtatcgc 300
tttaaacatc tggatcgtgt gtccatccag cgcctgaacc tgtatgatga cggctcaatg 360
gaatacgttg atctggaaaa agaagaaaac aaagacatct cggcagaaat taaacaagct 420
gaaaaacagc tgagccatta tctgctgacg ggtaaaatca aattcgataa cccgaccatt 480
gcgcgctacg tttggcagtc tgcctttccg gtcaaatatc acttcctgag tacggactac 540
tttgaaaaag cagaatttct gcaaccgctg aaagaatatc tggcggaaaa ttaccagaaa 600
atggattgga cggcctatca gcaactgacc ccggaacagc aagcatttta cctgaccctg 660
gttggcttca acgacgaagt caaacagagt ctggaagtgc agcaagcgaa atttattttc 720
acgggcacca cgacctggga aggtaatacc gatgttcgtg aatattacgc ccagcaacag 780
ctgaacctgc tgaatcattt tacccaggcg ggcggcgacc tgtttattgg tgaccattac 840
aaaatttact tcaaaggtca cccgcgcggc ggtgaaatca acgattacat cctgaacaac 900
gcaaaaaaca tcacgaatat cccggctaat atctctttcg aagtgctgat gatgaccggc 960
ctgctgccgg ataaagtcgg cggtgtggct agctctctgt acttcagtct gccgaaagaa 1020
aaaattagtc acatcatctt caccagcaac aaacaggtca aatcaaaaga agatgccctg 1080
aacaatccgt acgtgaaagt tatgcgtcgc ctgggtatta tcgatgaatc gcaagtgatc 1140
ttttgggaca gcctgaaaca gctgtaa 1167
<210> 5
<211> 1116
<212> DNA
<213> Neisseria meningitidis
<400> 5
atgggcctga aaaaagcctg cctgaccgtg ctgtgtctga tcgtgttttg cttcggcatc 60
ttttatacgt tcgatcgtgt gaaccagggt gaacgcaatg cagttagtct gctgaaagaa 120
aaactgttta acgaagaagg cgaaccggtg aatctgatct tctgttacac cattctgcaa 180
atgaaagttg ccgaacgtat tatggcacag catccgggtg aacgctttta tgtggttctg 240
atgagcgaaa accgtaacga aaaatacgat tactacttca accagatcaa agataaagcg 300
gaacgcgcct atttctttca cctgccgtac ggcctgaaca aaagttttaa tttcattccg 360
acgatggcgg aactgaaagt gaaaagcatg ctgctgccga aagttaaacg tatctatctg 420
gcaagcctgg aaaaagtgtc tattgcggcc tttctgagca cctacccgga tgcggaaatc 480
aaaaccttcg atgatggcac gggtaatctg attcagagct ctagttatct gggcgatgaa 540
ttttctgtta acggtacgat caaacgtaat ttcgcccgca tgatgatcgg tgattggtct 600
attgcgaaaa cccgcaacgc cagtgatgaa cattacacga tcttcaaagg cctgaaaaac 660
atcatggatg atggtcgtcg caaaatgacc tacctgccgc tgttcgatgc gtctgaactg 720
aaaacgggcg atgaaaccgg cggtacggtg cgtattctgc tgggtagccc ggataaagaa 780
atgaaagaaa tctctgaaaa agcagcgaaa aacttcaaaa tccagtatgt tgccccgcac 840
ccgcgtcaga cctacggcct gagtggtgtg accacgctga acagcccgta tgttattgaa 900
gattacatcc tgcgtgaaat taagaaaaac ccgcataccc gctatgaaat ctacacgttt 960
ttcagcggcg ccgcactgac catgaaagat tttccgaacg tgcacgttta tgcactgaaa 1020
ccggcgtctc tgccggaaga ttattggctg aaaccggtgt acgcgctgtt tacccagagt 1080
ggtattccga tcctgacgtt cgatgataaa aattaa 1116
<210> 6
<211> 852
<212> DNA
<213> Pasteurella multocida
<400> 6
atggataaat ttgcagaaca tgaaattccg aaagcagtga tcgttgctgg caacggtgaa 60
agtctgtccc agattgatta tcgtctgctg ccgaaaaact acgacgtctt ccgttgcaac 120
caattctact tcgaagaacg ctacttcctg ggcaataaaa tcaaagccgt gtttttcacc 180
ccgggtgttt ttctggaaca gtattacacg ctgtatcatc tgaaacgcaa caatgaatac 240
tttgtcgata acgtgattct gagctctttc aatcacccga ccgtggacct ggaaaaatca 300
cagaaaatcc aagcactgtt catcgatgtt atcaacggct acgaaaaata cctgtcgaaa 360
ctgaccgctt tcgatgttta tctgcgttac aaagaactgt atgaaaatca gcgcattacg 420
agcggtgttt acatgtgcgc tgtcgcgatc gccatgggct ataccgatat ttacctgacg 480
ggtatcgact tttatcaagc gtctgaagaa aactacgcct tcgataacaa aaaaccgaat 540
attatccgtc tgctgccgga ctttcgcaaa gaaaaaaccc tgttcagcta tcattctaaa 600
gatattgacc tggaagcgct gtcatttctg cagcaacatt accacgtgaa cttctactca 660
atctcgccga tgagtccgct gtccaaacat tttccgatcc cgacggttga agatgactgt 720
gaaaccacgt tcgtcgcccc gctgaaagaa aactatatta atgacatcct gctgccgccg 780
cactttgtct atgaaaaact gggcgtggat aaactggcgg ccgcactgga acatcaccat 840
caccatcact aa 852
<210> 7
<211> 1158
<212> DNA
<213> Pasteurella dagmatis
<400> 7
atgaccattt acctggaccc ggcgtctctg ccgaccctga accaactgat gcattttacg 60
aaagaaagcg aagacaaaga aaccgcacgt atttttggct tctctcgctt taaactgccg 120
gaaaaaatca cggaacagta caacaacatc catttcgtgg aaatcaaaaa caatcgtccg 180
acggaagata ttttcaccat cctggaccag tacccggaaa aactggaact ggatctgcat 240
ctgaacattg cacacagcat ccagctgttt catccgattc tgcaatatcg tttcaaacac 300
ccggatcgca ttagtatcaa atccctgaac ctgtatgatg acggcaccat ggaatacgtt 360
gatctggaaa aagaagaaaa caaagacatc aaaagtgcga tcaaaaaagc cgaaaaacag 420
ctgtccgatt atctgctgac gggtaaaatt aactttgaca atccgaccct ggcacgctac 480
gtttggcagt cacaatatcc ggtcaaatac catttcctgt cgacggaata ttttgaaaaa 540
gctgaattcc tgcagccgct gaaaacctat ctggcgggca aataccaaaa aatggattgg 600
tcagcctatg aaaaactgtc gccggaacag caaacgtttt acctgaaact ggtcggtttc 660
agtgatgaaa ccaaacagct gtttcacacg gaacaaacca aatttatttt cacgggcacc 720
acgacctggg agggtaacac cgatatccgt gaatattacg cgaaacagca actgaatctg 780
ctgaaacatt ttacccacag cgaaggcgac ctgtttatcg gtgaccagta caaaatctac 840
ttcaaaggcc atccgcgcgg cggtgatatt aacgactata tcctgaaaca cgcaaaagat 900
attacgaaca tcccggctaa tattagcttc gaaatcctga tgatgaccgg tctgctgccg 960
gacaaagtcg gcggtgtggc gagctctctg tacttctctc tgccgaaaga aaaaatcagc 1020
cacattatct tcacctctaa caagaaaatt aaaaacaaag aagatgccct gaatgacccg 1080
tacgtgcgtg ttatgctgcg tctgggtatg attgacaaaa gccaaattat cttctgggat 1140
tctctgaaac aactgtaa 1158
<210> 8
<211> 1173
<212> DNA
<213> Photobacterium phosphoreum
<400> 8
atgggctgta actccgatag caaacacaat aacagtgatg gcaatattac caaaaacaaa 60
acgatcgaag tctatgtgga ccgtgcgacc ctgccgacga ttcagcaaat gacccagatc 120
atcaacgaaa atagcaacaa caaaaaactg atttcatggt cgcgttaccc gatcaatgat 180
gaaacgctgc tggaatcaat taatggctcg tttttcaaaa accgcccgga actgatcaaa 240
agtctggatt ccatgattct gaccaacgaa attaagaaag tgatcatcaa cggtaacacg 300
ctgtgggcag ttgacgtggt taatattatc aaaagcattg aagctctggg caagaaaacc 360
gaaatcgaac tgaacttcta tgatgacggt tctgcggaat atgtgcgtct gtacgatttt 420
agccgcctgc cggaatctga acaggaatac aaaattagcc tgtctaaaga taacattcag 480
agcagcatca acggcaccca accgttcgac aacagcatcg aaaacatcta cggtttctct 540
cagctgtatc cgaccacgta ccacatgctg cgtgccgata tctttgaaac caatctgccg 600
ctgacgagtc tgaaacgcgt tatctccaac aacatcaaac agatgaaatg ggattacttc 660
accacgttca attcccagca gaaaaacaaa ttttacaact tcaccggctt caacccggaa 720
aaaatcaaag aacaatacaa agcgagtccg cacgaaaatt ttattttcat tggcaccaac 780
tccggcaccg ccaccgcaga acagcaaatt gatatcctga ccgaagccaa aaaaccggac 840
tcaccgatta tcaccaacag cattcagggc ctggacctgt ttttcaaagg tcatccgtct 900
gcgacctata accagcaaat tatcgacgcc cacaacatga tcgaaatcta caacaaaatc 960
ccgttcgaag cactgatcat gaccgatgca ctgccggacg ctgttggcgg tatgggtagt 1020
tccgtctttt tctcactgcc gaataccgtc gaaaacaaat tcattttcta taaatcggat 1080
acggacattg aaaacaatgc tctgatccag gttatgatcg aactgaatat cgtgaaccgc 1140
aatgatgtga aactgattag tgacctgcaa taa 1173
<210> 9
<211> 1254
<212> DNA
<213> Avibacterium paragallinarum
<400> 9
atgcgtaaaa tcatcacctt cttcagcctg ttcttctcga tctcagcgtg gtgtcaaaaa 60
atggaaatct acctggacta tgcgtcgctg ccgagcctga acatgatcct gaacctggtt 120
gaaaacaaaa acaacgaaaa agtcgaacgt attatcggct tcgaacgctt tgatttcaac 180
aaagaaattc tgaatagctt ctctaaagaa cgtatcgaat ttagtaaagt ctccattctg 240
gatatcaaag aattttcaga caaactgtac ctgaacattg aaaaatcgga tacgccggtg 300
gacctgatta tccataccaa tctggatcac tcagttcgtt cgctgctgag catctttaaa 360
accctgagtc cgctgttcca taaaatcaac atcgaaaaac tgtacctgta cgatgacggc 420
agcggtaact atgttgatct gtaccagcac cgccaagaaa atatttctgc gattctgatc 480
gaagcccaga aaaaactgaa agacgcgctg gaaaatcgtg aaacggatac cgacaaactg 540
catagcctga cgcgctatac ctggcacaaa atctttccga cggaatatat cctgctgcgt 600
ccggattacc tggatattga cgaaaaaatg caaccgctga aacatttcct gagcgatacc 660
atcgtgtcta tggacctgtc tcgctttagt catttctcca aaaaccagaa agaactgttt 720
ctgaaaatca cgcacttcga tcaaaacatc ttcaacgaac tgaacatcgg caccaaaaac 780
aaagaataca aaacgttcat cttcaccggc accacgacct gggaaaaaga taagaaaaaa 840
cgtctgaaca acgcgaaact gcagacggaa attctggaat cttttatcaa accgaacggc 900
aaattctacc tgggtaacga tatcaaaatc tttttcaaag gccacccgaa aggtgatgac 960
attaacgact acattatccg caaaaccggc gcagaaaaaa ttccggctaa catcccgttt 1020
gaagttctga tgatgacgaa tagtctgccg gattatgtcg gcggtattat gagtaccgtg 1080
tacttttccc tgccgccgaa aaatattgat aaagtggttt tcctgggttc cgaaaaaatc 1140
aaaaacgaaa acgacgccaa atcacagacc ctgtcgaaac tgatgctgat gctgaacgtc 1200
atcacgccgg aacagatttt ctttgaagaa atgccgaacc cgattaactt ttaa 1254
<210> 10
<211> 1293
<212> DNA
<213> Campylobacter jejuni
<400> 10
atgacccgca cccgtatgga aaacgaactg attgtgagca aaaacatgca gaacattatt 60
atcgccggta acggtccgag cctgaaaaat attaactata aacgtctgcc gcgcgaatac 120
gatgtgttcc gttgcaacca gttctacttc gaagacaaat actacctggg caagaaaatt 180
aaagccgtgt ttttcaatcc gggcgtgttt ctgcaacaat atcataccgc aaaacagctg 240
attctgaaaa acgaatacga aatcaaaaac atcttttgta gcaccttcaa tctgccgttt 300
atcgaatcta acgatttcct gcaccaattt tataactttt tcccggacgc taaactgggc 360
tacgaagtca tcgaaaacct gaaagaattt tacgcgtaca tcaaatacaa cgaaatctac 420
ttcaacaaac gcatcacctc tggcgtgtat atgtgcgcga ttgccatcgc actgggttat 480
aaaacgattt acctgtgtgg catcgatttc tatgaaggtg acgttattta cccgtttgaa 540
gcaatgagta ccaacattaa aacgatcttc ccgggtatca aagatttcaa accgagtaac 600
tgccattcca aagaatatga catcgaagcg ctgaaactgc tgaaaagcat ctacaaagtt 660
aacatctacg ccctgtgtga tgacagtatt ctggcaaatc atttcccgct gtccattaac 720
atcaacaaca acttcaccct ggaaaacaaa cacaacaact caatcaacga tattctgctg 780
accgacaata cgccgggcgt ctcgttttat aaaaatcagc tgaaagccga taacaaaatc 840
atgctgaact tctacaacat cctgcatagc aaagataacc tgatcaaatt cctgaacaaa 900
gaaatcgctg ttctgaaaaa acagaccacg caacgtgcta aagcgcgcat tcagaaccac 960
ctgagctata aactgggcca agccctgatt atcaatagca aatctgtcct gggtttcctg 1020
tctctgccgt ttattatcct gtcaattgtg atctcgcaca aacaggaaca aaaagcgtat 1080
aaattcaaag tgaagaaaaa cccgaacctg gcactgccgc cgctggaaac ctatccggat 1140
tacaacgaag ccctgaaaga aaaagaatgc ttcacgtaca aactgggcga agaatttatc 1200
aaagcaggta aaaactggta tggcgaaggt tacatcaaat ttatcttcaa agatgttccg 1260
cgtctgaaac gtgaatttga aaaaggcgaa taa 1293
<210> 11
<211> 1188
<212> DNA
<213> Heliobacter acinonychis
<400> 11
atgaataaga aaccgctgat tattgctggc aacgggccaa gcatcaaaga cttagattat 60
gcgttgttcc cgaaagactt tgatgtattc cgatgtaatc aattctactt cgaggacaaa 120
tactatttag ggcgggaaat aaaaggggtg ttctttaacg cgcacgtctt cgatctccaa 180
atgaagatca ctaaagccat agtcaaaaac ggggaatatc acccggacca catatattgc 240
acacatgtcg aaccgtacgg ttacgttaac ggaaaccagc aactcatgca agagtacctg 300
gaaaaacatt ttgtgggagt ccgaagcacg tacgcatacc tgaaagatct agagccattc 360
tttattctgc acagtaagta tcgcaacttc tacgaccagc acttcacaac gggcatcatg 420
atgctactgg tggccatcca attgggatac aaagaaatat acctgtgcgg aatagacttc 480
tacgaaaacg gattcggaca tttctacgag aaccaagggg gattctttga agaggatagc 540
gatccgatgc acgataagaa catagacatc caagcactgg aactggcaaa gaaatacgcg 600
aaaatctacg cactggtacc gaacagcgcc ctagtgaaaa tgattccgtt gagcagccaa 660
aaaggagttc tggaaaaggt gaaggaccgg atcgggttgg gcgagtttaa gagagagaaa 720
ttcgggcaaa aagaattgga aagacagaag gaattagaac gacaaaaaga gctcgaacgc 780
caaaaggagc ttgaacgtca aaaggaactt gaacgacaaa aagagttgga gaggcagaaa 840
gaactcgaac gccaaaaaga attagagaga cagaaggaat tagagcgcca aaaggagctt 900
gagcgtcaaa aagaattaga gaggcagaag gagttagaaa ggcagaaaga actggagaga 960
cagaaagaac tcgaaaggca gaaggagttg gaacgccaaa aagaactaga attagaacga 1020
tccttaaaag cacgattgaa agcggtactc gcgagcaaag gcatccgcgg cgacaacctg 1080
ataatcgtaa gtttaaaaga cacctaccga ctgtttaaag ggggatttgc gttactcttg 1140
gacctgaagg cgctaaagtc aatcattaaa gcattcctga agagataa 1188
<210> 12
<211> 783
<212> DNA
<213> Campylobacter jejuni
<400> 12
atgggcaaaa aagtgattat tgcgggcaac ggcccgagcc tgaaagaaat tgattatagc 60
cgtctgccga acgattttga tgtgtttcgc tgcaaccagt tttatttcga agataaatat 120
tacctgggca aaaaatgcaa agcggtgttc tataatccga tcctgttctt cgaacagtat 180
tacaccctga aacatctgat tcagaaccag gaatatgaaa ccgaactgat catgtgcagc 240
aactataacc aggcgcatct ggaaaacgaa aactttgtga aaaccttcta cgattatttt 300
ccggatgcgc atctgggcta tgattttttc aaacagctga aagatttcaa cgcgtacttc 360
aaattccacg aaatctattt caaccagcgt attaccagcg gcgtgtatat gtgcgcggtg 420
gcgattgcgc tgggctataa agaaatttat ctgagcggca tcgattttta tcagaacggc 480
agcagctatg cgtttgatac caaacagaaa aacctgctga aactggcccc gaactttaaa 540
aacgataaca gccactatat tggccatagc aaaaacaccg atatcaaagc gctggaattt 600
ctggaaaaaa cctataaaat caaactgtat tgcctgtgcc cgaacagcct gctggccaac 660
tttattgaac tggcaccgaa tctgaacagc aacttcatca tccaggaaaa aaacaactat 720
accaaagata ttctgattcc gagcagcgaa gcgtatggca aattcagcaa aaacatcaac 780
taa 783
<210> 13
<211> 897
<212> DNA
<213> Streptococcus entericus
<400> 13
atgaagaaag tctacttctg ccatacggtc taccatctgc tgattaccct gtgcaaaatt 60
agcgttgaag aacaagttga aattattgtg ttcgataccg ttagtaatca tgaactgatt 120
gtccagaaaa tccgcgacgt gtttgttaac accacggtgc tgttcgcaga acaaaatacc 180
gatttttcca ttctggaaat cgatcgcgct acggacattt atgtgttcaa cgactggacc 240
ccgatcggcg cgtatctgcg taaaaacaaa ctgttttacc atctgatcga agatggttat 300
aactaccacg aatataacgt ttacgcgaat gccctgacca tgaaacgtcg cctgctgaac 360
ttcgtgctgc gtcgcgaaga accgtcaggc ttttcgcgtt atgttcgcag cattgaagtt 420
aaccgtgtca aatacctgcc gaatgattgc cgcaaaagca aatgggttga aaaaccgcgt 480
tctgccctgt tcgaaaatct ggtcccggaa cataaacaga aaatcatcac gatcttcggc 540
ctggaaaact atcaagatag cctgcgcggt gtcctggtgc tgacccagcc gctggtgcaa 600
gactactggg atcgcgacat taccacggaa gaagaacagc tggaatttta tcgtcaaatc 660
gtggaatctt acggcgaagg tgaacaggtg tttttcaaaa ttcacccgcg tgataaagtt 720
gactatagct ctctgaccaa cgtcattttt ctgaagaaaa acgtcccgat ggaagtgtac 780
gaactgattg ccgattgtca ttttaccaaa ggtatcacgc acagttccac cgcactggac 840
ttcctgtcct gtgtggataa gaaaatcacc ctgaaacaaa tgaaagcaaa tagttaa 897
<210> 14
<211> 888
<212> DNA
<213> Haemophilus ducreyi
<400> 14
atgaaagaaa tcgccatcat ctccaaccaa cgcatgttct tcctgtactg tctgctgacc 60
aataaaaatg tcgaagacgt gttcttcatt tttgaaaaag gcgcgatgcc gaacaatctg 120
accagcattt ctcatttcat cgtgctggat cacagtaaat ccgaatgcta tgactttttc 180
tacttcaact tcatcagttg taaatatcgt ctgcgcggcc tggatgttta cggtgcagac 240
catatcaaag gcgctaaatt tttcctggaa cgtcaccgct ttttcgtggt tgaagatggt 300
atgatgaact acagcaaaaa catgtacgca ttctctctgt tccgtacccg caatccggtg 360
attctgccgg gcggttttca tccgaacgtt aaaaccatct tcctgacgaa agataatccg 420
attccggacc agatcgctca caaacgtgaa atcatcaaca tcaaaaccct gtggcaagcg 480
aaaaccgcca cggaaaaaac gaaaattctg agctttttcg aaatcgatat gcaggaaatt 540
tcagttatca aaaaccgctc gtttgtcctg tatacccaac cgctgtcaga agataaactg 600
ctgacggaag cggaaaaaat tgacatctat cgtaccattc tgacgaaata caaccattcg 660
cagaccgtta tcaaaccgca cccgcgcgat aaaacggact ataaacaact gtttccggat 720
gcctatgtca tgaaaggcac ctacccgagt gaactgctga cgctgctggg tgtcaacttc 780
aacaaagtga tcaccctgtt ttccacggcg gtcttcgatt atccgaaaga aaaaatcgac 840
ttctacggca ccgcggtgca tccgaaactg ctggatttct ttgactaa 888
<210> 15
<211> 1467
<212> DNA
<213> Alistipes sp.
<400> 15
atggccctgc tgagcggtac cgccgcatgc tcagatgacg aagtctcgca gaacctgatc 60
gtgattaatg gcggtgaaca ttttctgagc ctggatggtc tggcccgtgc aggtaaaatt 120
agcgtgctgg caccggctcc gtggcgtgtt acgaaagcag ctggtgatac ctggtttcgc 180
ctgagcgcaa ccgaaggtcc ggctggttac agcgaagtgg aactgtctct ggatgaaaat 240
ccgggtgccg cacgtagcgc acagctggcg tttgcctgtg gtgatgcgat tgtgccgttc 300
cgcctgagtc aaggcgcact gtccgctggt tatgattcac cggactatta cttttacgtt 360
accttcggca cgatgccgac cctgtatgcc ggtatccatc tgctgagcca cgataaaccg 420
ggctatgtct tttactcacg ttcgaaaacg tttgacccgg ccgaattccc ggcacgtgct 480
gaagttacca ccgcagctga tcgtaccgcc gatgcaaccc aggccgaaat ggaagcaatg 540
gctcgcgaaa tgaaacgtcg catcctggaa attaactctg cggatccgac cgccgtgttt 600
ggcctgtatg ttgatgacct gcgttgccgc attggctacg attggttcgt ggcgcagggt 660
atcgacagtg cccgtgtcaa agtgagcatg ctgtctgatg gcaccggcac gtacaacaat 720
ttttataact acttcggtga cgcggccacg gcggaacaaa attgggaaag ttatgcgtcc 780
gaagttgaag ccctggattg gaatcacggc ggtcgttatc cggaaacccg ctcgctgccg 840
gaatttgaaa gctacacgtg gccgtattac ctgtctaccc gtccggatta tcgcctggtg 900
gttcaggacg gcagtctgct ggaaagctct tgtccgttta ttaccgaaaa actgggtgaa 960
atggaaatcg aatccattca accgtatgaa atgctgtcag ccctgccgga aagttcccgt 1020
aaacgctttt atgatatggc aggcttcgat tacgacaaat ttgcagctct gttcgatgcg 1080
tccccgaaga aaaacctgat tatcattggt acctctcatg cggatgatgc cagtgcacgt 1140
ctgcagcgtg attacgttgc acgcatcatg gaacagtatg gcgctcaata cgatgtcttt 1200
ttcaaaccgc acccggcaga caccacgtca gctggttatg aaacggaatt tccgggcctg 1260
accctgctgc cgggtcaaat gccgtttgaa atcttcgttt ggtccctgat tgatcgtgtc 1320
gacatgatcg gcggttatcc gtcaacggtc tttctgaccg ttccggtcga taaagtgcgc 1380
tttatttttg ccgcggatgc agcttctctg gtgcgtccgc tgaatatcct gttccgcgat 1440
gcgaccgacg ttgaatggat gcagtaa 1467
<210> 16
<211> 876
<212> DNA
<213> Campylobacter jejuni
<400> 16
atgaagaaag tgattatcgc cggcaatggt ccgagcctga aagaaattga ttattctcgt 60
ctgccgaatg atttcgacgt ctttcgctgc aaccagttct actttgaaga caaatattac 120
ctgggcaaaa aatgtaaagc cgtgttttat accccgaact ttttctttga acagtattac 180
acgctgaaac atctgattca gaaccaagaa tatgaaaccg aactgatcat gtgctcaaac 240
tacaatcaag cacatctgga aaacgaaaac ttcgtcaaaa cgttctacga ttacttcccg 300
gacgctcacc tgggttacga tttctttaaa cagctgaaag aattcaacgc gtacttcaaa 360
ttccacgaaa tctacttcaa ccaacgtatc acctcaggcg tgtatatgtg tgcggttgcc 420
attgcactgg gttataaaga aatttacctg tcgggcatcg atttttatca gaatggtagc 480
tcttacgcct tcgacacgaa acaagaaaat ctgctgaaac tggcaccgga ttttaaaaac 540
gaccgctcac attatattgg ccactcgaaa aacaccgata tcaaagctct ggaattcctg 600
gaaaaaacgt acaaaatcaa actgtactgc ctgtgtccga atagtctgct ggctaacttt 660
atcgaactgg cgccgaacct gaattccaac ttcatcatcc aggagaaaaa caactacacc 720
aaagatatcc tgatcccgag ttccgaagcg tacggcaaat ttagcaaaaa catcaacttc 780
aagaaaatta aaatcaaaga aaacgtgtat tacaaactga ttaaagatct gctgcgtctg 840
ccgtctgaca tcaaacatta ttttaaaggt aaataa 876
<210> 17
<211> 939
<212> DNA
<213> Streptococcus agalactiae
<400> 17
atgacgaatc gcaaaatcta tgtctgccac accctgtacc atctgctgat ctgcctgtat 60
aaagaagaaa tctactcaaa tctggaaatt atcctgagca gcagcattcc ggatgtggac 120
aacctggaga aaaaactgaa aagcaaaacc atcaacatcc atattctgga agaatcctca 180
ggcgaatctg aagaactgct gagtgttctg aaagatgcag gtctgtctta cagtaaattc 240
gatagcaact gcttcatctt caacgacgct accccgattg gccgtacgct gatcaaacac 300
ggtatttatt acaatctgat cgaagatggc ctgaactgtt ttacctactc gattttcagc 360
cagaaactgt ggaaatacta cgtgaaaaaa tacatcctgc ataaaattca accgcacggc 420
ttttcccgct actgcctggg tatcgaagtg aacagtctgg ttaatctgcc gaaagatccg 480
cgttacaaaa aattcatcga agtcccgcgc aaagaactgt tcgacaatgt tacggaatac 540
cagaaagaaa tggcgatcaa cctgtttggc gccgtccgtg tgtctattaa atccccgtca 600
gttctggtcc tgacccagcc gctgtccatc gataaagaat ttatgtcata caacaacaaa 660
atcgaaacgt cggaagaaca attcaacttc tacaaaagca tcgtgaacga atacatcaac 720
aaaggttaca acgtctacct gaaagtgcat ccgcgtgatg tggttgacta ttctaaactg 780
ccggttgaac tgctgccgag taacgtcccg atggaaatta tcgaactgat gctgaccggc 840
cgctttgaat gcggtattac ccatagcagc accgccctgg atttcctgac ctgtgtggac 900
aagaaaatta cgctggttga tctgaaagac attaaataa 939
<210> 18
<211> 1233
<212> DNA
<213> Bibersteinia trehalosi
<400> 18
atggaattct gcaaaatggc aacgacgcaa aaaatctgtg tctacctgga ctatgctacg 60
atcccgagcc tgaactacat cctgcacttt gcgcaacatt tcgaagatca ggaaaccatt 120
cgtctgtttg gcctgtcccg cttccacatt ccggaatcag tcatccagcg ctatccgaaa 180
ggtgtggttc aattttaccc gaaccaggaa aaagacttca gcgcgctgct gctggccctg 240
aaaaacatcc tgatcgaagt taaacagcaa cagcgtaaat gcgaaatcga actgcatctg 300
aacctgtttc actatcagct gctgctgctg ccgttcctga gtctgtatct ggatacccag 360
gactactgtc atctgacgct gaaattttac gatgacggct ctgaagcgat tagtgccctg 420
caggaactgg cactggctcc ggatctggcg gcccaaatcc agtttgaaaa acaacagttc 480
gacgaactgg tcgtgaaaaa atcgtttaaa ctgtcgctgc tgagccgcta tttttggggt 540
aaactgttcg aaagcgaata catttggttc aatcaagcaa tcctgcagaa agctgaactg 600
caaattctga aacaggaaat cagctctagt cgtcagatgg attttgcaat ttatcaacag 660
atgtccgacg aacaaaaaca gctggtgctg gaaattctga acatcgatct gaataaagtt 720
gcttacctga aacaactgat ggaaaaccag ccgtcttttc tgttcctggg caccacgctg 780
tttaatatta cccaggaaac caaaacgtgg ctgatgcaga tgcatgtgga tctgatccaa 840
cagtattgcc tgccgagcgg ccagtttttc aacaataaag ccggctatct gtgtttttac 900
aaaggtcacc cgaacgaaaa agaaatgaac caaatgatcc tgtctcagtt caaaaacctg 960
atcgcgctgc cggatgacat tccgctggaa atcctgctgc tgctgggcgt tattccgagt 1020
aaagtcggcg gttttgcatc ctcagctctg tttaacttca ccccggcgca gatcgaaaat 1080
attatctttt tcacgccgcg ttatttcgaa aaagataatc gcctgcacgc cacgcaatac 1140
cgtctgatgc agggcctgat tgaactgggt tatctggacg ctgaaaaatc tgtgacccac 1200
tttgaaatca tgcaactgct gacgaaagaa taa 1233
<210> 19
<211> 1221
<212> DNA
<213> Haemophilus parahaemolyticus
<400> 19
atgaccgaac agtacatcaa aaacgtggaa gtttacctgg attacgcgac catcccgacg 60
ctgaactact tctaccattt caccgaaaac aaagatgaca tcgccacgat tcgtctgttt 120
ggcctgggtc gcttcaacat cagtaaatcc atcatcgaaa gctacccgga aggcattatc 180
cgttactgcc cgattatctt tgaagatcaa accgcatttc agcaactgtt cattaccctg 240
ctgacggaag acagtttttg tcagtatcgc tttaacttcc atattaacct gtttcactcc 300
tggaaaatgc tgatcccgct gctgcatatt atctggcagt ttaaacacaa agtcctggat 360
attaaactga acttctatga tgacggcagt gaaggtctgg tgacgctgtc caaaatcgaa 420
cagaactaca gctctgaaat cctgcaaaaa atcatcgata tcgactcaca gtcgttttat 480
gcagataaac tgtctttcct ggatgaagac attgctcgtt acctgtggaa cagtctgttt 540
gaatcccatt attacctgct gaacgacttc ctgctgaaaa acgaaaaact gtcactgctg 600
aaaaactcga tcaaatactg ccacatcatg gatctggaac gctacctgca gtttacccaa 660
gaagaaaaag actttttcaa cgaactgctg ggcatcaaca tccagagtct ggaagataaa 720
atcaaaatct tccagcagaa gaaaaccttt attttcacgg gtaccacgat cttcagcctg 780
ccgaaagaag aagaagaaac cctgtatcgt ctgcatctga acgcaatcct gaattatatt 840
cacccgaacg gcaaatactt tattggcgat ggtttcacgc tggttatcaa aggtcatccg 900
caccagaaag aaatgaacag ccgcctggaa aaatcttttg aaaaagctgt catgctgccg 960
gataatatcc cgttcgaaat tctgtatctg atcggctgca aaccggacaa aattggcggt 1020
tttgtgagca cctcttactt cagctgtgat aagaaaaaca ttgcggacct gctgtttatc 1080
tctgcccgtc aagaagaagt tcgcaaaaac gattacctgt ttaacatcca gtaccaactg 1140
cgtgacatga tgattaaaac cggttttatc caggaagaaa aaacgcactt ctactcagat 1200
atcccgatct tcatctcgta a 1221
<210> 20
<211> 903
<212> DNA
<213> Haemophilus somnus
<400> 20
atgaaatata acatcaaaat taaagctatc gtcatcgtgt cgagcctgcg tatgctgctg 60
atcttcctga tgctgaataa ataccacctg gatgaagttc tgtttgtctt caacgaaggc 120
ttcgaactgc ataaaaaata caaaatcaaa cactatgtgg cgattaaaaa gaaaattacc 180
aaattctggc gtctgtacta caaactgtac ttctaccgtt tcaaaattga ccgcatcccg 240
gtttatggcg cagatcatct gggttggacc gactattttc tgaaatactt cgatttctac 300
ctgattgaag acggcatcgc taacttctcc ccgaaacgtt acgaaattaa cctgacgcgc 360
aatatcccgg tctttggttt ccataaaacc gtgaagaaaa tttacctgac gagtctggaa 420
aatgttccgt ccgatattcg tcataaagtc gaactgatca gcctggaaca cctgtggaaa 480
acccgcacgg cgcaggaaca acacaacatc ctggatttct ttgcctttaa tctggacagc 540
ctgatctctc tgaaaatgaa aaaatacatc ctgttcaccc agtgcctgtc agaagatcgc 600
gtcatttcgg aacaggaaaa aatcgcgatc taccaacata tcatcaaaaa ctacgatgaa 660
cgtctgctgg ttatcaaacc gcacccgcgc gaaaccacgg actatcagaa atactttgaa 720
aatgtcttcg tgtaccaaga tgtggttccg agcgaactgt ttgaactgct ggacgtgaac 780
ttcgaacgtg ttattaccct gttttctacg gccgtgttca aatatgatcg caatatcgtt 840
gacttctacg gtacgcgcat ccacgacaaa atctatcaat ggttcggcga catcaaattc 900
taa 903
<210> 21
<211> 1146
<212> DNA
<213> Vibrio harveyi
<400> 21
atggattctt cgccggaaaa caccagctct acgctggaaa tttacatcga ttcagcaacc 60
ctgccgtcgc tgcagcacat ggtgaaaatt atcgacgaac aaagtggcaa caaaaaactg 120
atcaactgga aacgttatcc gatcgatgac gaactgctgc tggataaaat caacgctctg 180
agcttttctg ataccacgga cctgacccgt tatatggaaa gtattctgct gatcggcgat 240
attaaacgcg tggttattaa cggtaatagt ctgtccaact acaatattgt cggcgtgatg 300
cgctccatca acgccctggg tctggatctg gacgttgaaa tcaattttta tgatgacggt 360
tcagcagaat atgtccgtct gtacaacttc tcgcagctgc cggaagctga acgcgaactg 420
ctggtgtcaa tgtcgaaaaa caatattctg gcggccgtta acggcatcgg ttcttatgat 480
agcggctctc cggaaaatat ttacggtttt gcgcagattt atccggccac ctaccacatg 540
ctgcgtgcgg acattttcga tacggacctg gaaatcggcc tgattcgcga tatcctgggt 600
gacaacgtca aacagatgaa atggggccaa tttctgggtt tcaacgaaga acagaaagaa 660
ctgttttatc aactgaccag cttcaacccg gataaaatcc aggcgcaata caaagaatct 720
ccgaacaaaa acttcgtttt cgtcggcacc aacagtcgtt ccgcaacggc tgaacagcaa 780
atcaacatca tcaaagaagc caaaaaactg gatagcgaaa ttatcccgaa cagcatcgat 840
ggctatgacc tgtttttcaa aggtcatccg agcgcgacct acaaccagca aattgttgat 900
gcccacgaca tgaccgaaat ctataatcgc acgccgtttg aagtcctggc aatgacgagt 960
tccctgccgg atgctgtggg cggtatgggc tcatcgctgt ttttctcact gccgaaaacc 1020
gtggaaacga aattcatttt ctataaaagt ggcaccgata ttgaatccaa tgcgctgatc 1080
caggttatgc tgaaactggg tatcattacg gacgaaaaag tgcgctttac gacggacatc 1140
aaataa 1146
<210> 22
<211> 1452
<212> DNA
<213> Alistipes sp.
<400> 22
atggccagct gttctgatga cgataaagaa cagacgggtt ttcaaatcga cgatggctct 60
ggtttcctga gtctggatgc agctgcgcgt agtggctcca ttgccatcac cgcaaacaat 120
tcatggtcgg tgacgcagga taaagacagc gaatggctga ccctgagcac cacgtctggt 180
gcagcaggtc gtaccgaaat tggtatcatg ctggaagcga acccgggcga agctcgtaat 240
gcgggtctga cctttaactc tggcggtcgc acgtatccgt tcgtgattac ccagagtgcc 300
catgttacgg cagattttga cgatgctgac cactgctttt atatcacctt tggtaccctg 360
ccgaccctgt atgcaggtct gcatgtgctg tcccacgata aaccgtcata tgtgtttttc 420
cagcgttccc aaacctttcg cccggaagaa ttcccggccc atgcagaagt tacgattgct 480
gcggatccgt cagctaatgc gaccgatgaa gacatggaac gtatgcgcac ggccatgaaa 540
cagcaaattc tgaaaatcaa cgttgaagat ccgaccgcag tttttggcct gtatgtcgac 600
gatctgcgtt gtggcattgg ttacgattgg ttcgtcgccc agggtatcga cagtacccgc 660
gtgaaagtta gtatgctgtc cgatggcacc ggcacgtaca acaacttcta caactacttc 720
ggcgatccgg ccaccgcaga acaaaactgg gaaaattacg ccgcacaggt ggaagcgctg 780
gattggcaac acggcggtcg ttttccggaa acccgcatgc cggatggttt tgacttctat 840
gaatggccgt attacctggc aacgcgtccg aactaccgcc tggttctgca ggacgatgac 900
ctgctggaag cgacgtctcc gtttatgacc gaacgtctgc agcaaatgcg caccgaatcg 960
aaacagccgt atgaactgct ggccagcctg ccggctgaag cccgtcaacg ctttttccgt 1020
atggctggct ttgattacga cgcgtttgct gcgctgttcg atgccagccc gaagaaaaac 1080
ctggtcatta tcggcacgtc acatacctcg gaagaaagcg aagcacagca agccgcatat 1140
gtggaacgta ttatcggcga ttatggtacc gcctacgaca ttttctttaa accgcacccg 1200
gcagatagct ctagttccaa ctacgaagaa cgctttgaag gtctgaccct gctgccgggt 1260
cagatgccgt ttgaaatttt cgtctggtcg ctgctggata aagtggacct gatcggcggt 1320
tattcatcga cggtgtttct gaccgtcccg gtggaaaaaa ccggctttat tttcgctgcg 1380
aatgctgaaa gcctgccgcg cccgctgaac gttctgttcc gtaatgcgga acatgtccgc 1440
tggatccagt aa 1452
<210> 23
<211> 1452
<212> DNA
<213> Alistipes shahii
<400> 23
atggacgatg gcaccccgag tgtcagcatc aacggcggca ccgacttcct gagcctggac 60
cacctggcac gcagcggcaa aatcacggtc aacgcaccgg ctccgtggtc tgtgaccctg 120
gccccggaaa attacggcca ggatgaaaaa ccggactggc tgaccctgag cgccgaagaa 180
ggcccggcag gttatagcga aatcgatgtt acctttgcgg aaaacccggg tccggcccgt 240
tccgcatcac tgctgttcag ctgcgatggt aaaaccctgg cctttacggt ttcgcagagc 300
gcaggcggta cgggtttcga tgctccggac tattactttt atatttcggt cggcaccatg 360
ccgacgctgt actcgggtct gcatctgctg agccacgata aaccgtctta tgttagttac 420
gaacgtgcga gcacctttga tgcggccgaa ttcccggacc gcgcgtttgt ctatccggtg 480
gccgatccga ccggtcatgc aaccaacgaa gaactgcgtg cgatgagcga agccatgaaa 540
cgtcgcatcc tggaaattaa tgcagaagat ccgaccgctg ttttcggtct gtgggtcgat 600
gacctgcgtt gccgcctggg ctacgattgg tttgtggctc aaggtatcga ctctgcgcgc 660
gtgaaagtta cgatgctgag tgatggcacc gcgacgtata acaattttca taactacttc 720
ggtgacgcag ctaccgccga acagaactgg aatgattatg cggccgaagt tgaagcactg 780
gactggaatc atggcggtcg ttatccggaa acccgtgccc cggaagaatt cgcctcctac 840
acctggccgt attacctgtc aacgcgtccg gattatcgcc tgatgctgca aaacagctct 900
ctgatggaaa gttcctgtcc gtttatcgca gatcgcctgg cagctatgaa aatggaatcc 960
gtgcagccgt atgaactgct gacggcactg ccggaagctt caaaacagca attctatcgt 1020
atggccaaat ttgattacgc acgctttgct ggcctgttcg acctgtctcc gaagaaaaac 1080
ctgattatca ttggtacctc tcattcatcg gcggccagtg aacagcaaca ggcagcttac 1140
gtcgaacgta tcattcaaca gtatggcagt gattacgaca ttttctttaa accgcacccg 1200
gcagatagct ctagtgctgg ttatccggac cgctttgaag gtctgaccct gctgccgggt 1260
cagatgccgt ttgaaatctt cgtttgggcg ctgctggata aaatcgacat gattggcggt 1320
tatccgtcca ccacgtttat ttcagtgccg ctggataaag ttggctttct gttcgcggcc 1380
gatgccgacg gtctggtccg cccgctgaat atcctgttcc gtgacgctgc aaatgtcgaa 1440
tggattcaat aa 1452
<210> 24
<211> 1206
<212> DNA
<213> Actinobacillus suis
<400> 24
atggaacgca cgccgcaact gcaagcggtg gacatttaca ttgacttcgc aacgatcccg 60
agcctgagct actttctgca ctttctgaaa cataaacacg atgatcagcg tctgcgtctg 120
ttcagcctgg cccgttttga aatgccgcaa accctgattg aacagtatga aggcattatc 180
cagttctcgc gcaacgtgga acataatgtt gaaccgctgc tggaacagct gcaaacgatc 240
ctgtcacaag aaggtaaaca gtttgaactg catctgcacc tgaacctgtt tcattcgttc 300
gaaatgtttc tgaatctgag cccgacctac acgcagtaca aagaaaaaat ctctaaaatc 360
gttctgcacc tgtatgatga cggcagtgaa ggtgtcatga aacagtacca actgcagaaa 420
agctctagtc tggtgcagga tctggcggcc accaaagcat ctctggttag cctgttcgaa 480
aacggcgaag gttcgtttag ccagattgat ctgatccgtt atgtctggaa tgctgtgctg 540
gaaacccatt attacctgct gtctgatcac tttctgctgg acgaaaaact gcagccgctg 600
aaagcagaac tgggccatta ccaactgctg aacctgagtg cttatcagta cctgtcctca 660
gaagatctgc tgtggctgaa acagattctg aaaatcgaca ccgaactgga aagcctgatg 720
caaaaactga cggcgcagcc ggtgtatttc tttagcggta ccacgttttt caacatcagt 780
ttcgaagata aacaacgtct ggcgaatatc catgccattc tgatccgcga acacctggac 840
ccgaactccc agctgtttat tggcgaaccg tacctgtttg tcttcaaagg tcatccgaac 900
tcaccggaaa ttaatcaggc cctgcgtgaa tattacccga acgttatctt cctgccggaa 960
aatattccgt ttgaaatcct gaccctgctg ggcttctccc cgcaaaaaat tggcggtttt 1020
gcgtcaacga tccacgttaa ttccgaacag tcaaaactgg ccaaactgtt tttcctgacc 1080
tcgacggatg aacaagaacg ccagctgagc gacggttata ttaaacaata cgcactggct 1140
caggctatgc tggaaatgca actggtctcg caagaacaag tctattactg ctcgctgtcg 1200
tcgtaa 1206
<210> 25
<211> 1206
<212> DNA
<213> Actinobacillus capsulatus
<400> 25
atggaacgca tcccgcaact gcaagctgtc gatatttaca ttgacttcgc cacgatcccg 60
agcctgtcct actttctgca ctttctgaaa cataaacacg atcatcagcg tctgcgcctg 120
ttcagcctgg cgcgttttga aatgccgcag accgtcattg aacaatatga aggcattatc 180
cagttctcac gcaacgtgga acacaatgtt gaacaactgc tggaacagct gcaaacgatc 240
ctgtcgcagg aaggtaaaca atttgaactg cacctgcatc tgaacctgtt tcacagtttc 300
gaaatgtttc tgaatctgtc cccgacctac acgaaataca aagaaaaaat ctcaaaaatc 360
gttctgcatc tgtatgatga cggctcggaa ggtgtcatga aacagtacca actgcagcaa 420
agtaactccc tggcacagga tctggctagc accaaagcgt cactggtttc gctgttcaaa 480
aacggcgaag gtgccttttc tcagattgat ctgatccgtt atgtctggaa tgcagtgctg 540
gaaacccact attacctgct gtcagaccac tttctggccc atgaaaaact gcagccgctg 600
aaaattgaac tgggccatta ccagctgctg aatctgtctg cctatcaata cctgagctct 660
gaagatctgc tgtggctgaa acaaattctg aaaatcgacg cagaactgga aagtctgatg 720
cataaactga ccacgcagcc ggtgtatttc tttagcggta ccacgttttt caacatttcg 780
ttcgaagata aacagcgtct ggccaatatc cacgcaattc tgatccgcga acatctggac 840
ccgaacagtc agctgtttat cggcgaaccg tacctgtttg ttttcaaagg tcacccgaac 900
tccccggaaa ttaatcaggc tctgcgcgaa tattacccga acgcgatctt cctgccggaa 960
aatattccgt ttgaaatcct gaccctgctg ggcttcagcc cgcagaaaat tggcggtttt 1020
gcttctacga tccatgtgaa cagcgaacaa tctaaactgg cgaaactgtt tttcctgacc 1080
agtacggatg aacaggaacg taatcgctcc gacggttata ttaaacagta cgcgctggcc 1140
caagcaatgc tggaaatgca actggtctcg caagaacaag tctactactg ctcgctgtcg 1200
tcgtaa 1206
<210> 26
<211> 936
<212> DNA
<213> Haemophilus somnus
<400> 26
atgttccgtg aagacaatat gaacctgatt atctgctgta cgccgctgca agtgattatc 60
gccgaaaaaa ttatcgaacg ctatccggaa cagaaatttt atggcgttat gctggaatca 120
ttctacaacg ataaattcga cttctacgaa aacaaactga aacatctgtg ccacgaattt 180
ttctgtatca aaatcgcacg tttcaaactg gaacgctata aaaacctgct gtcactgctg 240
aaaatcaaaa acaaaacctt cgatcgtgtc ttcctggcta acatcgaaaa acgctacatc 300
catatcatcc tgtcgaacat tttctttaaa gaactgtaca ccttcgatga cggcacggcg 360
aacatcgccc cgaatagtca tctgtatcaa gaatacgatc actccctgaa aaaacgtatt 420
accgacatcc tgctgccgaa ccattacaac agcaacaaag tgaaaaacat cagcaaactg 480
cactactcta tctaccgctg caaaaacaac atcatcgata acatcgaata catgccgctg 540
tttaacctgg agaaaaaata cacggcacag gataaaagta tttccatcct gctgggtcaa 600
ccgattttct atgacgaaga gaaaaacatt cgtctgatca aagaagtcat cgccaaattc 660
aaaatcgatt actacttccc gcacccgcgc gaagattact acatcgacaa cgtgtcttac 720
atcaaaaccc cgctgatctt tgaagaattt tacgcggaac gttcaatcga aaattcgatc 780
aaaatctata cctttttcag ctctgccgtg ctgaacatcg ttacgaaaga aaatattgat 840
cgcatctacg cactgaaacc gaaactgacg gaaaaagcgt atctggattg ttacgacatc 900
ctgaaagatt tcggtatcaa agttatcgac atctaa 936
<210> 27
<211> 1200
<212> DNA
<213> Haemophilus ducreyi
<400> 27
atgctgattc aacagaacct ggaaatctac ctggactacg caaccatccc gagcctggcc 60
tgctttatgc acttcattca acacaaagat gacgtcgata gtattcgtct gtttggcctg 120
gcacgcttcg atatcccgca gtccattatc gaccgttacc cggctaacca cctgttttat 180
cacaacatcg ataatcgcga cctgaccgca gtgctgaacc agctggcgga tattctggcc 240
caggaaaata aacgttttca aatcaacctg catctgaacc tgtttcacag cattgacctg 300
tttttcgcta tttatccgat ctaccagcaa tatcagcata aaatttctac catccagctg 360
caactgtacg atgacggcag cgaaggtatt gttacgcagc attctctgtg caaaattgcg 420
gatctggaac agctgatcct gcaacacaaa aacgtgctgc tggaactgct gaccaaaggc 480
acggccaacg ttccgaatcc gaccctgctg cgttatctgt ggaacaatat tatcgattca 540
cagtttcatc tgatctcgga ccattttctg caacacccga aactgcaacc gctgaaacgt 600
ctgctgaaac gctacaccat tctggatttt acgtgttatc cgcgcttcaa tgccgaacag 660
aaacaactgc tgaaagaaat tctgcatatc tcaaacgaac tggaaaatct gctgaaactg 720
ctgaaacagc acaacacctt tctgttcacg ggcaccacgg cgtttaatct ggatcaggaa 780
aaactggacc tgctgaccca actgcatatc ctgctgctga acgaacacca gaatccgcat 840
tcaacgcact acattggcaa caattatctg ctgctgatca aaggtcatgc aaactcgccg 900
gctctgaatc ataccctggc gctgcacttt ccggatgcga ttttcctgcc ggccaatatt 960
ccgtttgaaa tcttcgcgat gctgggcttt acgccgaaca aaatgggcgg tttcgccagc 1020
acctcttaca ttaattatcc gacggaaaac atcaatcacc tgtttttcct gaccagtgat 1080
cagccgtcca ttcgcacgaa atggctggac tacgaaaaac aatttggtct gatgtattcc 1140
ctgctggcaa tgcagaaaat caacgaagat caggcgttta tgtgcaccat tcacaattaa 1200
<210> 28
<211> 1494
<212> DNA
<213> Photobacterium leiognathi
<400> 28
atgtgtaacg ataatcaaaa tacggtcgat gttgttgtga gcaccgttaa cgataacgtc 60
atcgaaaaca acacgtacca agttaaaccg atcgataccc cgaccacgtt tgacagttac 120
tcctggattc agacgtgcgg caccccgatc ctgaaagatg acgaaaaata ttcactgtcg 180
tttgatttcg tcgccccgga actggatcag gacgaaaaat tctgtttcga atttaccggc 240
gatgttgacg gtaaacgtta tgtcacgcag accaacctga cggtggttgc accgaccctg 300
gaagtttacg tcgatcatgc tagtctgccg tccctgcagc aactgatgaa aatcatccag 360
cagaaaaacg aatactcaca gaatgaacgt ttcatttcgt ggggccgcat cggtctgacg 420
gaagataacg cggaaaaact gaatgcccat atttatccgc tggcaggcaa caatacctca 480
caggaactgg tggatgcagt gatcgattac gctgactcga aaaaccgtct gaatctggaa 540
ctgaacacga ataccgcgca cagctttccg aacctggccc cgattctgcg cattatcagc 600
tctaaaagca acatcctgat ctctaacatc aacctgtacg atgacggcag tgctgaatat 660
gtgaacctgt acaattggaa agataccgaa gacaaatccg tgaaactgag cgattctttc 720
ctggttctga aagactactt taacggtatt agttccgaaa aaccgagcgg catctatggt 780
cgctacaact ggcatcaact gtataatacg tcttattact tcctgcgtaa agattacctg 840
accgttgaac cgcagctgca cgacctgcgc gaatatctgg gcggtagtct gaaacaaatg 900
tcctgggatg gcttttcaca gctgtcgaaa ggtgacaaag aactgttcct gaacattgtc 960
ggctttgatc aggaaaaact gcagcaagaa taccagcaat cagaactgcc gaatttcgtg 1020
tttacgggca ccacgacctg ggcaggcggt gaaaccaaag aatattacgc tcagcaacag 1080
gtgaacgtcg tgaacaatgc gattaatgaa accagcccgt attacctggg ccgtgaacat 1140
gacctgtttt tcaaaggtca cccgcgcggc ggtattatca atgatattat cctgggcagt 1200
ttcaacaata tgattgacat cccggccaaa gtgtcctttg aagttctgat gatgacgggt 1260
atgctgccgg ataccgtggg cggtattgcg tcatcgctgt attttagcat cccggccgaa 1320
aaagtctctt tcattgtgtt taccagctct gatacgatca ccgatcgtga agacgcgctg 1380
aaatctccgc tggtgcaggt tatgatgacc ctgggcattg ttaaagaaaa agatgtgctg 1440
ttctggtcgg atctgccgga ttgttcctcg ggtgtttgta ttgctcagta ttaa 1494
<210> 29
<211> 1497
<212> DNA
<213> Photobacterium sp.
<400> 29
atgagtgaag aaaacaccca gtccattatt aaaaacgaca tcaacaaaac catcatcgat 60
gaagaatacg ttaacctgga accgatcaac cagtctaaca tcagttttac caaacatagc 120
tgggtccaga cctgcggtac gcagcaactg ctgacggaac aaaacaaaga atcaatttcg 180
ctgagcgtgg ttgcgccgcg tctggatgac gatgaaaaat actgtttcga tttcaacggt 240
gttagtaata aaggcgaaaa atacatcacc aaagtcacgc tgaatgtcgt ggcaccgtct 300
ctggaagttt atgtggatca tgctagtctg ccgaccctgc aacaactgat ggatattatc 360
aaatcggaag aagaaaaccc gaccgcacag cgttacattg cttggggccg catcgtgccg 420
acggacgaac agatgaaaga actgaatatt accagctttg cgctgatcaa caatcacacg 480
ccggccgatc tggttcagga aattgtcaaa caggcgcaaa ccaaacatcg tctgaacgtg 540
aaactgagca gcaatacggc ccactcgttt gacaatctgg ttccgattct gaaagaactg 600
aacagcttca acaatgtgac cgttacgaat atcgatctgt atgacgatgg cagcgcggaa 660
tatgttaacc tgtacaattg gcgcgacacc ctgaacaaaa cggataatct gaaaattggc 720
aaagactatc tggaagatgt cattaacggt atcaatgaag ataccagcaa caccggcacg 780
agttccgtgt acaattggca gaaactgtat ccggctaact accattttct gcgtaaagat 840
tatctgaccc tggaaccgtc cctgcacgaa ctgcgcgact acattggtga ttcactgaaa 900
cagatgcaat gggacggctt caaaaaattc aactcgaaac agcaagaact gtttctgagc 960
atcgtgaatt tcgataaaca gaaactgcaa aacgaataca attcatcgaa cctgccgaat 1020
tttgtgttca ccggtaccac ggtttgggca ggcaaccacg aacgcgaata ctacgctaaa 1080
cagcaaatca acgttatcaa caacgccatc aacgaaagct ctccgcatta tctgggtaat 1140
tcctacgacc tgtttttcaa aggccacccg ggcggtggca ttatcaacac cctgatcatg 1200
cagaattatc cgtcaatggt cgatattccg tccaaaatct catttgaagt gctgatgatg 1260
accgacatgc tgccggatgc cgtggcaggt attgcgagtt ccctgtactt cacgatcccg 1320
gccgaaaaaa tcaaattcat cgttttcacc tctacggaaa ccattacgga tcgtgaaacc 1380
gccctgcgta gtccgctggt ccaggtgatg attaaactgg gcatcgtgaa agaagaaaat 1440
gtgctgttct gggcggacct gccgaattgc gaaacgggtg tctgtattgc tgtctga 1497
<210> 30
<211> 1449
<212> DNA
<213> Photobacterium leiognathi
<400> 30
atgaacgata atcaaaatac ggtggacgtg gtggtctcaa ccgtcaacga taacgtgatc 60
gaaaacaaca cgtaccaagt caaaccgatc gataccccga ccacgttcga ctcatactcg 120
tggattcaga cgtgcggcac cccgatcctg aaagatgacg aaaaatatag cctgtctttt 180
gatttcgttg ccccggaact ggatcaagac gaaaaattct gtttcgaatt taccggcgat 240
gtggatggta aacgttatgt gacgcagacc aacctgacgg tggttgcacc gaccctggaa 300
gtttacgtcg atcatgcttc actgccgtcg ctgcagcaac tgatgaaaat catccagcag 360
aaaaacgaat acagccagaa tgaacgcttt atttcttggg gccgtatccg cctgacggaa 420
gataacgcgg aaaaactgaa tgcccatatt tatccgctgg caggcaacaa taccagccag 480
gaactggtgg acgcagttat cgattacgct gactctaaaa accgtctgaa tctggaactg 540
aacacgaata ccggccacag tttccgtaac attgcgccga tcctgcgcgc caccagctct 600
aaaaacaaca tcctgatctc caacatcaac ctgtacgatg acggtagtgc tgaatatgtg 660
tccctgtaca actggaaaga taccgacaat aaatcacaga aactgagtga ttcctttctg 720
gttctgaaag actacctgaa tggcatcagt tccgaaaaac cgaacggtat ttatagcatc 780
tacaattggc atcagctgta tcactcatcg tattacttcc tgcgtaaaga ttacctgacg 840
gtggaaacca aactgcacga cctgcgcgaa tatctgggcg gttcactgaa acaaatgtcg 900
tgggatacct ttagccagct gtctaaaggc gacaaagaac tgttcctgaa cattgttggt 960
tttgatcagg aaaaactgca gcaagaatac cagcaaagcg aactgccgaa tttcgtcttt 1020
acgggcacca cgacctgggc aggcggtgaa accaaagaat attacgctca gcaacaggtg 1080
aacgtcgtga acaatgcgat taatgaaacc tctccgtatt acctgggccg tgaacatgac 1140
ctgtttttca aaggtcaccc gcgcggcggt attatcaatg atattatcct gggctcattc 1200
aacaatatga ttgacatccc ggccaaagtt tcgtttgaag tcctgatgat gacgggtatg 1260
ctgccggata ccgttggcgg tattgcgagc agcctgtatt ttagtatccc ggccgaaaaa 1320
gtgtccttca ttgtttttac cagttccgat acgatcaccg atcgcgaaga cgcgctgaaa 1380
agtccgctgg tccaagtgat gatgaccctg ggcattgtga aagaaaaaga tgtgctgttc 1440
tggtgctaa 1449
<210> 31
<211> 2028
<212> DNA
<213> Photobacterium damsela
<400> 31
atgaaaaaga tcctgaccgt cctgagcatc tttatcctga gcgcctgtaa tagcgacaac 60
acctctctga aagaaaccgt ctccagcaac agcgcggatg tggttgaaac ggaaacctat 120
cagctgaccc cgattgacgc cccgagcagc tttctgagcc attcttggga acagacgtgc 180
ggcaccccga tcctgaatga aagtgataaa caagcgattt cctttgactt cgtggccccg 240
gaactgaaac aggatgaaaa atactgtttc acgttcaaag gcatcaccgg tgaccaccgc 300
tacattacga acaccaccct gaccgttgtg gcaccgacgc tggaagtgta tatcgatcat 360
gctagtctgc cgagcctgca acaactgatt cacattatcc aggcgaaaga tgaatacccg 420
tcaaaccaac gctttgtttc gtggaaacgt gttaccgtcg atgcggacaa cgccaataaa 480
ctgaatattc atacctatcc gctgaaaggc aacaatacgt caccggaaat ggttgcggcc 540
atcgatgaat atgcacaatc gaaaaaccgc ctgaatattg aattttacac gaataccgct 600
catgtcttca acaatctgcc gccgattatc cagccgctgt acaacaacga aaaagtcaaa 660
atttcacaca tctcgctgta cgatgacggt agttccgaat atgtgagtct gtaccagtgg 720
aaagataccc cgaacaaaat tgaaacgctg gaaggcgaag tgagcctgct ggcaaattat 780
ctggctggca ccagcccgga tgcaccgaaa ggcatgggta accgttataa ttggcataaa 840
ctgtacgata ccgactatta ctttctgcgc gaagattatc tggacgtgga agcgaacctg 900
cacgatctgc gtgactacct gggttcatcg gcaaaacaga tgccgtggga tgaatttgct 960
aaactgagtg actcccagca aaccctgttt ctggatatcg ttggcttcga caaagaacag 1020
ctgcaacaac agtattcaca atcgccgctg ccgaatttta tttttaccgg caccaccacc 1080
tgggcgggcg gtgaaacgaa agaatattac gcccaacagc aagtgaacgt tattaacaat 1140
gccatcaatg aaaccagccc gtattacctg ggcaaagatt acgacctgtt tttcaaaggt 1200
catccggcag gcggtgtgat caacgatatt atcctgggca gttttccgga catgattaat 1260
atcccggcta aaatttcctt cgaagtgctg atgatgaccg atatgctgcc ggacacggtt 1320
gcaggtatcg ctagctctct gtattttacc attccggcgg ataaagtgaa ctttatcgtt 1380
ttcacgagtt ccgatacgat taccgaccgt gaagaagccc tgaaaagccc gctggtccag 1440
gtgatgctga ccctgggcat cgtcaaagaa aaagatgtgc tgttctgggc agaccacaaa 1500
gttaatagca tggaagtcgc gattgatgaa gcctgcaccc gcattatcgc aaaacgtcag 1560
ccgacggctt ctgatctgcg cctggtgatt gcgattatca aaacgatcac cgatctggaa 1620
cgtattggcg acgttgccga atctattgcg aaagtcgcgc tggaatcttt ttctaacaaa 1680
cagtacaatc tgctggttag cctggaatct ctgggtcaac ataccgtgcg catgctgcac 1740
gaagttctgg atgcattcgc tcgtatggac gtcaaagcag ctatcgaagt gtatcaggaa 1800
gatgaccgca tcgatcaaga atacgaaagt attgtccgtc agctgatggc ccacatgatg 1860
gaagatccgt catcgattcc gaacgttatg aaagtcatgt gggcggcccg ttccatcgaa 1920
cgcgttggtg atcgttgcca gaatatttgt gaatacatca tctacttcgt gaaaggcaaa 1980
gatgttcgcc acaccaaacc ggatgacttc ggtacgatgc tggactaa 2028
<210> 32
<211> 1533
<212> DNA
<213> Photobacterium damsela
<400> 32
atgaaaaaga tcctgaccgt cctgagcatc tttatcctga gcgcctgtaa tagcgacaac 60
acctctctga aagaaaccgt ctccagcaac agcgcggatg tggttgaaac ggaaacctat 120
cagctgaccc cgattgacgc cccgagcagc tttctgagcc attcttggga acagacgtgc 180
ggcaccccga tcctgaatga aagtgataaa caagcgattt cctttgactt cgtggccccg 240
gaactgaaac aggatgaaaa atactgtttc acgttcaaag gcatcaccgg tgaccaccgc 300
tacattacga acaccaccct gaccgttgtg gcaccgacgc tggaagtgta tatcgatcat 360
gctagtctgc cgagcctgca acaactgatt cacattatcc aggcgaaaga tgaatacccg 420
tcaaaccaac gctttgtttc gtggaaacgt gttaccgtcg atgcggacaa cgccaataaa 480
ctgaatattc atacctatcc gctgaaaggc aacaatacgt caccggaaat ggttgcggcc 540
atcgatgaat atgcacaatc gaaaaaccgc ctgaatattg aattttacac gaataccgct 600
catgtcttca acaatctgcc gccgattatc cagccgctgt acaacaacga aaaagtcaaa 660
atttcacaca tctcgctgta cgatgacggt agttccgaat atgtgagtct gtaccagtgg 720
aaagataccc cgaacaaaat tgaaacgctg gaaggcgaag tgagcctgct ggcaaattat 780
ctggctggca ccagcccgga tgcaccgaaa ggcatgggta accgttataa ttggcataaa 840
ctgtacgata ccgactatta ctttctgcgc gaagattatc tggacgtgga agcgaacctg 900
cacgatctgc gtgactacct gggttcatcg gcaaaacaga tgccgtggga tgaatttgct 960
aaactgagtg actcccagca aaccctgttt ctggatatcg ttggcttcga caaagaacag 1020
ctgcaacaac agtattcaca atcgccgctg ccgaatttta tttttaccgg caccaccacc 1080
tgggcgggcg gtgaaacgaa agaatattac gcccaacagc aagtgaacgt tattaacaat 1140
gccatcaatg aaaccagccc gtattacctg ggcaaagatt acgacctgtt tttcaaaggt 1200
catccggcag gcggtgtgat caacgatatt atcctgggca gttttccgga catgattaat 1260
atcccggcta aaatttcctt cgaagtgctg atgatgaccg atatgctgcc ggacacggtt 1320
gcaggtatcg ctagctctct gtattttacc attccggcgg ataaagtgaa ctttatcgtt 1380
ttcacgagtt ccgatacgat taccgaccgt gaagaagccc tgaaaagccc gctggtccag 1440
gtgatgctga ccctgggcat cgtcaaagaa aaagatgtgc tgttctgggc agacctgccg 1500
gactgctcgt ctggtgtgtg tatcgacaaa taa 1533
<210> 33
<211> 1269
<212> DNA
<213> Heliobacter acinonychis
<400> 33
atggggacca ttaaaaagcc cttaatcata gcaggaaatg gtccatcaat taaggaccta 60
gactatgctt tatttccaaa agacttcgat gtctttcgct gcaaccagtt ttacttcgag 120
gataaatatt acctaggacg cgaaataaaa ggagtgttct ttaacccttg tgtattaagc 180
agtcaaatgc aaacagtgca ataccttatg gacaatggcg aatatagcat agaacgcttc 240
ttttgcagtg tttcaacaga tcgccacgat tttgatgggg attaccaaac gattttaccg 300
gtagacggtt atttaaaagc acactatccg ttcgtctgcg atacattcag cttattcaaa 360
ggtcacgaag aaatcttaaa acacgtgaaa taccacctga aaacgtacag caaagaactt 420
agtgcgggtg tcttaatgtt attgagtgca gtggtattag gatacaaaga aatataccta 480
gtaggaatcg acttcggcgc ctcatcttgg gggcacttct atgacgaaag ccaatcccaa 540
cactttagca atcacatggc agattgtcac aatatctatt acgacatgct gactatttgt 600
ctctgtcaaa agtatgcaaa attgtacgca ttagcaccca attcaccatt atcacatttg 660
cttacactaa atccacaggc caaataccca tttgaactat tagataaacc tatcgggtat 720
actagcgacc taattattag tagcccgttg gaagagaagt tgctcgaatt taagaatatc 780
gaagagaagt tgcttgagtt caaaaacata gaagagaaac tcttagagtt caagaatatt 840
gaagagaaac tattagaatt taaaaacatc gaggaaaaac ttttggagtt caaaaatata 900
gaagagaaac tcctagagtt caagaacatt gaggaaaagt tgcttgagtt caaaaatatt 960
gaggaaaagt tgctcgaatt taagaatatc gaggaaaaac ttttggaatt taagaacata 1020
gaagaaaagt tactcgaatt taaaaacatt gaagagaaac tattggaatt taaaaatata 1080
gaggaaaagt tacttgagtt caaaaacata gaggaaaagt tacttgaatt taagaacata 1140
gaagagaaac ttctcgcaag ccgactgaac aacattctac gtaaaatcaa gcggaaaata 1200
cttccattct tttggggcgg aggtgtaacc ccaacattaa aagttagttt ccgttgggga 1260
gctgcataa 1269
<210> 34
<211> 469
<212> PRT
<213> Campylobacter coli
<400> 34
Met Gln Asn Val Ile Ile Ala Gly Asn Gly Pro Ser Leu Gln Ser Ile
1 5 10 15
Asn Tyr Gln Arg Leu Pro Lys Glu Tyr Asp Ile Phe Arg Cys Asn Gln
20 25 30
Phe Tyr Phe Glu Asp Lys Tyr Tyr Leu Gly Lys Asn Ile Lys Ala Ala
35 40 45
Phe Phe Asn Pro Tyr Pro Phe Leu Gln Gln Tyr His Thr Ala Lys Gln
50 55 60
Leu Val Phe Asn Asn Glu Tyr Lys Ile Glu Asn Ile Phe Cys Ser Thr
65 70 75 80
Phe Asn Leu Pro Phe Ile Glu Lys Asp Asn Phe Ile Asn Lys Phe Tyr
85 90 95
Asp Phe Phe Pro Asp Ala Lys Leu Gly His Lys Ile Ile Glu Asn Leu
100 105 110
Lys Glu Phe Tyr Ala Tyr Ile Lys Tyr Asn Glu Ile Tyr Leu Asn Lys
115 120 125
Arg Ile Thr Ser Gly Ile Tyr Met Cys Ala Ile Ala Ile Ala Leu Gly
130 135 140
Tyr Lys Asn Ile Tyr Leu Cys Gly Ile Asp Phe Tyr Glu Gly Glu Thr
145 150 155 160
Ile Tyr Pro Phe Lys Ala Met Ser Lys Asn Ile Lys Lys Ile Phe Pro
165 170 175
Trp Ile Lys Asp Phe Asn Pro Ser Asn Phe His Ser Lys Glu Tyr Asp
180 185 190
Ile Glu Ile Leu Lys Leu Leu Glu Ser Ile Tyr Lys Val Asn Ile Tyr
195 200 205
Ala Leu Cys Asp Asn Ser Ala Leu Ala Asn Tyr Phe Pro Leu Leu Val
210 215 220
Asn Thr Asp Asn Ser Phe Val Leu Glu Asn Lys Ser Asp Asp Cys Ile
225 230 235 240
Asn Asp Ile Leu Leu Thr Asn Asn Thr Pro Gly Ile Asn Phe Tyr Lys
245 250 255
Ser Gln Ile Gln Val Asn Asn Thr Glu Ile Leu Leu Leu Asn Phe Gln
260 265 270
Asn Met Ile Ser Ala Lys Glu Asn Glu Ile Ser Asn Leu Asn Lys Ile
275 280 285
Leu Gln Asp Ser Tyr Lys Thr Ile Asn Thr Lys Glu Asn Glu Ile Ser
290 295 300
Asn Leu Asn Lys Ile Leu Gln Asp Ser Tyr Lys Thr Ile Asn Thr Lys
305 310 315 320
Glu Asn Glu Ile Ser Asn Leu Asn Lys Ile Leu Gln Asp Lys Asp Lys
325 330 335
Leu Leu Ile Val Lys Glu Asn Leu Leu Asn Phe Lys Ser Arg His Gly
340 345 350
Lys Ala Lys Phe Arg Ile Gln Asn Gln Leu Ser Tyr Lys Leu Gly Gln
355 360 365
Ala Met Met Val Asn Ser Lys Ser Leu Leu Gly Tyr Ile Arg Met Pro
370 375 380
Phe Val Leu Ser Tyr Ile Lys Asp Lys His Lys Gln Glu Gln Lys Ile
385 390 395 400
Tyr Gln Glu Lys Ile Lys Lys Asp Pro Ser Leu Thr Leu Pro Pro Leu
405 410 415
Glu Asp Tyr Pro Asp Tyr Lys Glu Ala Leu Lys Glu Lys Glu Cys Leu
420 425 430
Thr Tyr Arg Leu Gly Gln Thr Leu Ile Lys Ala Asp Gln Glu Trp Tyr
435 440 445
Lys Gly Gly Tyr Val Lys Met Trp Phe Glu Ile Lys Lys Leu Lys Lys
450 455 460
Glu Tyr Lys Lys Lys
465
<210> 35
<211> 381
<212> PRT
<213> Vibrio sp.
<400> 35
Met Asn Asn Asp Asn Ser Thr Thr Thr Asn Asn Asn Ala Ile Glu Ile
1 5 10 15
Tyr Val Asp Arg Ala Thr Leu Pro Thr Ile Gln Gln Met Thr Lys Ile
20 25 30
Val Ser Gln Lys Thr Ser Asn Lys Lys Leu Ile Ser Trp Ser Arg Tyr
35 40 45
Pro Ile Thr Asp Lys Ser Leu Leu Lys Lys Ile Asn Ala Glu Phe Phe
50 55 60
Lys Glu Gln Phe Glu Leu Thr Glu Ser Leu Lys Asn Ile Ile Leu Ser
65 70 75 80
Glu Asn Ile Asp Asn Leu Ile Ile His Gly Asn Thr Leu Trp Ser Ile
85 90 95
Asp Val Val Asp Ile Ile Lys Glu Val Asn Leu Leu Gly Lys Asn Ile
100 105 110
Pro Ile Glu Leu His Phe Tyr Asp Asp Gly Ser Ala Glu Tyr Val Arg
115 120 125
Ile Tyr Glu Phe Ser Lys Leu Pro Glu Ser Glu Gln Lys Tyr Lys Thr
130 135 140
Ser Leu Ser Lys Asn Asn Ile Lys Phe Ser Ile Asp Gly Thr Asp Ser
145 150 155 160
Phe Lys Asn Thr Ile Glu Asn Ile Tyr Gly Phe Ser Gln Leu Tyr Pro
165 170 175
Thr Thr Tyr His Met Leu Arg Ala Asp Ile Phe Asp Thr Thr Leu Lys
180 185 190
Ile Asn Pro Leu Arg Glu Leu Leu Ser Asn Asn Ile Lys Gln Met Lys
195 200 205
Trp Asp Tyr Phe Lys Asp Phe Asn Tyr Lys Gln Lys Asp Ile Phe Tyr
210 215 220
Ser Leu Thr Asn Phe Asn Pro Lys Glu Ile Gln Glu Asp Phe Asn Lys
225 230 235 240
Asn Ser Asn Lys Asn Phe Ile Phe Ile Gly Ser Asn Ser Ala Thr Ala
245 250 255
Thr Ala Glu Glu Gln Ile Asn Ile Ile Ser Glu Ala Lys Lys Glu Asn
260 265 270
Ser Ser Ile Ile Thr Asn Ser Ile Ser Asp Tyr Asp Leu Phe Phe Lys
275 280 285
Gly His Pro Ser Ala Thr Phe Asn Glu Gln Ile Ile Asn Ala His Asp
290 295 300
Met Ile Glu Ile Asn Asn Lys Ile Pro Phe Glu Ala Leu Ile Met Thr
305 310 315 320
Gly Ile Leu Pro Asp Ala Val Gly Gly Met Gly Ser Ser Val Phe Phe
325 330 335
Ser Ile Pro Lys Glu Val Lys Asn Lys Phe Val Phe Tyr Lys Ser Gly
340 345 350
Thr Asp Ile Glu Asn Asn Ser Leu Ile Gln Val Met Leu Lys Leu Asn
355 360 365
Leu Ile Asn Arg Asp Asn Ile Lys Leu Ile Ser Asp Ile
370 375 380
<210> 36
<211> 390
<212> PRT
<213> Photobacterium sp.
<400> 36
Met Gly Cys Asn Ser Asp Ser Asn His Asn Asn Ser Asp Gly Asn Ile
1 5 10 15
Thr Lys Asn Lys Thr Ile Glu Val Tyr Val Asp Arg Ala Thr Leu Pro
20 25 30
Thr Ile Gln Gln Met Thr Gln Ile Ile Asn Glu Asn Ser Asn Asn Lys
35 40 45
Lys Leu Ile Ser Trp Ser Arg Tyr Pro Ile Asn Asp Glu Glu Leu Leu
50 55 60
Glu Ser Ile Asn Gly Ser Phe Phe Lys Asn Asn Ser Glu Leu Ile Lys
65 70 75 80
Ser Leu Asp Ser Met Ile Leu Thr Asn Asp Ile Lys Lys Val Ile Ile
85 90 95
Asn Gly Asn Thr Leu Trp Ala Ala Asp Val Val Asn Ile Ile Lys Ser
100 105 110
Ile Glu Ala Phe Gly Lys Lys Thr Glu Ile Glu Leu Asn Phe Tyr Asp
115 120 125
Asp Gly Ser Ala Glu Tyr Val Arg Leu Tyr Asp Phe Ser Lys Leu Pro
130 135 140
Glu Ser Glu Gln Glu Tyr Lys Ile Ser Leu Ser Lys Asp Asn Ile Leu
145 150 155 160
Ser Ser Ile Asn Gly Thr Gln Pro Phe Glu Asn Val Val Glu Asn Ile
165 170 175
Tyr Gly Phe Ser Gln Leu Tyr Pro Thr Thr Tyr His Met Leu Arg Ala
180 185 190
Asp Ile Phe Glu Thr Asn Leu Pro Leu Arg Ser Leu Lys Gly Val Leu
195 200 205
Ser Asn Asn Ile Lys Gln Met Lys Trp Asp Tyr Phe Lys Thr Phe Asn
210 215 220
Ser Gln Gln Lys Asp Lys Phe Tyr Asn Phe Thr Gly Phe Asn Pro Asp
225 230 235 240
Glu Ile Met Glu Gln Tyr Lys Ala Ser Pro Asn Lys Asn Phe Ile Phe
245 250 255
Val Gly Thr Asn Ser Gly Thr Ala Thr Ala Glu Gln Gln Ile Asp Ile
260 265 270
Leu Thr Glu Ala Lys Asn Pro Asn Ser Pro Ile Ile Thr Lys Ser Ile
275 280 285
Gln Gly Phe Asp Leu Phe Phe Lys Gly His Pro Ser Ala Thr Tyr Asn
290 295 300
Lys Gln Ile Ile Asp Ala His Asn Met Ile Glu Ile Tyr Asn Lys Ile
305 310 315 320
Pro Phe Glu Ala Leu Ile Met Thr Asp Ala Leu Pro Asp Ala Val Gly
325 330 335
Gly Met Gly Ser Ser Val Phe Phe Ser Leu Pro Asn Thr Val Glu Asn
340 345 350
Lys Phe Ile Phe Tyr Lys Ser Asp Thr Asp Ile Glu Asn Asn Ala Leu
355 360 365
Ile Gln Val Met Ile Glu Leu Asn Ile Val Asn Arg Asn Asp Val Lys
370 375 380
Leu Ile Ser Asp Leu Gln
385 390
<210> 37
<211> 388
<212> PRT
<213> Pasteurella multocida
<400> 37
Met Lys Thr Ile Thr Leu Tyr Leu Asp Pro Ala Ser Leu Pro Ala Leu
1 5 10 15
Asn Gln Leu Met Asp Phe Thr Gln Asn Asn Glu Asp Lys Thr His Pro
20 25 30
Arg Ile Phe Gly Leu Ser Arg Phe Lys Ile Pro Asp Asn Ile Ile Thr
35 40 45
Gln Tyr Gln Asn Ile His Phe Val Glu Leu Lys Asp Asn Arg Pro Thr
50 55 60
Glu Ala Leu Phe Thr Ile Leu Asp Gln Tyr Pro Gly Asn Ile Glu Leu
65 70 75 80
Asp Ile His Leu Asn Ile Ala His Ser Val Gln Leu Ile Arg Pro Ile
85 90 95
Leu Ala Tyr Arg Phe Lys His Leu Asp Arg Val Ser Ile Gln Arg Leu
100 105 110
Asn Leu Tyr Asp Asp Gly Ser Met Glu Tyr Val Asp Leu Glu Lys Glu
115 120 125
Glu Asn Lys Asp Ile Ser Ala Glu Ile Lys Gln Ala Glu Lys Gln Leu
130 135 140
Ser His Tyr Leu Leu Thr Gly Lys Ile Lys Phe Asp Asn Pro Thr Ile
145 150 155 160
Ala Arg Tyr Val Trp Gln Ser Ala Phe Pro Val Lys Tyr His Phe Leu
165 170 175
Ser Thr Asp Tyr Phe Glu Lys Ala Glu Phe Leu Gln Pro Leu Lys Glu
180 185 190
Tyr Leu Ala Glu Asn Tyr Gln Lys Met Asp Trp Thr Ala Tyr Gln Gln
195 200 205
Leu Thr Pro Glu Gln Gln Ala Phe Tyr Leu Thr Leu Val Gly Phe Asn
210 215 220
Asp Glu Val Lys Gln Ser Leu Glu Val Gln Gln Ala Lys Phe Ile Phe
225 230 235 240
Thr Gly Thr Thr Thr Trp Glu Gly Asn Thr Asp Val Arg Glu Tyr Tyr
245 250 255
Ala Gln Gln Gln Leu Asn Leu Leu Asn His Phe Thr Gln Ala Gly Gly
260 265 270
Asp Leu Phe Ile Gly Asp His Tyr Lys Ile Tyr Phe Lys Gly His Pro
275 280 285
Arg Gly Gly Glu Ile Asn Asp Tyr Ile Leu Asn Asn Ala Lys Asn Ile
290 295 300
Thr Asn Ile Pro Ala Asn Ile Ser Phe Glu Val Leu Met Met Thr Gly
305 310 315 320
Leu Leu Pro Asp Lys Val Gly Gly Val Ala Ser Ser Leu Tyr Phe Ser
325 330 335
Leu Pro Lys Glu Lys Ile Ser His Ile Ile Phe Thr Ser Asn Lys Gln
340 345 350
Val Lys Ser Lys Glu Asp Ala Leu Asn Asn Pro Tyr Val Lys Val Met
355 360 365
Arg Arg Leu Gly Ile Ile Asp Glu Ser Gln Val Ile Phe Trp Asp Ser
370 375 380
Leu Lys Gln Leu
385
<210> 38
<211> 371
<212> PRT
<213> Neisseria meningitidis
<400> 38
Met Gly Leu Lys Lys Ala Cys Leu Thr Val Leu Cys Leu Ile Val Phe
1 5 10 15
Cys Phe Gly Ile Phe Tyr Thr Phe Asp Arg Val Asn Gln Gly Glu Arg
20 25 30
Asn Ala Val Ser Leu Leu Lys Glu Lys Leu Phe Asn Glu Glu Gly Glu
35 40 45
Pro Val Asn Leu Ile Phe Cys Tyr Thr Ile Leu Gln Met Lys Val Ala
50 55 60
Glu Arg Ile Met Ala Gln His Pro Gly Glu Arg Phe Tyr Val Val Leu
65 70 75 80
Met Ser Glu Asn Arg Asn Glu Lys Tyr Asp Tyr Tyr Phe Asn Gln Ile
85 90 95
Lys Asp Lys Ala Glu Arg Ala Tyr Phe Phe His Leu Pro Tyr Gly Leu
100 105 110
Asn Lys Ser Phe Asn Phe Ile Pro Thr Met Ala Glu Leu Lys Val Lys
115 120 125
Ser Met Leu Leu Pro Lys Val Lys Arg Ile Tyr Leu Ala Ser Leu Glu
130 135 140
Lys Val Ser Ile Ala Ala Phe Leu Ser Thr Tyr Pro Asp Ala Glu Ile
145 150 155 160
Lys Thr Phe Asp Asp Gly Thr Gly Asn Leu Ile Gln Ser Ser Ser Tyr
165 170 175
Leu Gly Asp Glu Phe Ser Val Asn Gly Thr Ile Lys Arg Asn Phe Ala
180 185 190
Arg Met Met Ile Gly Asp Trp Ser Ile Ala Lys Thr Arg Asn Ala Ser
195 200 205
Asp Glu His Tyr Thr Ile Phe Lys Gly Leu Lys Asn Ile Met Asp Asp
210 215 220
Gly Arg Arg Lys Met Thr Tyr Leu Pro Leu Phe Asp Ala Ser Glu Leu
225 230 235 240
Lys Thr Gly Asp Glu Thr Gly Gly Thr Val Arg Ile Leu Leu Gly Ser
245 250 255
Pro Asp Lys Glu Met Lys Glu Ile Ser Glu Lys Ala Ala Lys Asn Phe
260 265 270
Lys Ile Gln Tyr Val Ala Pro His Pro Arg Gln Thr Tyr Gly Leu Ser
275 280 285
Gly Val Thr Thr Leu Asn Ser Pro Tyr Val Ile Glu Asp Tyr Ile Leu
290 295 300
Arg Glu Ile Lys Lys Asn Pro His Thr Arg Tyr Glu Ile Tyr Thr Phe
305 310 315 320
Phe Ser Gly Ala Ala Leu Thr Met Lys Asp Phe Pro Asn Val His Val
325 330 335
Tyr Ala Leu Lys Pro Ala Ser Leu Pro Glu Asp Tyr Trp Leu Lys Pro
340 345 350
Val Tyr Ala Leu Phe Thr Gln Ser Gly Ile Pro Ile Leu Thr Phe Asp
355 360 365
Asp Lys Asn
370
<210> 39
<211> 283
<212> PRT
<213> Pasteurella multocida
<400> 39
Met Asp Lys Phe Ala Glu His Glu Ile Pro Lys Ala Val Ile Val Ala
1 5 10 15
Gly Asn Gly Glu Ser Leu Ser Gln Ile Asp Tyr Arg Leu Leu Pro Lys
20 25 30
Asn Tyr Asp Val Phe Arg Cys Asn Gln Phe Tyr Phe Glu Glu Arg Tyr
35 40 45
Phe Leu Gly Asn Lys Ile Lys Ala Val Phe Phe Thr Pro Gly Val Phe
50 55 60
Leu Glu Gln Tyr Tyr Thr Leu Tyr His Leu Lys Arg Asn Asn Glu Tyr
65 70 75 80
Phe Val Asp Asn Val Ile Leu Ser Ser Phe Asn His Pro Thr Val Asp
85 90 95
Leu Glu Lys Ser Gln Lys Ile Gln Ala Leu Phe Ile Asp Val Ile Asn
100 105 110
Gly Tyr Glu Lys Tyr Leu Ser Lys Leu Thr Ala Phe Asp Val Tyr Leu
115 120 125
Arg Tyr Lys Glu Leu Tyr Glu Asn Gln Arg Ile Thr Ser Gly Val Tyr
130 135 140
Met Cys Ala Val Ala Ile Ala Met Gly Tyr Thr Asp Ile Tyr Leu Thr
145 150 155 160
Gly Ile Asp Phe Tyr Gln Ala Ser Glu Glu Asn Tyr Ala Phe Asp Asn
165 170 175
Lys Lys Pro Asn Ile Ile Arg Leu Leu Pro Asp Phe Arg Lys Glu Lys
180 185 190
Thr Leu Phe Ser Tyr His Ser Lys Asp Ile Asp Leu Glu Ala Leu Ser
195 200 205
Phe Leu Gln Gln His Tyr His Val Asn Phe Tyr Ser Ile Ser Pro Met
210 215 220
Ser Pro Leu Ser Lys His Phe Pro Ile Pro Thr Val Glu Asp Asp Cys
225 230 235 240
Glu Thr Thr Phe Val Ala Pro Leu Lys Glu Asn Tyr Ile Asn Asp Ile
245 250 255
Leu Leu Pro Pro His Phe Val Tyr Glu Lys Leu Gly Val Asp Lys Leu
260 265 270
Ala Ala Ala Leu Glu His His His His His His
275 280
<210> 40
<211> 385
<212> PRT
<213> Pasteurella dagmatis
<400> 40
Met Thr Ile Tyr Leu Asp Pro Ala Ser Leu Pro Thr Leu Asn Gln Leu
1 5 10 15
Met His Phe Thr Lys Glu Ser Glu Asp Lys Glu Thr Ala Arg Ile Phe
20 25 30
Gly Phe Ser Arg Phe Lys Leu Pro Glu Lys Ile Thr Glu Gln Tyr Asn
35 40 45
Asn Ile His Phe Val Glu Ile Lys Asn Asn Arg Pro Thr Glu Asp Ile
50 55 60
Phe Thr Ile Leu Asp Gln Tyr Pro Glu Lys Leu Glu Leu Asp Leu His
65 70 75 80
Leu Asn Ile Ala His Ser Ile Gln Leu Phe His Pro Ile Leu Gln Tyr
85 90 95
Arg Phe Lys His Pro Asp Arg Ile Ser Ile Lys Ser Leu Asn Leu Tyr
100 105 110
Asp Asp Gly Thr Met Glu Tyr Val Asp Leu Glu Lys Glu Glu Asn Lys
115 120 125
Asp Ile Lys Ser Ala Ile Lys Lys Ala Glu Lys Gln Leu Ser Asp Tyr
130 135 140
Leu Leu Thr Gly Lys Ile Asn Phe Asp Asn Pro Thr Leu Ala Arg Tyr
145 150 155 160
Val Trp Gln Ser Gln Tyr Pro Val Lys Tyr His Phe Leu Ser Thr Glu
165 170 175
Tyr Phe Glu Lys Ala Glu Phe Leu Gln Pro Leu Lys Thr Tyr Leu Ala
180 185 190
Gly Lys Tyr Gln Lys Met Asp Trp Ser Ala Tyr Glu Lys Leu Ser Pro
195 200 205
Glu Gln Gln Thr Phe Tyr Leu Lys Leu Val Gly Phe Ser Asp Glu Thr
210 215 220
Lys Gln Leu Phe His Thr Glu Gln Thr Lys Phe Ile Phe Thr Gly Thr
225 230 235 240
Thr Thr Trp Glu Gly Asn Thr Asp Ile Arg Glu Tyr Tyr Ala Lys Gln
245 250 255
Gln Leu Asn Leu Leu Lys His Phe Thr His Ser Glu Gly Asp Leu Phe
260 265 270
Ile Gly Asp Gln Tyr Lys Ile Tyr Phe Lys Gly His Pro Arg Gly Gly
275 280 285
Asp Ile Asn Asp Tyr Ile Leu Lys His Ala Lys Asp Ile Thr Asn Ile
290 295 300
Pro Ala Asn Ile Ser Phe Glu Ile Leu Met Met Thr Gly Leu Leu Pro
305 310 315 320
Asp Lys Val Gly Gly Val Ala Ser Ser Leu Tyr Phe Ser Leu Pro Lys
325 330 335
Glu Lys Ile Ser His Ile Ile Phe Thr Ser Asn Lys Lys Ile Lys Asn
340 345 350
Lys Glu Asp Ala Leu Asn Asp Pro Tyr Val Arg Val Met Leu Arg Leu
355 360 365
Gly Met Ile Asp Lys Ser Gln Ile Ile Phe Trp Asp Ser Leu Lys Gln
370 375 380
Leu
385
<210> 41
<211> 390
<212> PRT
<213> Photobacterium phosphoreum
<400> 41
Met Gly Cys Asn Ser Asp Ser Lys His Asn Asn Ser Asp Gly Asn Ile
1 5 10 15
Thr Lys Asn Lys Thr Ile Glu Val Tyr Val Asp Arg Ala Thr Leu Pro
20 25 30
Thr Ile Gln Gln Met Thr Gln Ile Ile Asn Glu Asn Ser Asn Asn Lys
35 40 45
Lys Leu Ile Ser Trp Ser Arg Tyr Pro Ile Asn Asp Glu Thr Leu Leu
50 55 60
Glu Ser Ile Asn Gly Ser Phe Phe Lys Asn Arg Pro Glu Leu Ile Lys
65 70 75 80
Ser Leu Asp Ser Met Ile Leu Thr Asn Glu Ile Lys Lys Val Ile Ile
85 90 95
Asn Gly Asn Thr Leu Trp Ala Val Asp Val Val Asn Ile Ile Lys Ser
100 105 110
Ile Glu Ala Leu Gly Lys Lys Thr Glu Ile Glu Leu Asn Phe Tyr Asp
115 120 125
Asp Gly Ser Ala Glu Tyr Val Arg Leu Tyr Asp Phe Ser Arg Leu Pro
130 135 140
Glu Ser Glu Gln Glu Tyr Lys Ile Ser Leu Ser Lys Asp Asn Ile Gln
145 150 155 160
Ser Ser Ile Asn Gly Thr Gln Pro Phe Asp Asn Ser Ile Glu Asn Ile
165 170 175
Tyr Gly Phe Ser Gln Leu Tyr Pro Thr Thr Tyr His Met Leu Arg Ala
180 185 190
Asp Ile Phe Glu Thr Asn Leu Pro Leu Thr Ser Leu Lys Arg Val Ile
195 200 205
Ser Asn Asn Ile Lys Gln Met Lys Trp Asp Tyr Phe Thr Thr Phe Asn
210 215 220
Ser Gln Gln Lys Asn Lys Phe Tyr Asn Phe Thr Gly Phe Asn Pro Glu
225 230 235 240
Lys Ile Lys Glu Gln Tyr Lys Ala Ser Pro His Glu Asn Phe Ile Phe
245 250 255
Ile Gly Thr Asn Ser Gly Thr Ala Thr Ala Glu Gln Gln Ile Asp Ile
260 265 270
Leu Thr Glu Ala Lys Lys Pro Asp Ser Pro Ile Ile Thr Asn Ser Ile
275 280 285
Gln Gly Leu Asp Leu Phe Phe Lys Gly His Pro Ser Ala Thr Tyr Asn
290 295 300
Gln Gln Ile Ile Asp Ala His Asn Met Ile Glu Ile Tyr Asn Lys Ile
305 310 315 320
Pro Phe Glu Ala Leu Ile Met Thr Asp Ala Leu Pro Asp Ala Val Gly
325 330 335
Gly Met Gly Ser Ser Val Phe Phe Ser Leu Pro Asn Thr Val Glu Asn
340 345 350
Lys Phe Ile Phe Tyr Lys Ser Asp Thr Asp Ile Glu Asn Asn Ala Leu
355 360 365
Ile Gln Val Met Ile Glu Leu Asn Ile Val Asn Arg Asn Asp Val Lys
370 375 380
Leu Ile Ser Asp Leu Gln
385 390
<210> 42
<211> 417
<212> PRT
<213> Avibacterium paragallinarum
<400> 42
Met Arg Lys Ile Ile Thr Phe Phe Ser Leu Phe Phe Ser Ile Ser Ala
1 5 10 15
Trp Cys Gln Lys Met Glu Ile Tyr Leu Asp Tyr Ala Ser Leu Pro Ser
20 25 30
Leu Asn Met Ile Leu Asn Leu Val Glu Asn Lys Asn Asn Glu Lys Val
35 40 45
Glu Arg Ile Ile Gly Phe Glu Arg Phe Asp Phe Asn Lys Glu Ile Leu
50 55 60
Asn Ser Phe Ser Lys Glu Arg Ile Glu Phe Ser Lys Val Ser Ile Leu
65 70 75 80
Asp Ile Lys Glu Phe Ser Asp Lys Leu Tyr Leu Asn Ile Glu Lys Ser
85 90 95
Asp Thr Pro Val Asp Leu Ile Ile His Thr Asn Leu Asp His Ser Val
100 105 110
Arg Ser Leu Leu Ser Ile Phe Lys Thr Leu Ser Pro Leu Phe His Lys
115 120 125
Ile Asn Ile Glu Lys Leu Tyr Leu Tyr Asp Asp Gly Ser Gly Asn Tyr
130 135 140
Val Asp Leu Tyr Gln His Arg Gln Glu Asn Ile Ser Ala Ile Leu Ile
145 150 155 160
Glu Ala Gln Lys Lys Leu Lys Asp Ala Leu Glu Asn Arg Glu Thr Asp
165 170 175
Thr Asp Lys Leu His Ser Leu Thr Arg Tyr Thr Trp His Lys Ile Phe
180 185 190
Pro Thr Glu Tyr Ile Leu Leu Arg Pro Asp Tyr Leu Asp Ile Asp Glu
195 200 205
Lys Met Gln Pro Leu Lys His Phe Leu Ser Asp Thr Ile Val Ser Met
210 215 220
Asp Leu Ser Arg Phe Ser His Phe Ser Lys Asn Gln Lys Glu Leu Phe
225 230 235 240
Leu Lys Ile Thr His Phe Asp Gln Asn Ile Phe Asn Glu Leu Asn Ile
245 250 255
Gly Thr Lys Asn Lys Glu Tyr Lys Thr Phe Ile Phe Thr Gly Thr Thr
260 265 270
Thr Trp Glu Lys Asp Lys Lys Lys Arg Leu Asn Asn Ala Lys Leu Gln
275 280 285
Thr Glu Ile Leu Glu Ser Phe Ile Lys Pro Asn Gly Lys Phe Tyr Leu
290 295 300
Gly Asn Asp Ile Lys Ile Phe Phe Lys Gly His Pro Lys Gly Asp Asp
305 310 315 320
Ile Asn Asp Tyr Ile Ile Arg Lys Thr Gly Ala Glu Lys Ile Pro Ala
325 330 335
Asn Ile Pro Phe Glu Val Leu Met Met Thr Asn Ser Leu Pro Asp Tyr
340 345 350
Val Gly Gly Ile Met Ser Thr Val Tyr Phe Ser Leu Pro Pro Lys Asn
355 360 365
Ile Asp Lys Val Val Phe Leu Gly Ser Glu Lys Ile Lys Asn Glu Asn
370 375 380
Asp Ala Lys Ser Gln Thr Leu Ser Lys Leu Met Leu Met Leu Asn Val
385 390 395 400
Ile Thr Pro Glu Gln Ile Phe Phe Glu Glu Met Pro Asn Pro Ile Asn
405 410 415
Phe
<210> 43
<211> 430
<212> PRT
<213> Campylobacter jejuni
<400> 43
Met Thr Arg Thr Arg Met Glu Asn Glu Leu Ile Val Ser Lys Asn Met
1 5 10 15
Gln Asn Ile Ile Ile Ala Gly Asn Gly Pro Ser Leu Lys Asn Ile Asn
20 25 30
Tyr Lys Arg Leu Pro Arg Glu Tyr Asp Val Phe Arg Cys Asn Gln Phe
35 40 45
Tyr Phe Glu Asp Lys Tyr Tyr Leu Gly Lys Lys Ile Lys Ala Val Phe
50 55 60
Phe Asn Pro Gly Val Phe Leu Gln Gln Tyr His Thr Ala Lys Gln Leu
65 70 75 80
Ile Leu Lys Asn Glu Tyr Glu Ile Lys Asn Ile Phe Cys Ser Thr Phe
85 90 95
Asn Leu Pro Phe Ile Glu Ser Asn Asp Phe Leu His Gln Phe Tyr Asn
100 105 110
Phe Phe Pro Asp Ala Lys Leu Gly Tyr Glu Val Ile Glu Asn Leu Lys
115 120 125
Glu Phe Tyr Ala Tyr Ile Lys Tyr Asn Glu Ile Tyr Phe Asn Lys Arg
130 135 140
Ile Thr Ser Gly Val Tyr Met Cys Ala Ile Ala Ile Ala Leu Gly Tyr
145 150 155 160
Lys Thr Ile Tyr Leu Cys Gly Ile Asp Phe Tyr Glu Gly Asp Val Ile
165 170 175
Tyr Pro Phe Glu Ala Met Ser Thr Asn Ile Lys Thr Ile Phe Pro Gly
180 185 190
Ile Lys Asp Phe Lys Pro Ser Asn Cys His Ser Lys Glu Tyr Asp Ile
195 200 205
Glu Ala Leu Lys Leu Leu Lys Ser Ile Tyr Lys Val Asn Ile Tyr Ala
210 215 220
Leu Cys Asp Asp Ser Ile Leu Ala Asn His Phe Pro Leu Ser Ile Asn
225 230 235 240
Ile Asn Asn Asn Phe Thr Leu Glu Asn Lys His Asn Asn Ser Ile Asn
245 250 255
Asp Ile Leu Leu Thr Asp Asn Thr Pro Gly Val Ser Phe Tyr Lys Asn
260 265 270
Gln Leu Lys Ala Asp Asn Lys Ile Met Leu Asn Phe Tyr Asn Ile Leu
275 280 285
His Ser Lys Asp Asn Leu Ile Lys Phe Leu Asn Lys Glu Ile Ala Val
290 295 300
Leu Lys Lys Gln Thr Thr Gln Arg Ala Lys Ala Arg Ile Gln Asn His
305 310 315 320
Leu Ser Tyr Lys Leu Gly Gln Ala Leu Ile Ile Asn Ser Lys Ser Val
325 330 335
Leu Gly Phe Leu Ser Leu Pro Phe Ile Ile Leu Ser Ile Val Ile Ser
340 345 350
His Lys Gln Glu Gln Lys Ala Tyr Lys Phe Lys Val Lys Lys Asn Pro
355 360 365
Asn Leu Ala Leu Pro Pro Leu Glu Thr Tyr Pro Asp Tyr Asn Glu Ala
370 375 380
Leu Lys Glu Lys Glu Cys Phe Thr Tyr Lys Leu Gly Glu Glu Phe Ile
385 390 395 400
Lys Ala Gly Lys Asn Trp Tyr Gly Glu Gly Tyr Ile Lys Phe Ile Phe
405 410 415
Lys Asp Val Pro Arg Leu Lys Arg Glu Phe Glu Lys Gly Glu
420 425 430
<210> 44
<211> 395
<212> PRT
<213> Heliobacter acinonychis
<400> 44
Met Asn Lys Lys Pro Leu Ile Ile Ala Gly Asn Gly Pro Ser Ile Lys
1 5 10 15
Asp Leu Asp Tyr Ala Leu Phe Pro Lys Asp Phe Asp Val Phe Arg Cys
20 25 30
Asn Gln Phe Tyr Phe Glu Asp Lys Tyr Tyr Leu Gly Arg Glu Ile Lys
35 40 45
Gly Val Phe Phe Asn Ala His Val Phe Asp Leu Gln Met Lys Ile Thr
50 55 60
Lys Ala Ile Val Lys Asn Gly Glu Tyr His Pro Asp His Ile Tyr Cys
65 70 75 80
Thr His Val Glu Pro Tyr Gly Tyr Val Asn Gly Asn Gln Gln Leu Met
85 90 95
Gln Glu Tyr Leu Glu Lys His Phe Val Gly Val Arg Ser Thr Tyr Ala
100 105 110
Tyr Leu Lys Asp Leu Glu Pro Phe Phe Ile Leu His Ser Lys Tyr Arg
115 120 125
Asn Phe Tyr Asp Gln His Phe Thr Thr Gly Ile Met Met Leu Leu Val
130 135 140
Ala Ile Gln Leu Gly Tyr Lys Glu Ile Tyr Leu Cys Gly Ile Asp Phe
145 150 155 160
Tyr Glu Asn Gly Phe Gly His Phe Tyr Glu Asn Gln Gly Gly Phe Phe
165 170 175
Glu Glu Asp Ser Asp Pro Met His Asp Lys Asn Ile Asp Ile Gln Ala
180 185 190
Leu Glu Leu Ala Lys Lys Tyr Ala Lys Ile Tyr Ala Leu Val Pro Asn
195 200 205
Ser Ala Leu Val Lys Met Ile Pro Leu Ser Ser Gln Lys Gly Val Leu
210 215 220
Glu Lys Val Lys Asp Arg Ile Gly Leu Gly Glu Phe Lys Arg Glu Lys
225 230 235 240
Phe Gly Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys
245 250 255
Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg
260 265 270
Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu
275 280 285
Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys
290 295 300
Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg
305 310 315 320
Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu Glu Arg Gln Lys Glu Leu
325 330 335
Glu Leu Glu Arg Ser Leu Lys Ala Arg Leu Lys Ala Val Leu Ala Ser
340 345 350
Lys Gly Ile Arg Gly Asp Asn Leu Ile Ile Val Ser Leu Lys Asp Thr
355 360 365
Tyr Arg Leu Phe Lys Gly Gly Phe Ala Leu Leu Leu Asp Leu Lys Ala
370 375 380
Leu Lys Ser Ile Ile Lys Ala Phe Leu Lys Arg
385 390 395
<210> 45
<211> 260
<212> PRT
<213> Campylobacter jejuni
<400> 45
Met Gly Lys Lys Val Ile Ile Ala Gly Asn Gly Pro Ser Leu Lys Glu
1 5 10 15
Ile Asp Tyr Ser Arg Leu Pro Asn Asp Phe Asp Val Phe Arg Cys Asn
20 25 30
Gln Phe Tyr Phe Glu Asp Lys Tyr Tyr Leu Gly Lys Lys Cys Lys Ala
35 40 45
Val Phe Tyr Asn Pro Ile Leu Phe Phe Glu Gln Tyr Tyr Thr Leu Lys
50 55 60
His Leu Ile Gln Asn Gln Glu Tyr Glu Thr Glu Leu Ile Met Cys Ser
65 70 75 80
Asn Tyr Asn Gln Ala His Leu Glu Asn Glu Asn Phe Val Lys Thr Phe
85 90 95
Tyr Asp Tyr Phe Pro Asp Ala His Leu Gly Tyr Asp Phe Phe Lys Gln
100 105 110
Leu Lys Asp Phe Asn Ala Tyr Phe Lys Phe His Glu Ile Tyr Phe Asn
115 120 125
Gln Arg Ile Thr Ser Gly Val Tyr Met Cys Ala Val Ala Ile Ala Leu
130 135 140
Gly Tyr Lys Glu Ile Tyr Leu Ser Gly Ile Asp Phe Tyr Gln Asn Gly
145 150 155 160
Ser Ser Tyr Ala Phe Asp Thr Lys Gln Lys Asn Leu Leu Lys Leu Ala
165 170 175
Pro Asn Phe Lys Asn Asp Asn Ser His Tyr Ile Gly His Ser Lys Asn
180 185 190
Thr Asp Ile Lys Ala Leu Glu Phe Leu Glu Lys Thr Tyr Lys Ile Lys
195 200 205
Leu Tyr Cys Leu Cys Pro Asn Ser Leu Leu Ala Asn Phe Ile Glu Leu
210 215 220
Ala Pro Asn Leu Asn Ser Asn Phe Ile Ile Gln Glu Lys Asn Asn Tyr
225 230 235 240
Thr Lys Asp Ile Leu Ile Pro Ser Ser Glu Ala Tyr Gly Lys Phe Ser
245 250 255
Lys Asn Ile Asn
260
<210> 46
<211> 298
<212> PRT
<213> Streptococcus entericus
<400> 46
Met Lys Lys Val Tyr Phe Cys His Thr Val Tyr His Leu Leu Ile Thr
1 5 10 15
Leu Cys Lys Ile Ser Val Glu Glu Gln Val Glu Ile Ile Val Phe Asp
20 25 30
Thr Val Ser Asn His Glu Leu Ile Val Gln Lys Ile Arg Asp Val Phe
35 40 45
Val Asn Thr Thr Val Leu Phe Ala Glu Gln Asn Thr Asp Phe Ser Ile
50 55 60
Leu Glu Ile Asp Arg Ala Thr Asp Ile Tyr Val Phe Asn Asp Trp Thr
65 70 75 80
Pro Ile Gly Ala Tyr Leu Arg Lys Asn Lys Leu Phe Tyr His Leu Ile
85 90 95
Glu Asp Gly Tyr Asn Tyr His Glu Tyr Asn Val Tyr Ala Asn Ala Leu
100 105 110
Thr Met Lys Arg Arg Leu Leu Asn Phe Val Leu Arg Arg Glu Glu Pro
115 120 125
Ser Gly Phe Ser Arg Tyr Val Arg Ser Ile Glu Val Asn Arg Val Lys
130 135 140
Tyr Leu Pro Asn Asp Cys Arg Lys Ser Lys Trp Val Glu Lys Pro Arg
145 150 155 160
Ser Ala Leu Phe Glu Asn Leu Val Pro Glu His Lys Gln Lys Ile Ile
165 170 175
Thr Ile Phe Gly Leu Glu Asn Tyr Gln Asp Ser Leu Arg Gly Val Leu
180 185 190
Val Leu Thr Gln Pro Leu Val Gln Asp Tyr Trp Asp Arg Asp Ile Thr
195 200 205
Thr Glu Glu Glu Gln Leu Glu Phe Tyr Arg Gln Ile Val Glu Ser Tyr
210 215 220
Gly Glu Gly Glu Gln Val Phe Phe Lys Ile His Pro Arg Asp Lys Val
225 230 235 240
Asp Tyr Ser Ser Leu Thr Asn Val Ile Phe Leu Lys Lys Asn Val Pro
245 250 255
Met Glu Val Tyr Glu Leu Ile Ala Asp Cys His Phe Thr Lys Gly Ile
260 265 270
Thr His Ser Ser Thr Ala Leu Asp Phe Leu Ser Cys Val Asp Lys Lys
275 280 285
Ile Thr Leu Lys Gln Met Lys Ala Asn Ser
290 295
<210> 47
<211> 295
<212> PRT
<213> Haemophilus ducreyi
<400> 47
Met Lys Glu Ile Ala Ile Ile Ser Asn Gln Arg Met Phe Phe Leu Tyr
1 5 10 15
Cys Leu Leu Thr Asn Lys Asn Val Glu Asp Val Phe Phe Ile Phe Glu
20 25 30
Lys Gly Ala Met Pro Asn Asn Leu Thr Ser Ile Ser His Phe Ile Val
35 40 45
Leu Asp His Ser Lys Ser Glu Cys Tyr Asp Phe Phe Tyr Phe Asn Phe
50 55 60
Ile Ser Cys Lys Tyr Arg Leu Arg Gly Leu Asp Val Tyr Gly Ala Asp
65 70 75 80
His Ile Lys Gly Ala Lys Phe Phe Leu Glu Arg His Arg Phe Phe Val
85 90 95
Val Glu Asp Gly Met Met Asn Tyr Ser Lys Asn Met Tyr Ala Phe Ser
100 105 110
Leu Phe Arg Thr Arg Asn Pro Val Ile Leu Pro Gly Gly Phe His Pro
115 120 125
Asn Val Lys Thr Ile Phe Leu Thr Lys Asp Asn Pro Ile Pro Asp Gln
130 135 140
Ile Ala His Lys Arg Glu Ile Ile Asn Ile Lys Thr Leu Trp Gln Ala
145 150 155 160
Lys Thr Ala Thr Glu Lys Thr Lys Ile Leu Ser Phe Phe Glu Ile Asp
165 170 175
Met Gln Glu Ile Ser Val Ile Lys Asn Arg Ser Phe Val Leu Tyr Thr
180 185 190
Gln Pro Leu Ser Glu Asp Lys Leu Leu Thr Glu Ala Glu Lys Ile Asp
195 200 205
Ile Tyr Arg Thr Ile Leu Thr Lys Tyr Asn His Ser Gln Thr Val Ile
210 215 220
Lys Pro His Pro Arg Asp Lys Thr Asp Tyr Lys Gln Leu Phe Pro Asp
225 230 235 240
Ala Tyr Val Met Lys Gly Thr Tyr Pro Ser Glu Leu Leu Thr Leu Leu
245 250 255
Gly Val Asn Phe Asn Lys Val Ile Thr Leu Phe Ser Thr Ala Val Phe
260 265 270
Asp Tyr Pro Lys Glu Lys Ile Asp Phe Tyr Gly Thr Ala Val His Pro
275 280 285
Lys Leu Leu Asp Phe Phe Asp
290 295
<210> 48
<211> 488
<212> PRT
<213> Alistipes sp.
<400> 48
Met Ala Leu Leu Ser Gly Thr Ala Ala Cys Ser Asp Asp Glu Val Ser
1 5 10 15
Gln Asn Leu Ile Val Ile Asn Gly Gly Glu His Phe Leu Ser Leu Asp
20 25 30
Gly Leu Ala Arg Ala Gly Lys Ile Ser Val Leu Ala Pro Ala Pro Trp
35 40 45
Arg Val Thr Lys Ala Ala Gly Asp Thr Trp Phe Arg Leu Ser Ala Thr
50 55 60
Glu Gly Pro Ala Gly Tyr Ser Glu Val Glu Leu Ser Leu Asp Glu Asn
65 70 75 80
Pro Gly Ala Ala Arg Ser Ala Gln Leu Ala Phe Ala Cys Gly Asp Ala
85 90 95
Ile Val Pro Phe Arg Leu Ser Gln Gly Ala Leu Ser Ala Gly Tyr Asp
100 105 110
Ser Pro Asp Tyr Tyr Phe Tyr Val Thr Phe Gly Thr Met Pro Thr Leu
115 120 125
Tyr Ala Gly Ile His Leu Leu Ser His Asp Lys Pro Gly Tyr Val Phe
130 135 140
Tyr Ser Arg Ser Lys Thr Phe Asp Pro Ala Glu Phe Pro Ala Arg Ala
145 150 155 160
Glu Val Thr Thr Ala Ala Asp Arg Thr Ala Asp Ala Thr Gln Ala Glu
165 170 175
Met Glu Ala Met Ala Arg Glu Met Lys Arg Arg Ile Leu Glu Ile Asn
180 185 190
Ser Ala Asp Pro Thr Ala Val Phe Gly Leu Tyr Val Asp Asp Leu Arg
195 200 205
Cys Arg Ile Gly Tyr Asp Trp Phe Val Ala Gln Gly Ile Asp Ser Ala
210 215 220
Arg Val Lys Val Ser Met Leu Ser Asp Gly Thr Gly Thr Tyr Asn Asn
225 230 235 240
Phe Tyr Asn Tyr Phe Gly Asp Ala Ala Thr Ala Glu Gln Asn Trp Glu
245 250 255
Ser Tyr Ala Ser Glu Val Glu Ala Leu Asp Trp Asn His Gly Gly Arg
260 265 270
Tyr Pro Glu Thr Arg Ser Leu Pro Glu Phe Glu Ser Tyr Thr Trp Pro
275 280 285
Tyr Tyr Leu Ser Thr Arg Pro Asp Tyr Arg Leu Val Val Gln Asp Gly
290 295 300
Ser Leu Leu Glu Ser Ser Cys Pro Phe Ile Thr Glu Lys Leu Gly Glu
305 310 315 320
Met Glu Ile Glu Ser Ile Gln Pro Tyr Glu Met Leu Ser Ala Leu Pro
325 330 335
Glu Ser Ser Arg Lys Arg Phe Tyr Asp Met Ala Gly Phe Asp Tyr Asp
340 345 350
Lys Phe Ala Ala Leu Phe Asp Ala Ser Pro Lys Lys Asn Leu Ile Ile
355 360 365
Ile Gly Thr Ser His Ala Asp Asp Ala Ser Ala Arg Leu Gln Arg Asp
370 375 380
Tyr Val Ala Arg Ile Met Glu Gln Tyr Gly Ala Gln Tyr Asp Val Phe
385 390 395 400
Phe Lys Pro His Pro Ala Asp Thr Thr Ser Ala Gly Tyr Glu Thr Glu
405 410 415
Phe Pro Gly Leu Thr Leu Leu Pro Gly Gln Met Pro Phe Glu Ile Phe
420 425 430
Val Trp Ser Leu Ile Asp Arg Val Asp Met Ile Gly Gly Tyr Pro Ser
435 440 445
Thr Val Phe Leu Thr Val Pro Val Asp Lys Val Arg Phe Ile Phe Ala
450 455 460
Ala Asp Ala Ala Ser Leu Val Arg Pro Leu Asn Ile Leu Phe Arg Asp
465 470 475 480
Ala Thr Asp Val Glu Trp Met Gln
485
<210> 49
<211> 291
<212> PRT
<213> Campylobacter jejuni
<400> 49
Met Lys Lys Val Ile Ile Ala Gly Asn Gly Pro Ser Leu Lys Glu Ile
1 5 10 15
Asp Tyr Ser Arg Leu Pro Asn Asp Phe Asp Val Phe Arg Cys Asn Gln
20 25 30
Phe Tyr Phe Glu Asp Lys Tyr Tyr Leu Gly Lys Lys Cys Lys Ala Val
35 40 45
Phe Tyr Thr Pro Asn Phe Phe Phe Glu Gln Tyr Tyr Thr Leu Lys His
50 55 60
Leu Ile Gln Asn Gln Glu Tyr Glu Thr Glu Leu Ile Met Cys Ser Asn
65 70 75 80
Tyr Asn Gln Ala His Leu Glu Asn Glu Asn Phe Val Lys Thr Phe Tyr
85 90 95
Asp Tyr Phe Pro Asp Ala His Leu Gly Tyr Asp Phe Phe Lys Gln Leu
100 105 110
Lys Glu Phe Asn Ala Tyr Phe Lys Phe His Glu Ile Tyr Phe Asn Gln
115 120 125
Arg Ile Thr Ser Gly Val Tyr Met Cys Ala Val Ala Ile Ala Leu Gly
130 135 140
Tyr Lys Glu Ile Tyr Leu Ser Gly Ile Asp Phe Tyr Gln Asn Gly Ser
145 150 155 160
Ser Tyr Ala Phe Asp Thr Lys Gln Glu Asn Leu Leu Lys Leu Ala Pro
165 170 175
Asp Phe Lys Asn Asp Arg Ser His Tyr Ile Gly His Ser Lys Asn Thr
180 185 190
Asp Ile Lys Ala Leu Glu Phe Leu Glu Lys Thr Tyr Lys Ile Lys Leu
195 200 205
Tyr Cys Leu Cys Pro Asn Ser Leu Leu Ala Asn Phe Ile Glu Leu Ala
210 215 220
Pro Asn Leu Asn Ser Asn Phe Ile Ile Gln Glu Lys Asn Asn Tyr Thr
225 230 235 240
Lys Asp Ile Leu Ile Pro Ser Ser Glu Ala Tyr Gly Lys Phe Ser Lys
245 250 255
Asn Ile Asn Phe Lys Lys Ile Lys Ile Lys Glu Asn Val Tyr Tyr Lys
260 265 270
Leu Ile Lys Asp Leu Leu Arg Leu Pro Ser Asp Ile Lys His Tyr Phe
275 280 285
Lys Gly Lys
290
<210> 50
<211> 312
<212> PRT
<213> Streptococcus agalactiae
<400> 50
Met Thr Asn Arg Lys Ile Tyr Val Cys His Thr Leu Tyr His Leu Leu
1 5 10 15
Ile Cys Leu Tyr Lys Glu Glu Ile Tyr Ser Asn Leu Glu Ile Ile Leu
20 25 30
Ser Ser Ser Ile Pro Asp Val Asp Asn Leu Glu Lys Lys Leu Lys Ser
35 40 45
Lys Thr Ile Asn Ile His Ile Leu Glu Glu Ser Ser Gly Glu Ser Glu
50 55 60
Glu Leu Leu Ser Val Leu Lys Asp Ala Gly Leu Ser Tyr Ser Lys Phe
65 70 75 80
Asp Ser Asn Cys Phe Ile Phe Asn Asp Ala Thr Pro Ile Gly Arg Thr
85 90 95
Leu Ile Lys His Gly Ile Tyr Tyr Asn Leu Ile Glu Asp Gly Leu Asn
100 105 110
Cys Phe Thr Tyr Ser Ile Phe Ser Gln Lys Leu Trp Lys Tyr Tyr Val
115 120 125
Lys Lys Tyr Ile Leu His Lys Ile Gln Pro His Gly Phe Ser Arg Tyr
130 135 140
Cys Leu Gly Ile Glu Val Asn Ser Leu Val Asn Leu Pro Lys Asp Pro
145 150 155 160
Arg Tyr Lys Lys Phe Ile Glu Val Pro Arg Lys Glu Leu Phe Asp Asn
165 170 175
Val Thr Glu Tyr Gln Lys Glu Met Ala Ile Asn Leu Phe Gly Ala Val
180 185 190
Arg Val Ser Ile Lys Ser Pro Ser Val Leu Val Leu Thr Gln Pro Leu
195 200 205
Ser Ile Asp Lys Glu Phe Met Ser Tyr Asn Asn Lys Ile Glu Thr Ser
210 215 220
Glu Glu Gln Phe Asn Phe Tyr Lys Ser Ile Val Asn Glu Tyr Ile Asn
225 230 235 240
Lys Gly Tyr Asn Val Tyr Leu Lys Val His Pro Arg Asp Val Val Asp
245 250 255
Tyr Ser Lys Leu Pro Val Glu Leu Leu Pro Ser Asn Val Pro Met Glu
260 265 270
Ile Ile Glu Leu Met Leu Thr Gly Arg Phe Glu Cys Gly Ile Thr His
275 280 285
Ser Ser Thr Ala Leu Asp Phe Leu Thr Cys Val Asp Lys Lys Ile Thr
290 295 300
Leu Val Asp Leu Lys Asp Ile Lys
305 310
<210> 51
<211> 410
<212> PRT
<213> Bibersteinia trehalosi
<400> 51
Met Glu Phe Cys Lys Met Ala Thr Thr Gln Lys Ile Cys Val Tyr Leu
1 5 10 15
Asp Tyr Ala Thr Ile Pro Ser Leu Asn Tyr Ile Leu His Phe Ala Gln
20 25 30
His Phe Glu Asp Gln Glu Thr Ile Arg Leu Phe Gly Leu Ser Arg Phe
35 40 45
His Ile Pro Glu Ser Val Ile Gln Arg Tyr Pro Lys Gly Val Val Gln
50 55 60
Phe Tyr Pro Asn Gln Glu Lys Asp Phe Ser Ala Leu Leu Leu Ala Leu
65 70 75 80
Lys Asn Ile Leu Ile Glu Val Lys Gln Gln Gln Arg Lys Cys Glu Ile
85 90 95
Glu Leu His Leu Asn Leu Phe His Tyr Gln Leu Leu Leu Leu Pro Phe
100 105 110
Leu Ser Leu Tyr Leu Asp Thr Gln Asp Tyr Cys His Leu Thr Leu Lys
115 120 125
Phe Tyr Asp Asp Gly Ser Glu Ala Ile Ser Ala Leu Gln Glu Leu Ala
130 135 140
Leu Ala Pro Asp Leu Ala Ala Gln Ile Gln Phe Glu Lys Gln Gln Phe
145 150 155 160
Asp Glu Leu Val Val Lys Lys Ser Phe Lys Leu Ser Leu Leu Ser Arg
165 170 175
Tyr Phe Trp Gly Lys Leu Phe Glu Ser Glu Tyr Ile Trp Phe Asn Gln
180 185 190
Ala Ile Leu Gln Lys Ala Glu Leu Gln Ile Leu Lys Gln Glu Ile Ser
195 200 205
Ser Ser Arg Gln Met Asp Phe Ala Ile Tyr Gln Gln Met Ser Asp Glu
210 215 220
Gln Lys Gln Leu Val Leu Glu Ile Leu Asn Ile Asp Leu Asn Lys Val
225 230 235 240
Ala Tyr Leu Lys Gln Leu Met Glu Asn Gln Pro Ser Phe Leu Phe Leu
245 250 255
Gly Thr Thr Leu Phe Asn Ile Thr Gln Glu Thr Lys Thr Trp Leu Met
260 265 270
Gln Met His Val Asp Leu Ile Gln Gln Tyr Cys Leu Pro Ser Gly Gln
275 280 285
Phe Phe Asn Asn Lys Ala Gly Tyr Leu Cys Phe Tyr Lys Gly His Pro
290 295 300
Asn Glu Lys Glu Met Asn Gln Met Ile Leu Ser Gln Phe Lys Asn Leu
305 310 315 320
Ile Ala Leu Pro Asp Asp Ile Pro Leu Glu Ile Leu Leu Leu Leu Gly
325 330 335
Val Ile Pro Ser Lys Val Gly Gly Phe Ala Ser Ser Ala Leu Phe Asn
340 345 350
Phe Thr Pro Ala Gln Ile Glu Asn Ile Ile Phe Phe Thr Pro Arg Tyr
355 360 365
Phe Glu Lys Asp Asn Arg Leu His Ala Thr Gln Tyr Arg Leu Met Gln
370 375 380
Gly Leu Ile Glu Leu Gly Tyr Leu Asp Ala Glu Lys Ser Val Thr His
385 390 395 400
Phe Glu Ile Met Gln Leu Leu Thr Lys Glu
405 410
<210> 52
<211> 406
<212> PRT
<213> Haemophilus parahaemolyticus
<400> 52
Met Thr Glu Gln Tyr Ile Lys Asn Val Glu Val Tyr Leu Asp Tyr Ala
1 5 10 15
Thr Ile Pro Thr Leu Asn Tyr Phe Tyr His Phe Thr Glu Asn Lys Asp
20 25 30
Asp Ile Ala Thr Ile Arg Leu Phe Gly Leu Gly Arg Phe Asn Ile Ser
35 40 45
Lys Ser Ile Ile Glu Ser Tyr Pro Glu Gly Ile Ile Arg Tyr Cys Pro
50 55 60
Ile Ile Phe Glu Asp Gln Thr Ala Phe Gln Gln Leu Phe Ile Thr Leu
65 70 75 80
Leu Thr Glu Asp Ser Phe Cys Gln Tyr Arg Phe Asn Phe His Ile Asn
85 90 95
Leu Phe His Ser Trp Lys Met Leu Ile Pro Leu Leu His Ile Ile Trp
100 105 110
Gln Phe Lys His Lys Val Leu Asp Ile Lys Leu Asn Phe Tyr Asp Asp
115 120 125
Gly Ser Glu Gly Leu Val Thr Leu Ser Lys Ile Glu Gln Asn Tyr Ser
130 135 140
Ser Glu Ile Leu Gln Lys Ile Ile Asp Ile Asp Ser Gln Ser Phe Tyr
145 150 155 160
Ala Asp Lys Leu Ser Phe Leu Asp Glu Asp Ile Ala Arg Tyr Leu Trp
165 170 175
Asn Ser Leu Phe Glu Ser His Tyr Tyr Leu Leu Asn Asp Phe Leu Leu
180 185 190
Lys Asn Glu Lys Leu Ser Leu Leu Lys Asn Ser Ile Lys Tyr Cys His
195 200 205
Ile Met Asp Leu Glu Arg Tyr Leu Gln Phe Thr Gln Glu Glu Lys Asp
210 215 220
Phe Phe Asn Glu Leu Leu Gly Ile Asn Ile Gln Ser Leu Glu Asp Lys
225 230 235 240
Ile Lys Ile Phe Gln Gln Lys Lys Thr Phe Ile Phe Thr Gly Thr Thr
245 250 255
Ile Phe Ser Leu Pro Lys Glu Glu Glu Glu Thr Leu Tyr Arg Leu His
260 265 270
Leu Asn Ala Ile Leu Asn Tyr Ile His Pro Asn Gly Lys Tyr Phe Ile
275 280 285
Gly Asp Gly Phe Thr Leu Val Ile Lys Gly His Pro His Gln Lys Glu
290 295 300
Met Asn Ser Arg Leu Glu Lys Ser Phe Glu Lys Ala Val Met Leu Pro
305 310 315 320
Asp Asn Ile Pro Phe Glu Ile Leu Tyr Leu Ile Gly Cys Lys Pro Asp
325 330 335
Lys Ile Gly Gly Phe Val Ser Thr Ser Tyr Phe Ser Cys Asp Lys Lys
340 345 350
Asn Ile Ala Asp Leu Leu Phe Ile Ser Ala Arg Gln Glu Glu Val Arg
355 360 365
Lys Asn Asp Tyr Leu Phe Asn Ile Gln Tyr Gln Leu Arg Asp Met Met
370 375 380
Ile Lys Thr Gly Phe Ile Gln Glu Glu Lys Thr His Phe Tyr Ser Asp
385 390 395 400
Ile Pro Ile Phe Ile Ser
405
<210> 53
<211> 300
<212> PRT
<213> Haemophilus somnus
<400> 53
Met Lys Tyr Asn Ile Lys Ile Lys Ala Ile Val Ile Val Ser Ser Leu
1 5 10 15
Arg Met Leu Leu Ile Phe Leu Met Leu Asn Lys Tyr His Leu Asp Glu
20 25 30
Val Leu Phe Val Phe Asn Glu Gly Phe Glu Leu His Lys Lys Tyr Lys
35 40 45
Ile Lys His Tyr Val Ala Ile Lys Lys Lys Ile Thr Lys Phe Trp Arg
50 55 60
Leu Tyr Tyr Lys Leu Tyr Phe Tyr Arg Phe Lys Ile Asp Arg Ile Pro
65 70 75 80
Val Tyr Gly Ala Asp His Leu Gly Trp Thr Asp Tyr Phe Leu Lys Tyr
85 90 95
Phe Asp Phe Tyr Leu Ile Glu Asp Gly Ile Ala Asn Phe Ser Pro Lys
100 105 110
Arg Tyr Glu Ile Asn Leu Thr Arg Asn Ile Pro Val Phe Gly Phe His
115 120 125
Lys Thr Val Lys Lys Ile Tyr Leu Thr Ser Leu Glu Asn Val Pro Ser
130 135 140
Asp Ile Arg His Lys Val Glu Leu Ile Ser Leu Glu His Leu Trp Lys
145 150 155 160
Thr Arg Thr Ala Gln Glu Gln His Asn Ile Leu Asp Phe Phe Ala Phe
165 170 175
Asn Leu Asp Ser Leu Ile Ser Leu Lys Met Lys Lys Tyr Ile Leu Phe
180 185 190
Thr Gln Cys Leu Ser Glu Asp Arg Val Ile Ser Glu Gln Glu Lys Ile
195 200 205
Ala Ile Tyr Gln His Ile Ile Lys Asn Tyr Asp Glu Arg Leu Leu Val
210 215 220
Ile Lys Pro His Pro Arg Glu Thr Thr Asp Tyr Gln Lys Tyr Phe Glu
225 230 235 240
Asn Val Phe Val Tyr Gln Asp Val Val Pro Ser Glu Leu Phe Glu Leu
245 250 255
Leu Asp Val Asn Phe Glu Arg Val Ile Thr Leu Phe Ser Thr Ala Val
260 265 270
Phe Lys Tyr Asp Arg Asn Ile Val Asp Phe Tyr Gly Thr Arg Ile His
275 280 285
Asp Lys Ile Tyr Gln Trp Phe Gly Asp Ile Lys Phe
290 295 300
<210> 54
<211> 381
<212> PRT
<213> Vibrio harveyi
<400> 54
Met Asp Ser Ser Pro Glu Asn Thr Ser Ser Thr Leu Glu Ile Tyr Ile
1 5 10 15
Asp Ser Ala Thr Leu Pro Ser Leu Gln His Met Val Lys Ile Ile Asp
20 25 30
Glu Gln Ser Gly Asn Lys Lys Leu Ile Asn Trp Lys Arg Tyr Pro Ile
35 40 45
Asp Asp Glu Leu Leu Leu Asp Lys Ile Asn Ala Leu Ser Phe Ser Asp
50 55 60
Thr Thr Asp Leu Thr Arg Tyr Met Glu Ser Ile Leu Leu Ile Gly Asp
65 70 75 80
Ile Lys Arg Val Val Ile Asn Gly Asn Ser Leu Ser Asn Tyr Asn Ile
85 90 95
Val Gly Val Met Arg Ser Ile Asn Ala Leu Gly Leu Asp Leu Asp Val
100 105 110
Glu Ile Asn Phe Tyr Asp Asp Gly Ser Ala Glu Tyr Val Arg Leu Tyr
115 120 125
Asn Phe Ser Gln Leu Pro Glu Ala Glu Arg Glu Leu Leu Val Ser Met
130 135 140
Ser Lys Asn Asn Ile Leu Ala Ala Val Asn Gly Ile Gly Ser Tyr Asp
145 150 155 160
Ser Gly Ser Pro Glu Asn Ile Tyr Gly Phe Ala Gln Ile Tyr Pro Ala
165 170 175
Thr Tyr His Met Leu Arg Ala Asp Ile Phe Asp Thr Asp Leu Glu Ile
180 185 190
Gly Leu Ile Arg Asp Ile Leu Gly Asp Asn Val Lys Gln Met Lys Trp
195 200 205
Gly Gln Phe Leu Gly Phe Asn Glu Glu Gln Lys Glu Leu Phe Tyr Gln
210 215 220
Leu Thr Ser Phe Asn Pro Asp Lys Ile Gln Ala Gln Tyr Lys Glu Ser
225 230 235 240
Pro Asn Lys Asn Phe Val Phe Val Gly Thr Asn Ser Arg Ser Ala Thr
245 250 255
Ala Glu Gln Gln Ile Asn Ile Ile Lys Glu Ala Lys Lys Leu Asp Ser
260 265 270
Glu Ile Ile Pro Asn Ser Ile Asp Gly Tyr Asp Leu Phe Phe Lys Gly
275 280 285
His Pro Ser Ala Thr Tyr Asn Gln Gln Ile Val Asp Ala His Asp Met
290 295 300
Thr Glu Ile Tyr Asn Arg Thr Pro Phe Glu Val Leu Ala Met Thr Ser
305 310 315 320
Ser Leu Pro Asp Ala Val Gly Gly Met Gly Ser Ser Leu Phe Phe Ser
325 330 335
Leu Pro Lys Thr Val Glu Thr Lys Phe Ile Phe Tyr Lys Ser Gly Thr
340 345 350
Asp Ile Glu Ser Asn Ala Leu Ile Gln Val Met Leu Lys Leu Gly Ile
355 360 365
Ile Thr Asp Glu Lys Val Arg Phe Thr Thr Asp Ile Lys
370 375 380
<210> 55
<211> 483
<212> PRT
<213> Alistipes sp.
<400> 55
Met Ala Ser Cys Ser Asp Asp Asp Lys Glu Gln Thr Gly Phe Gln Ile
1 5 10 15
Asp Asp Gly Ser Gly Phe Leu Ser Leu Asp Ala Ala Ala Arg Ser Gly
20 25 30
Ser Ile Ala Ile Thr Ala Asn Asn Ser Trp Ser Val Thr Gln Asp Lys
35 40 45
Asp Ser Glu Trp Leu Thr Leu Ser Thr Thr Ser Gly Ala Ala Gly Arg
50 55 60
Thr Glu Ile Gly Ile Met Leu Glu Ala Asn Pro Gly Glu Ala Arg Asn
65 70 75 80
Ala Gly Leu Thr Phe Asn Ser Gly Gly Arg Thr Tyr Pro Phe Val Ile
85 90 95
Thr Gln Ser Ala His Val Thr Ala Asp Phe Asp Asp Ala Asp His Cys
100 105 110
Phe Tyr Ile Thr Phe Gly Thr Leu Pro Thr Leu Tyr Ala Gly Leu His
115 120 125
Val Leu Ser His Asp Lys Pro Ser Tyr Val Phe Phe Gln Arg Ser Gln
130 135 140
Thr Phe Arg Pro Glu Glu Phe Pro Ala His Ala Glu Val Thr Ile Ala
145 150 155 160
Ala Asp Pro Ser Ala Asn Ala Thr Asp Glu Asp Met Glu Arg Met Arg
165 170 175
Thr Ala Met Lys Gln Gln Ile Leu Lys Ile Asn Val Glu Asp Pro Thr
180 185 190
Ala Val Phe Gly Leu Tyr Val Asp Asp Leu Arg Cys Gly Ile Gly Tyr
195 200 205
Asp Trp Phe Val Ala Gln Gly Ile Asp Ser Thr Arg Val Lys Val Ser
210 215 220
Met Leu Ser Asp Gly Thr Gly Thr Tyr Asn Asn Phe Tyr Asn Tyr Phe
225 230 235 240
Gly Asp Pro Ala Thr Ala Glu Gln Asn Trp Glu Asn Tyr Ala Ala Gln
245 250 255
Val Glu Ala Leu Asp Trp Gln His Gly Gly Arg Phe Pro Glu Thr Arg
260 265 270
Met Pro Asp Gly Phe Asp Phe Tyr Glu Trp Pro Tyr Tyr Leu Ala Thr
275 280 285
Arg Pro Asn Tyr Arg Leu Val Leu Gln Asp Asp Asp Leu Leu Glu Ala
290 295 300
Thr Ser Pro Phe Met Thr Glu Arg Leu Gln Gln Met Arg Thr Glu Ser
305 310 315 320
Lys Gln Pro Tyr Glu Leu Leu Ala Ser Leu Pro Ala Glu Ala Arg Gln
325 330 335
Arg Phe Phe Arg Met Ala Gly Phe Asp Tyr Asp Ala Phe Ala Ala Leu
340 345 350
Phe Asp Ala Ser Pro Lys Lys Asn Leu Val Ile Ile Gly Thr Ser His
355 360 365
Thr Ser Glu Glu Ser Glu Ala Gln Gln Ala Ala Tyr Val Glu Arg Ile
370 375 380
Ile Gly Asp Tyr Gly Thr Ala Tyr Asp Ile Phe Phe Lys Pro His Pro
385 390 395 400
Ala Asp Ser Ser Ser Ser Asn Tyr Glu Glu Arg Phe Glu Gly Leu Thr
405 410 415
Leu Leu Pro Gly Gln Met Pro Phe Glu Ile Phe Val Trp Ser Leu Leu
420 425 430
Asp Lys Val Asp Leu Ile Gly Gly Tyr Ser Ser Thr Val Phe Leu Thr
435 440 445
Val Pro Val Glu Lys Thr Gly Phe Ile Phe Ala Ala Asn Ala Glu Ser
450 455 460
Leu Pro Arg Pro Leu Asn Val Leu Phe Arg Asn Ala Glu His Val Arg
465 470 475 480
Trp Ile Gln
<210> 56
<211> 483
<212> PRT
<213> Alistipes shahii
<400> 56
Met Asp Asp Gly Thr Pro Ser Val Ser Ile Asn Gly Gly Thr Asp Phe
1 5 10 15
Leu Ser Leu Asp His Leu Ala Arg Ser Gly Lys Ile Thr Val Asn Ala
20 25 30
Pro Ala Pro Trp Ser Val Thr Leu Ala Pro Glu Asn Tyr Gly Gln Asp
35 40 45
Glu Lys Pro Asp Trp Leu Thr Leu Ser Ala Glu Glu Gly Pro Ala Gly
50 55 60
Tyr Ser Glu Ile Asp Val Thr Phe Ala Glu Asn Pro Gly Pro Ala Arg
65 70 75 80
Ser Ala Ser Leu Leu Phe Ser Cys Asp Gly Lys Thr Leu Ala Phe Thr
85 90 95
Val Ser Gln Ser Ala Gly Gly Thr Gly Phe Asp Ala Pro Asp Tyr Tyr
100 105 110
Phe Tyr Ile Ser Val Gly Thr Met Pro Thr Leu Tyr Ser Gly Leu His
115 120 125
Leu Leu Ser His Asp Lys Pro Ser Tyr Val Ser Tyr Glu Arg Ala Ser
130 135 140
Thr Phe Asp Ala Ala Glu Phe Pro Asp Arg Ala Phe Val Tyr Pro Val
145 150 155 160
Ala Asp Pro Thr Gly His Ala Thr Asn Glu Glu Leu Arg Ala Met Ser
165 170 175
Glu Ala Met Lys Arg Arg Ile Leu Glu Ile Asn Ala Glu Asp Pro Thr
180 185 190
Ala Val Phe Gly Leu Trp Val Asp Asp Leu Arg Cys Arg Leu Gly Tyr
195 200 205
Asp Trp Phe Val Ala Gln Gly Ile Asp Ser Ala Arg Val Lys Val Thr
210 215 220
Met Leu Ser Asp Gly Thr Ala Thr Tyr Asn Asn Phe His Asn Tyr Phe
225 230 235 240
Gly Asp Ala Ala Thr Ala Glu Gln Asn Trp Asn Asp Tyr Ala Ala Glu
245 250 255
Val Glu Ala Leu Asp Trp Asn His Gly Gly Arg Tyr Pro Glu Thr Arg
260 265 270
Ala Pro Glu Glu Phe Ala Ser Tyr Thr Trp Pro Tyr Tyr Leu Ser Thr
275 280 285
Arg Pro Asp Tyr Arg Leu Met Leu Gln Asn Ser Ser Leu Met Glu Ser
290 295 300
Ser Cys Pro Phe Ile Ala Asp Arg Leu Ala Ala Met Lys Met Glu Ser
305 310 315 320
Val Gln Pro Tyr Glu Leu Leu Thr Ala Leu Pro Glu Ala Ser Lys Gln
325 330 335
Gln Phe Tyr Arg Met Ala Lys Phe Asp Tyr Ala Arg Phe Ala Gly Leu
340 345 350
Phe Asp Leu Ser Pro Lys Lys Asn Leu Ile Ile Ile Gly Thr Ser His
355 360 365
Ser Ser Ala Ala Ser Glu Gln Gln Gln Ala Ala Tyr Val Glu Arg Ile
370 375 380
Ile Gln Gln Tyr Gly Ser Asp Tyr Asp Ile Phe Phe Lys Pro His Pro
385 390 395 400
Ala Asp Ser Ser Ser Ala Gly Tyr Pro Asp Arg Phe Glu Gly Leu Thr
405 410 415
Leu Leu Pro Gly Gln Met Pro Phe Glu Ile Phe Val Trp Ala Leu Leu
420 425 430
Asp Lys Ile Asp Met Ile Gly Gly Tyr Pro Ser Thr Thr Phe Ile Ser
435 440 445
Val Pro Leu Asp Lys Val Gly Phe Leu Phe Ala Ala Asp Ala Asp Gly
450 455 460
Leu Val Arg Pro Leu Asn Ile Leu Phe Arg Asp Ala Ala Asn Val Glu
465 470 475 480
Trp Ile Gln
<210> 57
<211> 401
<212> PRT
<213> Actinobacillus suis
<400> 57
Met Glu Arg Thr Pro Gln Leu Gln Ala Val Asp Ile Tyr Ile Asp Phe
1 5 10 15
Ala Thr Ile Pro Ser Leu Ser Tyr Phe Leu His Phe Leu Lys His Lys
20 25 30
His Asp Asp Gln Arg Leu Arg Leu Phe Ser Leu Ala Arg Phe Glu Met
35 40 45
Pro Gln Thr Leu Ile Glu Gln Tyr Glu Gly Ile Ile Gln Phe Ser Arg
50 55 60
Asn Val Glu His Asn Val Glu Pro Leu Leu Glu Gln Leu Gln Thr Ile
65 70 75 80
Leu Ser Gln Glu Gly Lys Gln Phe Glu Leu His Leu His Leu Asn Leu
85 90 95
Phe His Ser Phe Glu Met Phe Leu Asn Leu Ser Pro Thr Tyr Thr Gln
100 105 110
Tyr Lys Glu Lys Ile Ser Lys Ile Val Leu His Leu Tyr Asp Asp Gly
115 120 125
Ser Glu Gly Val Met Lys Gln Tyr Gln Leu Gln Lys Ser Ser Ser Leu
130 135 140
Val Gln Asp Leu Ala Ala Thr Lys Ala Ser Leu Val Ser Leu Phe Glu
145 150 155 160
Asn Gly Glu Gly Ser Phe Ser Gln Ile Asp Leu Ile Arg Tyr Val Trp
165 170 175
Asn Ala Val Leu Glu Thr His Tyr Tyr Leu Leu Ser Asp His Phe Leu
180 185 190
Leu Asp Glu Lys Leu Gln Pro Leu Lys Ala Glu Leu Gly His Tyr Gln
195 200 205
Leu Leu Asn Leu Ser Ala Tyr Gln Tyr Leu Ser Ser Glu Asp Leu Leu
210 215 220
Trp Leu Lys Gln Ile Leu Lys Ile Asp Thr Glu Leu Glu Ser Leu Met
225 230 235 240
Gln Lys Leu Thr Ala Gln Pro Val Tyr Phe Phe Ser Gly Thr Thr Phe
245 250 255
Phe Asn Ile Ser Phe Glu Asp Lys Gln Arg Leu Ala Asn Ile His Ala
260 265 270
Ile Leu Ile Arg Glu His Leu Asp Pro Asn Ser Gln Leu Phe Ile Gly
275 280 285
Glu Pro Tyr Leu Phe Val Phe Lys Gly His Pro Asn Ser Pro Glu Ile
290 295 300
Asn Gln Ala Leu Arg Glu Tyr Tyr Pro Asn Val Ile Phe Leu Pro Glu
305 310 315 320
Asn Ile Pro Phe Glu Ile Leu Thr Leu Leu Gly Phe Ser Pro Gln Lys
325 330 335
Ile Gly Gly Phe Ala Ser Thr Ile His Val Asn Ser Glu Gln Ser Lys
340 345 350
Leu Ala Lys Leu Phe Phe Leu Thr Ser Thr Asp Glu Gln Glu Arg Gln
355 360 365
Leu Ser Asp Gly Tyr Ile Lys Gln Tyr Ala Leu Ala Gln Ala Met Leu
370 375 380
Glu Met Gln Leu Val Ser Gln Glu Gln Val Tyr Tyr Cys Ser Leu Ser
385 390 395 400
Ser
<210> 58
<211> 401
<212> PRT
<213> Actinobacillus capsulatus
<400> 58
Met Glu Arg Ile Pro Gln Leu Gln Ala Val Asp Ile Tyr Ile Asp Phe
1 5 10 15
Ala Thr Ile Pro Ser Leu Ser Tyr Phe Leu His Phe Leu Lys His Lys
20 25 30
His Asp His Gln Arg Leu Arg Leu Phe Ser Leu Ala Arg Phe Glu Met
35 40 45
Pro Gln Thr Val Ile Glu Gln Tyr Glu Gly Ile Ile Gln Phe Ser Arg
50 55 60
Asn Val Glu His Asn Val Glu Gln Leu Leu Glu Gln Leu Gln Thr Ile
65 70 75 80
Leu Ser Gln Glu Gly Lys Gln Phe Glu Leu His Leu His Leu Asn Leu
85 90 95
Phe His Ser Phe Glu Met Phe Leu Asn Leu Ser Pro Thr Tyr Thr Lys
100 105 110
Tyr Lys Glu Lys Ile Ser Lys Ile Val Leu His Leu Tyr Asp Asp Gly
115 120 125
Ser Glu Gly Val Met Lys Gln Tyr Gln Leu Gln Gln Ser Asn Ser Leu
130 135 140
Ala Gln Asp Leu Ala Ser Thr Lys Ala Ser Leu Val Ser Leu Phe Lys
145 150 155 160
Asn Gly Glu Gly Ala Phe Ser Gln Ile Asp Leu Ile Arg Tyr Val Trp
165 170 175
Asn Ala Val Leu Glu Thr His Tyr Tyr Leu Leu Ser Asp His Phe Leu
180 185 190
Ala His Glu Lys Leu Gln Pro Leu Lys Ile Glu Leu Gly His Tyr Gln
195 200 205
Leu Leu Asn Leu Ser Ala Tyr Gln Tyr Leu Ser Ser Glu Asp Leu Leu
210 215 220
Trp Leu Lys Gln Ile Leu Lys Ile Asp Ala Glu Leu Glu Ser Leu Met
225 230 235 240
His Lys Leu Thr Thr Gln Pro Val Tyr Phe Phe Ser Gly Thr Thr Phe
245 250 255
Phe Asn Ile Ser Phe Glu Asp Lys Gln Arg Leu Ala Asn Ile His Ala
260 265 270
Ile Leu Ile Arg Glu His Leu Asp Pro Asn Ser Gln Leu Phe Ile Gly
275 280 285
Glu Pro Tyr Leu Phe Val Phe Lys Gly His Pro Asn Ser Pro Glu Ile
290 295 300
Asn Gln Ala Leu Arg Glu Tyr Tyr Pro Asn Ala Ile Phe Leu Pro Glu
305 310 315 320
Asn Ile Pro Phe Glu Ile Leu Thr Leu Leu Gly Phe Ser Pro Gln Lys
325 330 335
Ile Gly Gly Phe Ala Ser Thr Ile His Val Asn Ser Glu Gln Ser Lys
340 345 350
Leu Ala Lys Leu Phe Phe Leu Thr Ser Thr Asp Glu Gln Glu Arg Asn
355 360 365
Arg Ser Asp Gly Tyr Ile Lys Gln Tyr Ala Leu Ala Gln Ala Met Leu
370 375 380
Glu Met Gln Leu Val Ser Gln Glu Gln Val Tyr Tyr Cys Ser Leu Ser
385 390 395 400
Ser
<210> 59
<211> 311
<212> PRT
<213> Haemophilus somnus
<400> 59
Met Phe Arg Glu Asp Asn Met Asn Leu Ile Ile Cys Cys Thr Pro Leu
1 5 10 15
Gln Val Ile Ile Ala Glu Lys Ile Ile Glu Arg Tyr Pro Glu Gln Lys
20 25 30
Phe Tyr Gly Val Met Leu Glu Ser Phe Tyr Asn Asp Lys Phe Asp Phe
35 40 45
Tyr Glu Asn Lys Leu Lys His Leu Cys His Glu Phe Phe Cys Ile Lys
50 55 60
Ile Ala Arg Phe Lys Leu Glu Arg Tyr Lys Asn Leu Leu Ser Leu Leu
65 70 75 80
Lys Ile Lys Asn Lys Thr Phe Asp Arg Val Phe Leu Ala Asn Ile Glu
85 90 95
Lys Arg Tyr Ile His Ile Ile Leu Ser Asn Ile Phe Phe Lys Glu Leu
100 105 110
Tyr Thr Phe Asp Asp Gly Thr Ala Asn Ile Ala Pro Asn Ser His Leu
115 120 125
Tyr Gln Glu Tyr Asp His Ser Leu Lys Lys Arg Ile Thr Asp Ile Leu
130 135 140
Leu Pro Asn His Tyr Asn Ser Asn Lys Val Lys Asn Ile Ser Lys Leu
145 150 155 160
His Tyr Ser Ile Tyr Arg Cys Lys Asn Asn Ile Ile Asp Asn Ile Glu
165 170 175
Tyr Met Pro Leu Phe Asn Leu Glu Lys Lys Tyr Thr Ala Gln Asp Lys
180 185 190
Ser Ile Ser Ile Leu Leu Gly Gln Pro Ile Phe Tyr Asp Glu Glu Lys
195 200 205
Asn Ile Arg Leu Ile Lys Glu Val Ile Ala Lys Phe Lys Ile Asp Tyr
210 215 220
Tyr Phe Pro His Pro Arg Glu Asp Tyr Tyr Ile Asp Asn Val Ser Tyr
225 230 235 240
Ile Lys Thr Pro Leu Ile Phe Glu Glu Phe Tyr Ala Glu Arg Ser Ile
245 250 255
Glu Asn Ser Ile Lys Ile Tyr Thr Phe Phe Ser Ser Ala Val Leu Asn
260 265 270
Ile Val Thr Lys Glu Asn Ile Asp Arg Ile Tyr Ala Leu Lys Pro Lys
275 280 285
Leu Thr Glu Lys Ala Tyr Leu Asp Cys Tyr Asp Ile Leu Lys Asp Phe
290 295 300
Gly Ile Lys Val Ile Asp Ile
305 310
<210> 60
<211> 399
<212> PRT
<213> Haemophilus ducreyi
<400> 60
Met Leu Ile Gln Gln Asn Leu Glu Ile Tyr Leu Asp Tyr Ala Thr Ile
1 5 10 15
Pro Ser Leu Ala Cys Phe Met His Phe Ile Gln His Lys Asp Asp Val
20 25 30
Asp Ser Ile Arg Leu Phe Gly Leu Ala Arg Phe Asp Ile Pro Gln Ser
35 40 45
Ile Ile Asp Arg Tyr Pro Ala Asn His Leu Phe Tyr His Asn Ile Asp
50 55 60
Asn Arg Asp Leu Thr Ala Val Leu Asn Gln Leu Ala Asp Ile Leu Ala
65 70 75 80
Gln Glu Asn Lys Arg Phe Gln Ile Asn Leu His Leu Asn Leu Phe His
85 90 95
Ser Ile Asp Leu Phe Phe Ala Ile Tyr Pro Ile Tyr Gln Gln Tyr Gln
100 105 110
His Lys Ile Ser Thr Ile Gln Leu Gln Leu Tyr Asp Asp Gly Ser Glu
115 120 125
Gly Ile Val Thr Gln His Ser Leu Cys Lys Ile Ala Asp Leu Glu Gln
130 135 140
Leu Ile Leu Gln His Lys Asn Val Leu Leu Glu Leu Leu Thr Lys Gly
145 150 155 160
Thr Ala Asn Val Pro Asn Pro Thr Leu Leu Arg Tyr Leu Trp Asn Asn
165 170 175
Ile Ile Asp Ser Gln Phe His Leu Ile Ser Asp His Phe Leu Gln His
180 185 190
Pro Lys Leu Gln Pro Leu Lys Arg Leu Leu Lys Arg Tyr Thr Ile Leu
195 200 205
Asp Phe Thr Cys Tyr Pro Arg Phe Asn Ala Glu Gln Lys Gln Leu Leu
210 215 220
Lys Glu Ile Leu His Ile Ser Asn Glu Leu Glu Asn Leu Leu Lys Leu
225 230 235 240
Leu Lys Gln His Asn Thr Phe Leu Phe Thr Gly Thr Thr Ala Phe Asn
245 250 255
Leu Asp Gln Glu Lys Leu Asp Leu Leu Thr Gln Leu His Ile Leu Leu
260 265 270
Leu Asn Glu His Gln Asn Pro His Ser Thr His Tyr Ile Gly Asn Asn
275 280 285
Tyr Leu Leu Leu Ile Lys Gly His Ala Asn Ser Pro Ala Leu Asn His
290 295 300
Thr Leu Ala Leu His Phe Pro Asp Ala Ile Phe Leu Pro Ala Asn Ile
305 310 315 320
Pro Phe Glu Ile Phe Ala Met Leu Gly Phe Thr Pro Asn Lys Met Gly
325 330 335
Gly Phe Ala Ser Thr Ser Tyr Ile Asn Tyr Pro Thr Glu Asn Ile Asn
340 345 350
His Leu Phe Phe Leu Thr Ser Asp Gln Pro Ser Ile Arg Thr Lys Trp
355 360 365
Leu Asp Tyr Glu Lys Gln Phe Gly Leu Met Tyr Ser Leu Leu Ala Met
370 375 380
Gln Lys Ile Asn Glu Asp Gln Ala Phe Met Cys Thr Ile His Asn
385 390 395
<210> 61
<211> 497
<212> PRT
<213> Photobacterium leiognathi
<400> 61
Met Cys Asn Asp Asn Gln Asn Thr Val Asp Val Val Val Ser Thr Val
1 5 10 15
Asn Asp Asn Val Ile Glu Asn Asn Thr Tyr Gln Val Lys Pro Ile Asp
20 25 30
Thr Pro Thr Thr Phe Asp Ser Tyr Ser Trp Ile Gln Thr Cys Gly Thr
35 40 45
Pro Ile Leu Lys Asp Asp Glu Lys Tyr Ser Leu Ser Phe Asp Phe Val
50 55 60
Ala Pro Glu Leu Asp Gln Asp Glu Lys Phe Cys Phe Glu Phe Thr Gly
65 70 75 80
Asp Val Asp Gly Lys Arg Tyr Val Thr Gln Thr Asn Leu Thr Val Val
85 90 95
Ala Pro Thr Leu Glu Val Tyr Val Asp His Ala Ser Leu Pro Ser Leu
100 105 110
Gln Gln Leu Met Lys Ile Ile Gln Gln Lys Asn Glu Tyr Ser Gln Asn
115 120 125
Glu Arg Phe Ile Ser Trp Gly Arg Ile Gly Leu Thr Glu Asp Asn Ala
130 135 140
Glu Lys Leu Asn Ala His Ile Tyr Pro Leu Ala Gly Asn Asn Thr Ser
145 150 155 160
Gln Glu Leu Val Asp Ala Val Ile Asp Tyr Ala Asp Ser Lys Asn Arg
165 170 175
Leu Asn Leu Glu Leu Asn Thr Asn Thr Ala His Ser Phe Pro Asn Leu
180 185 190
Ala Pro Ile Leu Arg Ile Ile Ser Ser Lys Ser Asn Ile Leu Ile Ser
195 200 205
Asn Ile Asn Leu Tyr Asp Asp Gly Ser Ala Glu Tyr Val Asn Leu Tyr
210 215 220
Asn Trp Lys Asp Thr Glu Asp Lys Ser Val Lys Leu Ser Asp Ser Phe
225 230 235 240
Leu Val Leu Lys Asp Tyr Phe Asn Gly Ile Ser Ser Glu Lys Pro Ser
245 250 255
Gly Ile Tyr Gly Arg Tyr Asn Trp His Gln Leu Tyr Asn Thr Ser Tyr
260 265 270
Tyr Phe Leu Arg Lys Asp Tyr Leu Thr Val Glu Pro Gln Leu His Asp
275 280 285
Leu Arg Glu Tyr Leu Gly Gly Ser Leu Lys Gln Met Ser Trp Asp Gly
290 295 300
Phe Ser Gln Leu Ser Lys Gly Asp Lys Glu Leu Phe Leu Asn Ile Val
305 310 315 320
Gly Phe Asp Gln Glu Lys Leu Gln Gln Glu Tyr Gln Gln Ser Glu Leu
325 330 335
Pro Asn Phe Val Phe Thr Gly Thr Thr Thr Trp Ala Gly Gly Glu Thr
340 345 350
Lys Glu Tyr Tyr Ala Gln Gln Gln Val Asn Val Val Asn Asn Ala Ile
355 360 365
Asn Glu Thr Ser Pro Tyr Tyr Leu Gly Arg Glu His Asp Leu Phe Phe
370 375 380
Lys Gly His Pro Arg Gly Gly Ile Ile Asn Asp Ile Ile Leu Gly Ser
385 390 395 400
Phe Asn Asn Met Ile Asp Ile Pro Ala Lys Val Ser Phe Glu Val Leu
405 410 415
Met Met Thr Gly Met Leu Pro Asp Thr Val Gly Gly Ile Ala Ser Ser
420 425 430
Leu Tyr Phe Ser Ile Pro Ala Glu Lys Val Ser Phe Ile Val Phe Thr
435 440 445
Ser Ser Asp Thr Ile Thr Asp Arg Glu Asp Ala Leu Lys Ser Pro Leu
450 455 460
Val Gln Val Met Met Thr Leu Gly Ile Val Lys Glu Lys Asp Val Leu
465 470 475 480
Phe Trp Ser Asp Leu Pro Asp Cys Ser Ser Gly Val Cys Ile Ala Gln
485 490 495
Tyr
<210> 62
<211> 498
<212> PRT
<213> Photobacterium sp.
<400> 62
Met Ser Glu Glu Asn Thr Gln Ser Ile Ile Lys Asn Asp Ile Asn Lys
1 5 10 15
Thr Ile Ile Asp Glu Glu Tyr Val Asn Leu Glu Pro Ile Asn Gln Ser
20 25 30
Asn Ile Ser Phe Thr Lys His Ser Trp Val Gln Thr Cys Gly Thr Gln
35 40 45
Gln Leu Leu Thr Glu Gln Asn Lys Glu Ser Ile Ser Leu Ser Val Val
50 55 60
Ala Pro Arg Leu Asp Asp Asp Glu Lys Tyr Cys Phe Asp Phe Asn Gly
65 70 75 80
Val Ser Asn Lys Gly Glu Lys Tyr Ile Thr Lys Val Thr Leu Asn Val
85 90 95
Val Ala Pro Ser Leu Glu Val Tyr Val Asp His Ala Ser Leu Pro Thr
100 105 110
Leu Gln Gln Leu Met Asp Ile Ile Lys Ser Glu Glu Glu Asn Pro Thr
115 120 125
Ala Gln Arg Tyr Ile Ala Trp Gly Arg Ile Val Pro Thr Asp Glu Gln
130 135 140
Met Lys Glu Leu Asn Ile Thr Ser Phe Ala Leu Ile Asn Asn His Thr
145 150 155 160
Pro Ala Asp Leu Val Gln Glu Ile Val Lys Gln Ala Gln Thr Lys His
165 170 175
Arg Leu Asn Val Lys Leu Ser Ser Asn Thr Ala His Ser Phe Asp Asn
180 185 190
Leu Val Pro Ile Leu Lys Glu Leu Asn Ser Phe Asn Asn Val Thr Val
195 200 205
Thr Asn Ile Asp Leu Tyr Asp Asp Gly Ser Ala Glu Tyr Val Asn Leu
210 215 220
Tyr Asn Trp Arg Asp Thr Leu Asn Lys Thr Asp Asn Leu Lys Ile Gly
225 230 235 240
Lys Asp Tyr Leu Glu Asp Val Ile Asn Gly Ile Asn Glu Asp Thr Ser
245 250 255
Asn Thr Gly Thr Ser Ser Val Tyr Asn Trp Gln Lys Leu Tyr Pro Ala
260 265 270
Asn Tyr His Phe Leu Arg Lys Asp Tyr Leu Thr Leu Glu Pro Ser Leu
275 280 285
His Glu Leu Arg Asp Tyr Ile Gly Asp Ser Leu Lys Gln Met Gln Trp
290 295 300
Asp Gly Phe Lys Lys Phe Asn Ser Lys Gln Gln Glu Leu Phe Leu Ser
305 310 315 320
Ile Val Asn Phe Asp Lys Gln Lys Leu Gln Asn Glu Tyr Asn Ser Ser
325 330 335
Asn Leu Pro Asn Phe Val Phe Thr Gly Thr Thr Val Trp Ala Gly Asn
340 345 350
His Glu Arg Glu Tyr Tyr Ala Lys Gln Gln Ile Asn Val Ile Asn Asn
355 360 365
Ala Ile Asn Glu Ser Ser Pro His Tyr Leu Gly Asn Ser Tyr Asp Leu
370 375 380
Phe Phe Lys Gly His Pro Gly Gly Gly Ile Ile Asn Thr Leu Ile Met
385 390 395 400
Gln Asn Tyr Pro Ser Met Val Asp Ile Pro Ser Lys Ile Ser Phe Glu
405 410 415
Val Leu Met Met Thr Asp Met Leu Pro Asp Ala Val Ala Gly Ile Ala
420 425 430
Ser Ser Leu Tyr Phe Thr Ile Pro Ala Glu Lys Ile Lys Phe Ile Val
435 440 445
Phe Thr Ser Thr Glu Thr Ile Thr Asp Arg Glu Thr Ala Leu Arg Ser
450 455 460
Pro Leu Val Gln Val Met Ile Lys Leu Gly Ile Val Lys Glu Glu Asn
465 470 475 480
Val Leu Phe Trp Ala Asp Leu Pro Asn Cys Glu Thr Gly Val Cys Ile
485 490 495
Ala Val
<210> 63
<211> 482
<212> PRT
<213> Photobacterium leiognathi
<400> 63
Met Asn Asp Asn Gln Asn Thr Val Asp Val Val Val Ser Thr Val Asn
1 5 10 15
Asp Asn Val Ile Glu Asn Asn Thr Tyr Gln Val Lys Pro Ile Asp Thr
20 25 30
Pro Thr Thr Phe Asp Ser Tyr Ser Trp Ile Gln Thr Cys Gly Thr Pro
35 40 45
Ile Leu Lys Asp Asp Glu Lys Tyr Ser Leu Ser Phe Asp Phe Val Ala
50 55 60
Pro Glu Leu Asp Gln Asp Glu Lys Phe Cys Phe Glu Phe Thr Gly Asp
65 70 75 80
Val Asp Gly Lys Arg Tyr Val Thr Gln Thr Asn Leu Thr Val Val Ala
85 90 95
Pro Thr Leu Glu Val Tyr Val Asp His Ala Ser Leu Pro Ser Leu Gln
100 105 110
Gln Leu Met Lys Ile Ile Gln Gln Lys Asn Glu Tyr Ser Gln Asn Glu
115 120 125
Arg Phe Ile Ser Trp Gly Arg Ile Arg Leu Thr Glu Asp Asn Ala Glu
130 135 140
Lys Leu Asn Ala His Ile Tyr Pro Leu Ala Gly Asn Asn Thr Ser Gln
145 150 155 160
Glu Leu Val Asp Ala Val Ile Asp Tyr Ala Asp Ser Lys Asn Arg Leu
165 170 175
Asn Leu Glu Leu Asn Thr Asn Thr Gly His Ser Phe Arg Asn Ile Ala
180 185 190
Pro Ile Leu Arg Ala Thr Ser Ser Lys Asn Asn Ile Leu Ile Ser Asn
195 200 205
Ile Asn Leu Tyr Asp Asp Gly Ser Ala Glu Tyr Val Ser Leu Tyr Asn
210 215 220
Trp Lys Asp Thr Asp Asn Lys Ser Gln Lys Leu Ser Asp Ser Phe Leu
225 230 235 240
Val Leu Lys Asp Tyr Leu Asn Gly Ile Ser Ser Glu Lys Pro Asn Gly
245 250 255
Ile Tyr Ser Ile Tyr Asn Trp His Gln Leu Tyr His Ser Ser Tyr Tyr
260 265 270
Phe Leu Arg Lys Asp Tyr Leu Thr Val Glu Thr Lys Leu His Asp Leu
275 280 285
Arg Glu Tyr Leu Gly Gly Ser Leu Lys Gln Met Ser Trp Asp Thr Phe
290 295 300
Ser Gln Leu Ser Lys Gly Asp Lys Glu Leu Phe Leu Asn Ile Val Gly
305 310 315 320
Phe Asp Gln Glu Lys Leu Gln Gln Glu Tyr Gln Gln Ser Glu Leu Pro
325 330 335
Asn Phe Val Phe Thr Gly Thr Thr Thr Trp Ala Gly Gly Glu Thr Lys
340 345 350
Glu Tyr Tyr Ala Gln Gln Gln Val Asn Val Val Asn Asn Ala Ile Asn
355 360 365
Glu Thr Ser Pro Tyr Tyr Leu Gly Arg Glu His Asp Leu Phe Phe Lys
370 375 380
Gly His Pro Arg Gly Gly Ile Ile Asn Asp Ile Ile Leu Gly Ser Phe
385 390 395 400
Asn Asn Met Ile Asp Ile Pro Ala Lys Val Ser Phe Glu Val Leu Met
405 410 415
Met Thr Gly Met Leu Pro Asp Thr Val Gly Gly Ile Ala Ser Ser Leu
420 425 430
Tyr Phe Ser Ile Pro Ala Glu Lys Val Ser Phe Ile Val Phe Thr Ser
435 440 445
Ser Asp Thr Ile Thr Asp Arg Glu Asp Ala Leu Lys Ser Pro Leu Val
450 455 460
Gln Val Met Met Thr Leu Gly Ile Val Lys Glu Lys Asp Val Leu Phe
465 470 475 480
Trp Cys
<210> 64
<211> 675
<212> PRT
<213> Photobacterium damsela
<400> 64
Met Lys Lys Ile Leu Thr Val Leu Ser Ile Phe Ile Leu Ser Ala Cys
1 5 10 15
Asn Ser Asp Asn Thr Ser Leu Lys Glu Thr Val Ser Ser Asn Ser Ala
20 25 30
Asp Val Val Glu Thr Glu Thr Tyr Gln Leu Thr Pro Ile Asp Ala Pro
35 40 45
Ser Ser Phe Leu Ser His Ser Trp Glu Gln Thr Cys Gly Thr Pro Ile
50 55 60
Leu Asn Glu Ser Asp Lys Gln Ala Ile Ser Phe Asp Phe Val Ala Pro
65 70 75 80
Glu Leu Lys Gln Asp Glu Lys Tyr Cys Phe Thr Phe Lys Gly Ile Thr
85 90 95
Gly Asp His Arg Tyr Ile Thr Asn Thr Thr Leu Thr Val Val Ala Pro
100 105 110
Thr Leu Glu Val Tyr Ile Asp His Ala Ser Leu Pro Ser Leu Gln Gln
115 120 125
Leu Ile His Ile Ile Gln Ala Lys Asp Glu Tyr Pro Ser Asn Gln Arg
130 135 140
Phe Val Ser Trp Lys Arg Val Thr Val Asp Ala Asp Asn Ala Asn Lys
145 150 155 160
Leu Asn Ile His Thr Tyr Pro Leu Lys Gly Asn Asn Thr Ser Pro Glu
165 170 175
Met Val Ala Ala Ile Asp Glu Tyr Ala Gln Ser Lys Asn Arg Leu Asn
180 185 190
Ile Glu Phe Tyr Thr Asn Thr Ala His Val Phe Asn Asn Leu Pro Pro
195 200 205
Ile Ile Gln Pro Leu Tyr Asn Asn Glu Lys Val Lys Ile Ser His Ile
210 215 220
Ser Leu Tyr Asp Asp Gly Ser Ser Glu Tyr Val Ser Leu Tyr Gln Trp
225 230 235 240
Lys Asp Thr Pro Asn Lys Ile Glu Thr Leu Glu Gly Glu Val Ser Leu
245 250 255
Leu Ala Asn Tyr Leu Ala Gly Thr Ser Pro Asp Ala Pro Lys Gly Met
260 265 270
Gly Asn Arg Tyr Asn Trp His Lys Leu Tyr Asp Thr Asp Tyr Tyr Phe
275 280 285
Leu Arg Glu Asp Tyr Leu Asp Val Glu Ala Asn Leu His Asp Leu Arg
290 295 300
Asp Tyr Leu Gly Ser Ser Ala Lys Gln Met Pro Trp Asp Glu Phe Ala
305 310 315 320
Lys Leu Ser Asp Ser Gln Gln Thr Leu Phe Leu Asp Ile Val Gly Phe
325 330 335
Asp Lys Glu Gln Leu Gln Gln Gln Tyr Ser Gln Ser Pro Leu Pro Asn
340 345 350
Phe Ile Phe Thr Gly Thr Thr Thr Trp Ala Gly Gly Glu Thr Lys Glu
355 360 365
Tyr Tyr Ala Gln Gln Gln Val Asn Val Ile Asn Asn Ala Ile Asn Glu
370 375 380
Thr Ser Pro Tyr Tyr Leu Gly Lys Asp Tyr Asp Leu Phe Phe Lys Gly
385 390 395 400
His Pro Ala Gly Gly Val Ile Asn Asp Ile Ile Leu Gly Ser Phe Pro
405 410 415
Asp Met Ile Asn Ile Pro Ala Lys Ile Ser Phe Glu Val Leu Met Met
420 425 430
Thr Asp Met Leu Pro Asp Thr Val Ala Gly Ile Ala Ser Ser Leu Tyr
435 440 445
Phe Thr Ile Pro Ala Asp Lys Val Asn Phe Ile Val Phe Thr Ser Ser
450 455 460
Asp Thr Ile Thr Asp Arg Glu Glu Ala Leu Lys Ser Pro Leu Val Gln
465 470 475 480
Val Met Leu Thr Leu Gly Ile Val Lys Glu Lys Asp Val Leu Phe Trp
485 490 495
Ala Asp His Lys Val Asn Ser Met Glu Val Ala Ile Asp Glu Ala Cys
500 505 510
Thr Arg Ile Ile Ala Lys Arg Gln Pro Thr Ala Ser Asp Leu Arg Leu
515 520 525
Val Ile Ala Ile Ile Lys Thr Ile Thr Asp Leu Glu Arg Ile Gly Asp
530 535 540
Val Ala Glu Ser Ile Ala Lys Val Ala Leu Glu Ser Phe Ser Asn Lys
545 550 555 560
Gln Tyr Asn Leu Leu Val Ser Leu Glu Ser Leu Gly Gln His Thr Val
565 570 575
Arg Met Leu His Glu Val Leu Asp Ala Phe Ala Arg Met Asp Val Lys
580 585 590
Ala Ala Ile Glu Val Tyr Gln Glu Asp Asp Arg Ile Asp Gln Glu Tyr
595 600 605
Glu Ser Ile Val Arg Gln Leu Met Ala His Met Met Glu Asp Pro Ser
610 615 620
Ser Ile Pro Asn Val Met Lys Val Met Trp Ala Ala Arg Ser Ile Glu
625 630 635 640
Arg Val Gly Asp Arg Cys Gln Asn Ile Cys Glu Tyr Ile Ile Tyr Phe
645 650 655
Val Lys Gly Lys Asp Val Arg His Thr Lys Pro Asp Asp Phe Gly Thr
660 665 670
Met Leu Asp
675
<210> 65
<211> 510
<212> PRT
<213> Photobacterium damsela
<400> 65
Met Lys Lys Ile Leu Thr Val Leu Ser Ile Phe Ile Leu Ser Ala Cys
1 5 10 15
Asn Ser Asp Asn Thr Ser Leu Lys Glu Thr Val Ser Ser Asn Ser Ala
20 25 30
Asp Val Val Glu Thr Glu Thr Tyr Gln Leu Thr Pro Ile Asp Ala Pro
35 40 45
Ser Ser Phe Leu Ser His Ser Trp Glu Gln Thr Cys Gly Thr Pro Ile
50 55 60
Leu Asn Glu Ser Asp Lys Gln Ala Ile Ser Phe Asp Phe Val Ala Pro
65 70 75 80
Glu Leu Lys Gln Asp Glu Lys Tyr Cys Phe Thr Phe Lys Gly Ile Thr
85 90 95
Gly Asp His Arg Tyr Ile Thr Asn Thr Thr Leu Thr Val Val Ala Pro
100 105 110
Thr Leu Glu Val Tyr Ile Asp His Ala Ser Leu Pro Ser Leu Gln Gln
115 120 125
Leu Ile His Ile Ile Gln Ala Lys Asp Glu Tyr Pro Ser Asn Gln Arg
130 135 140
Phe Val Ser Trp Lys Arg Val Thr Val Asp Ala Asp Asn Ala Asn Lys
145 150 155 160
Leu Asn Ile His Thr Tyr Pro Leu Lys Gly Asn Asn Thr Ser Pro Glu
165 170 175
Met Val Ala Ala Ile Asp Glu Tyr Ala Gln Ser Lys Asn Arg Leu Asn
180 185 190
Ile Glu Phe Tyr Thr Asn Thr Ala His Val Phe Asn Asn Leu Pro Pro
195 200 205
Ile Ile Gln Pro Leu Tyr Asn Asn Glu Lys Val Lys Ile Ser His Ile
210 215 220
Ser Leu Tyr Asp Asp Gly Ser Ser Glu Tyr Val Ser Leu Tyr Gln Trp
225 230 235 240
Lys Asp Thr Pro Asn Lys Ile Glu Thr Leu Glu Gly Glu Val Ser Leu
245 250 255
Leu Ala Asn Tyr Leu Ala Gly Thr Ser Pro Asp Ala Pro Lys Gly Met
260 265 270
Gly Asn Arg Tyr Asn Trp His Lys Leu Tyr Asp Thr Asp Tyr Tyr Phe
275 280 285
Leu Arg Glu Asp Tyr Leu Asp Val Glu Ala Asn Leu His Asp Leu Arg
290 295 300
Asp Tyr Leu Gly Ser Ser Ala Lys Gln Met Pro Trp Asp Glu Phe Ala
305 310 315 320
Lys Leu Ser Asp Ser Gln Gln Thr Leu Phe Leu Asp Ile Val Gly Phe
325 330 335
Asp Lys Glu Gln Leu Gln Gln Gln Tyr Ser Gln Ser Pro Leu Pro Asn
340 345 350
Phe Ile Phe Thr Gly Thr Thr Thr Trp Ala Gly Gly Glu Thr Lys Glu
355 360 365
Tyr Tyr Ala Gln Gln Gln Val Asn Val Ile Asn Asn Ala Ile Asn Glu
370 375 380
Thr Ser Pro Tyr Tyr Leu Gly Lys Asp Tyr Asp Leu Phe Phe Lys Gly
385 390 395 400
His Pro Ala Gly Gly Val Ile Asn Asp Ile Ile Leu Gly Ser Phe Pro
405 410 415
Asp Met Ile Asn Ile Pro Ala Lys Ile Ser Phe Glu Val Leu Met Met
420 425 430
Thr Asp Met Leu Pro Asp Thr Val Ala Gly Ile Ala Ser Ser Leu Tyr
435 440 445
Phe Thr Ile Pro Ala Asp Lys Val Asn Phe Ile Val Phe Thr Ser Ser
450 455 460
Asp Thr Ile Thr Asp Arg Glu Glu Ala Leu Lys Ser Pro Leu Val Gln
465 470 475 480
Val Met Leu Thr Leu Gly Ile Val Lys Glu Lys Asp Val Leu Phe Trp
485 490 495
Ala Asp Leu Pro Asp Cys Ser Ser Gly Val Cys Ile Asp Lys
500 505 510
<210> 66
<211> 422
<212> PRT
<213> Heliobacter acinonychis
<400> 66
Met Gly Thr Ile Lys Lys Pro Leu Ile Ile Ala Gly Asn Gly Pro Ser
1 5 10 15
Ile Lys Asp Leu Asp Tyr Ala Leu Phe Pro Lys Asp Phe Asp Val Phe
20 25 30
Arg Cys Asn Gln Phe Tyr Phe Glu Asp Lys Tyr Tyr Leu Gly Arg Glu
35 40 45
Ile Lys Gly Val Phe Phe Asn Pro Cys Val Leu Ser Ser Gln Met Gln
50 55 60
Thr Val Gln Tyr Leu Met Asp Asn Gly Glu Tyr Ser Ile Glu Arg Phe
65 70 75 80
Phe Cys Ser Val Ser Thr Asp Arg His Asp Phe Asp Gly Asp Tyr Gln
85 90 95
Thr Ile Leu Pro Val Asp Gly Tyr Leu Lys Ala His Tyr Pro Phe Val
100 105 110
Cys Asp Thr Phe Ser Leu Phe Lys Gly His Glu Glu Ile Leu Lys His
115 120 125
Val Lys Tyr His Leu Lys Thr Tyr Ser Lys Glu Leu Ser Ala Gly Val
130 135 140
Leu Met Leu Leu Ser Ala Val Val Leu Gly Tyr Lys Glu Ile Tyr Leu
145 150 155 160
Val Gly Ile Asp Phe Gly Ala Ser Ser Trp Gly His Phe Tyr Asp Glu
165 170 175
Ser Gln Ser Gln His Phe Ser Asn His Met Ala Asp Cys His Asn Ile
180 185 190
Tyr Tyr Asp Met Leu Thr Ile Cys Leu Cys Gln Lys Tyr Ala Lys Leu
195 200 205
Tyr Ala Leu Ala Pro Asn Ser Pro Leu Ser His Leu Leu Thr Leu Asn
210 215 220
Pro Gln Ala Lys Tyr Pro Phe Glu Leu Leu Asp Lys Pro Ile Gly Tyr
225 230 235 240
Thr Ser Asp Leu Ile Ile Ser Ser Pro Leu Glu Glu Lys Leu Leu Glu
245 250 255
Phe Lys Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu
260 265 270
Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys
275 280 285
Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu Lys Leu
290 295 300
Leu Glu Phe Lys Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys Asn Ile
305 310 315 320
Glu Glu Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu Lys Leu Leu Glu
325 330 335
Phe Lys Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu
340 345 350
Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys
355 360 365
Asn Ile Glu Glu Lys Leu Leu Glu Phe Lys Asn Ile Glu Glu Lys Leu
370 375 380
Leu Ala Ser Arg Leu Asn Asn Ile Leu Arg Lys Ile Lys Arg Lys Ile
385 390 395 400
Leu Pro Phe Phe Trp Gly Gly Gly Val Thr Pro Thr Leu Lys Val Ser
405 410 415
Phe Arg Trp Gly Ala Ala
420
<210> 67
<211> 609
<212> PRT
<213> Escherichia coli
<400> 67
Met Cys Gly Ile Val Gly Ala Ile Ala Gln Arg Asp Val Ala Glu Ile
1 5 10 15
Leu Leu Glu Gly Leu Arg Arg Leu Glu Tyr Arg Gly Tyr Asp Ser Ala
20 25 30
Gly Leu Ala Val Val Asp Ala Glu Gly His Met Thr Arg Leu Arg Arg
35 40 45
Leu Gly Lys Val Gln Met Leu Ala Gln Ala Ala Glu Glu His Pro Leu
50 55 60
His Gly Gly Thr Gly Ile Ala His Thr Arg Trp Ala Thr His Gly Glu
65 70 75 80
Pro Ser Glu Val Asn Ala His Pro His Val Ser Glu His Ile Val Val
85 90 95
Val His Asn Gly Ile Ile Glu Asn His Glu Pro Leu Arg Glu Glu Leu
100 105 110
Lys Ala Arg Gly Tyr Thr Phe Val Ser Glu Thr Asp Thr Glu Val Ile
115 120 125
Ala His Leu Val Asn Trp Glu Leu Lys Gln Gly Gly Thr Leu Arg Glu
130 135 140
Ala Val Leu Arg Ala Ile Pro Gln Leu Arg Gly Ala Tyr Gly Thr Val
145 150 155 160
Ile Met Asp Ser Arg His Pro Asp Thr Leu Leu Ala Ala Arg Ser Gly
165 170 175
Ser Pro Leu Val Ile Gly Leu Gly Met Gly Glu Asn Phe Ile Ala Ser
180 185 190
Asp Gln Leu Ala Leu Leu Pro Val Thr Arg Arg Phe Ile Phe Leu Glu
195 200 205
Glu Gly Asp Ile Ala Glu Ile Thr Arg Arg Ser Val Asn Ile Phe Asp
210 215 220
Lys Thr Gly Ala Glu Val Lys Arg Gln Asp Ile Glu Ser Asn Leu Gln
225 230 235 240
Tyr Asp Ala Gly Asp Lys Gly Ile Tyr Arg His Tyr Met Gln Lys Glu
245 250 255
Ile Tyr Glu Gln Pro Asn Ala Ile Lys Asn Thr Leu Thr Gly Arg Ile
260 265 270
Ser His Gly Gln Val Asp Leu Ser Glu Leu Gly Pro Asn Ala Asp Glu
275 280 285
Leu Leu Ser Lys Val Glu His Ile Gln Ile Leu Ala Cys Gly Thr Ser
290 295 300
Tyr Asn Ser Gly Met Val Ser Arg Tyr Trp Phe Glu Ser Leu Ala Gly
305 310 315 320
Ile Pro Cys Asp Val Glu Ile Ala Ser Glu Phe Arg Tyr Arg Lys Ser
325 330 335
Ala Val Arg Arg Asn Ser Leu Met Ile Thr Leu Ser Gln Ser Gly Glu
340 345 350
Thr Ala Asp Thr Leu Ala Gly Leu Arg Leu Ser Lys Glu Leu Gly Tyr
355 360 365
Leu Gly Ser Leu Ala Ile Cys Asn Val Pro Gly Ser Ser Leu Val Arg
370 375 380
Glu Ser Asp Leu Ala Leu Met Thr Asn Ala Gly Thr Glu Ile Gly Val
385 390 395 400
Ala Ser Thr Lys Ala Phe Thr Thr Gln Leu Thr Val Leu Leu Met Leu
405 410 415
Val Ala Lys Leu Ser Arg Leu Lys Gly Leu Asp Ala Ser Ile Glu His
420 425 430
Asp Ile Val His Gly Leu Gln Ala Leu Pro Ser Arg Ile Glu Gln Met
435 440 445
Leu Ser Gln Asp Lys Arg Ile Glu Ala Leu Ala Glu Asp Phe Ser Asp
450 455 460
Lys His His Ala Leu Phe Leu Gly Arg Gly Asp Gln Tyr Pro Ile Ala
465 470 475 480
Leu Glu Gly Ala Leu Lys Leu Lys Glu Ile Ser Tyr Ile His Ala Glu
485 490 495
Ala Tyr Ala Ala Gly Glu Leu Lys His Gly Pro Leu Ala Leu Ile Asp
500 505 510
Ala Asp Met Pro Val Ile Val Val Ala Pro Asn Asn Glu Leu Leu Glu
515 520 525
Lys Leu Lys Ser Asn Ile Glu Glu Val Arg Ala Arg Gly Gly Gln Leu
530 535 540
Tyr Val Phe Ala Asp Gln Asp Ala Gly Phe Val Ser Ser Asp Asn Met
545 550 555 560
His Ile Ile Glu Met Pro His Val Glu Glu Val Ile Ala Pro Ile Phe
565 570 575
Tyr Thr Val Pro Leu Gln Leu Leu Ala Tyr His Val Ala Leu Ile Lys
580 585 590
Gly Thr Asp Val Asp Gln Pro Arg Asn Leu Ala Lys Ser Val Thr Val
595 600 605
Glu
<210> 68
<211> 1830
<212> DNA
<213> Escherichia coli
<400> 68
atgtgtggaa ttgttggcgc gatcgcgcaa cgtgatgtag cagaaatcct tcttgaaggt 60
ttacgtcgtc tggaataccg cggatatgac tctgccggtc tggccgttgt tgatgcagaa 120
ggtcatatga cccgcctgcg tcgcctcggt aaagtccaga tgctggcaca ggcagcggaa 180
gaacatcctc tgcatggcgg cactggtatt gctcacactc gctgggcgac ccacggtgaa 240
ccttcagaag tgaatgcgca tccgcatgtt tctgaacaca ttgtggtggt gcataacggc 300
atcatcgaaa accatgaacc gctgcgtgaa gagctaaaag cgcgtggcta taccttcgtt 360
tctgaaaccg acaccgaagt gattgcccat ctggtgaact gggagctgaa acaaggcggg 420
actctgcgtg aggccgttct gcgtgctatc ccgcagctgc gtggtgcgta cggtacagtg 480
atcatggact cccgtcaccc ggataccctg ctggcggcac gttctggtag tccgctggtg 540
attggcctgg ggatgggcga aaactttatc gcttctgacc agctggcgct gttgccggtg 600
acccgtcgct ttatcttcct tgaagagggc gatattgcgg aaatcactcg ccgttcggta 660
aacatcttcg ataaaactgg cgcggaagta aaacgtcagg atatcgaatc caatctgcaa 720
tatgacgcgg gcgataaagg catttaccgt cactacatgc agaaagagat ctacgaacag 780
ccgaacgcga tcaaaaacac ccttaccgga cgcatcagcc acggtcaggt tgatttaagc 840
gagctgggac cgaacgccga cgaactgctg tcgaaggttg agcatattca gatcctcgcc 900
tgtggtactt cttataactc cggtatggtt tcccgctact ggtttgaatc gctagcaggt 960
attccgtgcg acgtcgaaat cgcctctgaa ttccgctatc gcaaatctgc cgtgcgtcgt 1020
aacagcctga tgatcacctt gtcacagtct ggcgaaaccg cggataccct ggctggcctg 1080
cgtctgtcga aagagctggg ttaccttggt tcactggcaa tctgtaacgt tccgggttct 1140
tctctggtgc gcgaatccga tctggcgcta atgaccaacg cgggtacaga aatcggcgtg 1200
gcatccacta aagcattcac cactcagtta actgtgctgt tgatgctggt ggcgaagctg 1260
tctcgcctga aaggtctgga tgcctccatt gaacatgaca tcgtgcatgg tctgcaggcg 1320
ctgccgagcc gtattgagca gatgctgtct caggacaaac gcattgaagc gctggcagaa 1380
gatttctctg acaaacatca cgcgctgttc ctgggccgtg gcgatcagta cccaatcgcg 1440
ctggaaggcg cattgaagtt gaaagagatc tcttacattc acgctgaagc ctacgctgct 1500
ggcgaactga aacacggtcc gctggcgcta attgatgccg atatgccggt tattgttgtt 1560
gcaccgaaca acgaattgct ggaaaaactg aaatccaaca ttgaagaagt tcgcgcgcgt 1620
ggcggtcagt tgtatgtctt cgccgatcag gatgcgggtt ttgtaagtag cgataacatg 1680
cacatcatcg agatgccgca tgtggaagag gtgattgcac cgatcttcta caccgttccg 1740
ctgcagctgc tggcttacca tgtcgcgctg atcaaaggca ccgacgttga ccagccgcgt 1800
aacctggcaa aatcggttac ggttgagtaa 1830
<210> 69
<211> 609
<212> PRT
<213> Escherichia coli
<400> 69
Met Cys Gly Ile Val Gly Ala Ile Ala Gln Arg Asp Val Ala Lys Ile
1 5 10 15
Leu Leu Glu Gly Leu Arg Arg Leu Glu Tyr Arg Gly Tyr Asp Ser Ala
20 25 30
Gly Leu Ala Val Val Asp Ala Glu Gly His Met Thr Arg Leu Arg Arg
35 40 45
Leu Gly Lys Val Gln Met Leu Ala Gln Ala Ala Glu Glu His Pro Leu
50 55 60
His Gly Gly Thr Gly Ile Ala His Thr Arg Trp Ala Thr His Gly Glu
65 70 75 80
Pro Ser Glu Val Asn Ala His Pro His Val Ser Glu His Ile Val Val
85 90 95
Val His Asn Gly Ile Ile Glu Asn His Glu Pro Leu Arg Glu Glu Leu
100 105 110
Lys Ala Arg Gly Tyr Thr Phe Val Ser Glu Thr Asp Thr Glu Val Ile
115 120 125
Ala His Leu Val Asn Trp Glu Leu Lys Gln Gly Gly Thr Leu Arg Glu
130 135 140
Ala Val Leu Arg Ala Ile Pro Gln Leu Arg Gly Ala Tyr Gly Thr Val
145 150 155 160
Ile Met Asp Ser Arg His Pro Asp Thr Leu Leu Ala Ala Arg Ser Gly
165 170 175
Ser Pro Leu Val Ile Gly Leu Gly Met Gly Glu Asn Phe Ile Ala Ser
180 185 190
Asp Gln Leu Ala Leu Leu Pro Val Thr Arg Arg Phe Ile Phe Leu Glu
195 200 205
Glu Gly Asp Ile Ala Glu Ile Thr Arg Arg Ser Val Asn Ile Phe Asp
210 215 220
Lys Thr Gly Ala Glu Val Lys Arg Gln Asp Ile Glu Ser Asn Leu Gln
225 230 235 240
Tyr Asp Ala Gly Asp Lys Gly Ile Tyr Arg His Tyr Met Gln Lys Glu
245 250 255
Ile Tyr Glu Gln Pro Asn Ala Ile Lys Asn Thr Leu Thr Gly Arg Ile
260 265 270
Ser His Gly Gln Val Asp Leu Ser Glu Leu Gly Pro Asn Ala Asp Glu
275 280 285
Leu Leu Ser Lys Val Glu His Ile Gln Ile Leu Ala Cys Gly Thr Ser
290 295 300
Tyr Asn Ser Gly Met Val Ser Arg Tyr Trp Phe Glu Ser Leu Ala Gly
305 310 315 320
Ile Pro Cys Asp Val Glu Ile Ala Ser Glu Phe Arg Tyr Arg Lys Ser
325 330 335
Ala Val Arg Arg Asn Ser Leu Met Ile Thr Leu Ser Gln Ser Gly Glu
340 345 350
Thr Ala Asp Thr Leu Ala Gly Leu Arg Leu Ser Lys Glu Leu Gly Tyr
355 360 365
Leu Gly Ser Leu Ala Ile Cys Asn Val Pro Gly Ser Ser Leu Val Arg
370 375 380
Glu Ser Val Leu Ala Leu Met Thr Asn Ala Gly Thr Glu Ile Gly Val
385 390 395 400
Ala Ser Thr Lys Ala Phe Thr Thr Gln Leu Thr Val Leu Leu Met Leu
405 410 415
Val Ala Lys Leu Ser Arg Leu Lys Gly Leu Asp Ala Ser Ile Glu His
420 425 430
Asp Ile Val His Gly Leu Gln Ala Leu Pro Ser Arg Ile Glu Gln Met
435 440 445
Leu Pro Gln Asp Lys Arg Ile Glu Ala Leu Ala Glu Asp Phe Ser Asp
450 455 460
Lys His His Ala Leu Phe Leu Gly Arg Gly Asp Gln Tyr Pro Ile Ala
465 470 475 480
Leu Glu Gly Ala Leu Lys Leu Lys Glu Ile Ser Tyr Ile His Ala Glu
485 490 495
Ala Tyr Ala Ala Gly Glu Leu Lys His Gly Pro Leu Ala Leu Ile Asp
500 505 510
Ala Asp Met Pro Val Ile Val Val Ala Pro Asn Asn Gly Leu Leu Glu
515 520 525
Lys Leu Lys Ser Asn Ile Glu Glu Val Arg Ala Arg Gly Gly Gln Leu
530 535 540
Tyr Val Phe Ala Asp Gln Asp Ala Gly Phe Val Ser Ser Asp Asn Met
545 550 555 560
His Ile Ile Glu Met Pro His Val Glu Glu Val Ile Ala Pro Ile Phe
565 570 575
Tyr Thr Val Pro Leu Gln Leu Leu Ala Tyr His Val Ala Leu Ile Lys
580 585 590
Gly Thr Asp Val Asp Gln Pro Arg Asn Leu Ala Lys Ser Val Thr Val
595 600 605
Glu
<210> 70
<211> 1830
<212> DNA
<213> Escherichia coli
<400> 70
atgtgcggta tcgttggtgc tatcgcacag cgtgatgtag cgaaaatcct cctggaaggt 60
ctgcgtcgtc tcgaataccg tggttacgac tctgccggtc tggcagtagt ggatgcagaa 120
ggtcacatga ctcgtctgcg tcgtctgggt aaagtgcaga tgctcgcgca ggcggcggaa 180
gaacacccac tccacggtgg tacgggtatc gcacacactc gttgggcaac ccacggtgaa 240
ccgtctgagg tcaacgcaca cccgcatgtt agcgagcaca tcgtagtcgt tcacaacggt 300
atcatcgaga accacgaacc actccgtgag gaactcaaag cccgtggtta caccttcgta 360
agcgaaaccg acacggaagt tatcgcccac ctcgttaact gggaactcaa acagggtggt 420
actctgcgtg aagcagttct gcgtgccatt ccacagctgc gtggtgcata cggtaccgtg 480
atcatggact ctcgtcatcc ggataccctg ctcgccgcac gttctggttc tccactcgtt 540
atcggtctgg gtatgggtga gaacttcatc gcctctgatc agctggccct gctcccagtt 600
acccgtcgct tcatcttcct ggaagagggt gacatcgccg aaatcacccg tcgttccgtt 660
aacatcttcg acaaaacggg tgcggaagtt aaacgtcagg acatcgagtc taacctgcag 720
tatgacgctg gtgacaaagg catctaccgt cactacatgc agaaagagat ctacgaacag 780
ccgaacgcga tcaaaaacac cctgaccggt cgtatctctc acggtcaggt tgacctgtct 840
gagctgggtc caaacgcgga cgaactcctg tccaaagtcg agcacatcca gatcctggct 900
tgtggtacct cttacaactc cggtatggtt tctcgttact ggttcgaatc tctggcaggt 960
atcccatgcg acgttgaaat cgcctccgaa ttccgttatc gtaaatctgc ggtacgtcgt 1020
aactccctca tgatcaccct gtctcagtct ggtgaaaccg ctgatactct ggcaggtctg 1080
cgtctcagca aagaactggg ttacctgggt tctctggcca tctgcaacgt tccgggttct 1140
agcctggttc gtgagtctgt gctggctctg atgaccaacg cgggtacgga gatcggtgtt 1200
gcctctacca aagcgttcac tacccagctc actgtcctgc tgatgctggt tgccaaactg 1260
tctcgtctca aaggcctcga cgctagcatc gaacacgaca tcgtacacgg tctgcaggcc 1320
ctcccatctc gtatcgagca gatgctgccg caggacaaac gtatcgaagc actggcagaa 1380
gacttcagcg acaaacacca cgcgctgttt ctgggtcgtg gtgaccagta cccaattgcg 1440
ctggaaggtg ccctgaaact gaaagagatc agctacatcc atgcagaggc atacgcagcg 1500
ggtgagctga aacatggtcc actggccctg atcgacgcag atatgccggt tattgtggtt 1560
gctccgaaca acggcctgct ggagaaactg aaatccaaca tcgaggaagt acgtgcgcgt 1620
ggtggtcagc tgtacgtgtt tgctgaccag gacgcgggtt tcgtttccag cgacaacatg 1680
cacatcatcg aaatgccgca tgttgaagag gtaatcgcgc caatcttcta caccgtaccg 1740
ctgcagctgc tggcgtacca tgtagccctg atcaaaggta cggacgttga ccagccgcgt 1800
aacctggcga aatccgtgac cgtggaataa 1830
<210> 71
<211> 445
<212> PRT
<213> Escherichia coli
<400> 71
Met Ser Asn Arg Lys Tyr Phe Gly Thr Asp Gly Ile Arg Gly Arg Val
1 5 10 15
Gly Asp Ala Pro Ile Thr Pro Asp Phe Val Leu Lys Leu Gly Trp Ala
20 25 30
Ala Gly Lys Val Leu Ala Arg His Gly Ser Arg Lys Ile Ile Ile Gly
35 40 45
Lys Asp Thr Arg Ile Ser Gly Tyr Met Leu Glu Ser Ala Leu Glu Ala
50 55 60
Gly Leu Ala Ala Ala Gly Leu Ser Ala Leu Phe Thr Gly Pro Met Pro
65 70 75 80
Thr Pro Ala Val Ala Tyr Leu Thr Arg Thr Phe Arg Ala Glu Ala Gly
85 90 95
Ile Val Ile Ser Ala Ser His Asn Pro Phe Tyr Asp Asn Gly Ile Lys
100 105 110
Phe Phe Ser Ile Asp Gly Thr Lys Leu Pro Asp Ala Val Glu Glu Ala
115 120 125
Ile Glu Ala Glu Met Glu Lys Glu Ile Ser Cys Val Asp Ser Ala Glu
130 135 140
Leu Gly Lys Ala Ser Arg Ile Val Asp Ala Ala Gly Arg Tyr Ile Glu
145 150 155 160
Phe Cys Lys Ala Thr Phe Pro Asn Glu Leu Ser Leu Ser Glu Leu Lys
165 170 175
Ile Val Val Asp Cys Ala Asn Gly Ala Thr Tyr His Ile Ala Pro Asn
180 185 190
Val Leu Arg Glu Leu Gly Ala Asn Val Ile Ala Ile Gly Cys Glu Pro
195 200 205
Asn Gly Val Asn Ile Asn Ala Glu Val Gly Ala Thr Asp Val Arg Ala
210 215 220
Leu Gln Ala Arg Val Leu Ala Glu Lys Ala Asp Leu Gly Ile Ala Phe
225 230 235 240
Asp Gly Asp Gly Asp Arg Val Ile Met Val Asp His Glu Gly Asn Lys
245 250 255
Val Asp Gly Asp Gln Ile Met Tyr Ile Ile Ala Arg Glu Gly Leu Arg
260 265 270
Gln Gly Gln Leu Arg Gly Gly Ala Val Gly Thr Leu Met Ser Asn Met
275 280 285
Gly Leu Glu Leu Ala Leu Lys Gln Leu Gly Ile Pro Phe Ala Arg Ala
290 295 300
Lys Val Gly Asp Arg Tyr Val Leu Glu Lys Met Gln Glu Lys Gly Trp
305 310 315 320
Arg Ile Gly Ala Glu Asn Ser Gly His Val Ile Leu Leu Asp Lys Thr
325 330 335
Thr Thr Gly Asp Gly Ile Val Ala Gly Leu Gln Val Leu Ala Ala Met
340 345 350
Ala Arg Asn His Met Ser Leu His Asp Leu Cys Ser Gly Met Lys Met
355 360 365
Phe Pro Gln Ile Leu Val Asn Val Arg Tyr Thr Ala Gly Ser Gly Asp
370 375 380
Pro Leu Glu His Glu Ser Val Lys Ala Val Thr Ala Glu Val Glu Ala
385 390 395 400
Ala Leu Gly Asn Arg Gly Arg Val Leu Leu Arg Lys Ser Gly Thr Glu
405 410 415
Pro Leu Ile Arg Val Met Val Glu Gly Glu Asp Glu Ala Gln Val Thr
420 425 430
Glu Phe Ala His Arg Ile Ala Asp Ala Val Lys Ala Val
435 440 445
<210> 72
<211> 1338
<212> DNA
<213> Escherichia coli
<400> 72
atgagtaatc gtaaatattt cggtaccgat gggattcgtg gtcgtgtagg ggatgcgccg 60
atcacacctg attttgtgct taagctgggt tgggccgcgg gtaaagtgct ggcgcgccac 120
ggctcccgta agattattat tggtaaagac acgcgtattt ctggctatat gctggagtca 180
gcactggaag cgggtctggc ggcagcgggc ctttccgcac tcttcactgg cccgatgcca 240
acaccggccg tggcttatct gacgcgtacc ttccgcgcag aggccggaat tgtgatatct 300
gcatcgcata acccgttcta cgataatggc attaaattct tctctatcga cggcaccaaa 360
ctgccggatg cggtagaaga ggccatcgaa gcggaaatgg aaaaggagat cagctgcgtt 420
gattcggcag aactgggtaa agccagccgt atcgttgatg ccgcgggtcg ctatatcgag 480
ttttgcaaag ccacgttccc gaacgaactt agcctcagtg aactgaagat tgtggtggat 540
tgtgcaaacg gtgcgactta tcacatcgcg ccgaacgtgc tgcgcgaact gggggcgaac 600
gttatcgcta tcggttgtga gccaaacggt gtaaacatca atgccgaagt gggggctacc 660
gacgttcgcg cgctccaggc tcgtgtgctg gctgaaaaag cggatctcgg tattgccttc 720
gacggcgatg gcgatcgcgt gattatggtt gaccatgaag gcaataaagt cgatggcgat 780
cagatcatgt atatcatcgc gcgtgaaggt cttcgtcagg gccagctgcg tggtggcgct 840
gtgggtacat tgatgagcaa catggggctt gaactggcgc tgaaacagtt aggaattcca 900
tttgcgcgcg cgaaagtggg tgaccgctac gtactggaaa aaatgcagga gaaaggctgg 960
cgtatcggtg cagagaattc cggtcatgtg atcctgctgg ataaaactac taccggtgac 1020
ggcatcgttg ctggcttgca ggtgctggcg gcgatggcac gtaaccatat gagcctgcac 1080
gacctttgca gcggcatgaa aatgttcccg cagattctgg ttaacgtacg ttacaccgca 1140
ggtagcggcg atccacttga gcatgagtca gttaaagccg tgaccgcaga ggttgaagct 1200
gcgctgggca accgtggacg cgtgttgctg cgtaaatccg gcaccgaacc gttaattcgc 1260
gtgatggtgg aaggcgaaga cgaagcgcag gtgactgaat ttgcacaccg catcgccgat 1320
gcagtaaaag ccgtttaa 1338
<210> 73
<211> 456
<212> PRT
<213> Escherichia coli
<400> 73
Met Leu Asn Asn Ala Met Ser Val Val Ile Leu Ala Ala Gly Lys Gly
1 5 10 15
Thr Arg Met Tyr Ser Asp Leu Pro Lys Val Leu His Thr Leu Ala Gly
20 25 30
Lys Ala Met Val Gln His Val Ile Asp Ala Ala Asn Glu Leu Gly Ala
35 40 45
Ala His Val His Leu Val Tyr Gly His Gly Gly Asp Leu Leu Lys Gln
50 55 60
Ala Leu Lys Asp Asp Asn Leu Asn Trp Val Leu Gln Ala Glu Gln Leu
65 70 75 80
Gly Thr Gly His Ala Met Gln Gln Ala Ala Pro Phe Phe Ala Asp Asp
85 90 95
Glu Asp Ile Leu Met Leu Tyr Gly Asp Val Pro Leu Ile Ser Val Glu
100 105 110
Thr Leu Gln Arg Leu Arg Asp Ala Lys Pro Gln Gly Gly Ile Gly Leu
115 120 125
Leu Thr Val Lys Leu Asp Asp Pro Thr Gly Tyr Gly Arg Ile Thr Arg
130 135 140
Glu Asn Gly Lys Val Thr Gly Ile Val Glu His Lys Asp Ala Thr Asp
145 150 155 160
Glu Gln Arg Gln Ile Gln Glu Ile Asn Thr Gly Ile Leu Ile Ala Asn
165 170 175
Gly Ala Asp Met Lys Arg Trp Leu Ala Lys Leu Thr Asn Asn Asn Ala
180 185 190
Gln Gly Glu Tyr Tyr Ile Thr Asp Ile Ile Ala Leu Ala Tyr Gln Glu
195 200 205
Gly Arg Glu Ile Val Ala Val His Pro Gln Arg Leu Ser Glu Val Glu
210 215 220
Gly Val Asn Asn Arg Leu Gln Leu Ser Arg Leu Glu Arg Val Tyr Gln
225 230 235 240
Ser Glu Gln Ala Glu Lys Leu Leu Leu Ala Gly Val Met Leu Arg Asp
245 250 255
Pro Ala Arg Phe Asp Leu Arg Gly Thr Leu Thr His Gly Arg Asp Val
260 265 270
Glu Ile Asp Thr Asn Val Ile Ile Glu Gly Asn Val Thr Leu Gly His
275 280 285
Arg Val Lys Ile Gly Thr Gly Cys Val Ile Lys Asn Ser Val Ile Gly
290 295 300
Asp Asp Cys Glu Ile Ser Pro Tyr Thr Val Val Glu Asp Ala Asn Leu
305 310 315 320
Ala Ala Ala Cys Thr Ile Gly Pro Phe Ala Arg Leu Arg Pro Gly Ala
325 330 335
Glu Leu Leu Glu Gly Ala His Val Gly Asn Phe Val Glu Met Lys Lys
340 345 350
Ala Arg Leu Gly Lys Gly Ser Lys Ala Gly His Leu Thr Tyr Leu Gly
355 360 365
Asp Ala Glu Ile Gly Asp Asn Val Asn Ile Gly Ala Gly Thr Ile Thr
370 375 380
Cys Asn Tyr Asp Gly Ala Asn Lys Phe Lys Thr Ile Ile Gly Asp Asp
385 390 395 400
Val Phe Val Gly Ser Asp Thr Gln Leu Val Ala Pro Val Thr Val Gly
405 410 415
Lys Gly Ala Thr Ile Ala Ala Gly Thr Thr Val Thr Arg Asn Val Gly
420 425 430
Glu Asn Ala Leu Ala Ile Ser Arg Val Pro Gln Thr Gln Lys Glu Gly
435 440 445
Trp Arg Arg Pro Val Lys Lys Lys
450 455
<210> 74
<211> 1371
<212> DNA
<213> Escherichia coli
<400> 74
atgttgaata atgctatgag cgtagtgatc cttgccgcag gcaaaggcac gcgcatgtat 60
tccgatcttc cgaaagtgct gcataccctt gccgggaaag cgatggttca gcatgtcatt 120
gatgctgcga atgaattagg cgcagcgcac gttcacctgg tgtacggtca cggcggcgat 180
ctgctaaaac aggcgctgaa agacgacaac cttaactggg tgcttcaggc agagcagctg 240
ggtacgggtc atgcaatgca gcaggccgca cctttctttg ccgatgatga agacatttta 300
atgctctacg gcgacgtgcc gctgatctct gtcgaaacac tccagcgtct gcgtgatgct 360
aaaccgcagg gtggcattgg tctgctgacg gtgaaactgg atgatccgac cggttatgga 420
cgtatcaccc gtgaaaacgg caaagttacc ggcattgttg agcacaaaga tgccaccgac 480
gagcagcgtc agattcagga gatcaacacc ggcattctga ttgccaacgg cgcagatatg 540
aaacgctggc tggcgaagct gaccaacaat aatgctcagg gcgaatacta catcaccgac 600
attattgcgc tggcgtatca ggaagggcgt gaaatcgtcg ccgttcatcc gcaacgttta 660
agcgaagtag aaggcgtgaa taaccgcctg caactctccc gtctggagcg tgtttatcag 720
tccgaacagg ctgaaaaact gctgttagca ggcgttatgc tgcgcgatcc agcgcgtttt 780
gatctgcgtg gtacgctaac tcacgggcgc gatgttgaaa ttgatactaa cgttatcatc 840
gagggcaacg tgactctcgg tcatcgcgtg aaaattggca ccggttgcgt gattaaaaac 900
agcgtgattg gcgatgattg cgaaatcagt ccgtataccg ttgtggaaga tgcgaatctg 960
gcagcggcct gtaccattgg cccgtttgcc cgtttgcgtc ctggtgctga gttgctggaa 1020
ggtgctcacg tcggtaactt cgttgagatg aaaaaagcgc gtctgggtaa aggctcgaaa 1080
gctggtcatc tgacttacct gggcgatgcg gaaattggcg ataacgttaa catcggcgcg 1140
ggaaccatta cctgcaacta cgatggtgcg aataaattta agaccattat cggcgacgat 1200
gtgtttgttg gttccgacac tcagctggtg gccccggtaa cagtaggcaa aggcgcgacc 1260
attgctgcgg gtacaactgt gacgcgtaat gtcggcgaaa atgcattagc tatcagccgt 1320
gtgccgcaga ctcagaaaga aggctggcgt cgtccggtaa agaaaaagtg a 1371
<210> 75
<211> 391
<212> PRT
<213> Escherichia coli
<400> 75
Met Lys Lys Ile Leu Tyr Val Thr Gly Ser Arg Ala Glu Tyr Gly Ile
1 5 10 15
Val Arg Arg Leu Leu Thr Met Leu Arg Glu Thr Pro Glu Ile Gln Leu
20 25 30
Asp Leu Ala Val Thr Gly Met His Cys Asp Asn Ala Tyr Gly Asn Thr
35 40 45
Ile His Ile Ile Glu Gln Asp Asn Phe Asn Ile Ile Lys Val Val Asp
50 55 60
Ile Asn Ile Asn Thr Thr Ser His Thr His Ile Leu His Ser Met Ser
65 70 75 80
Val Cys Leu Asn Ser Phe Gly Asp Phe Phe Ser Asn Asn Thr Tyr Asp
85 90 95
Ala Val Met Val Leu Gly Asp Arg Tyr Glu Ile Phe Ser Val Ala Ile
100 105 110
Ala Ala Ser Met His Asn Ile Pro Leu Ile His Ile His Gly Gly Glu
115 120 125
Lys Thr Leu Ala Asn Tyr Asp Glu Phe Ile Arg His Ser Ile Thr Lys
130 135 140
Met Ser Lys Leu His Leu Thr Ser Thr Glu Glu Tyr Lys Lys Arg Val
145 150 155 160
Ile Gln Leu Gly Glu Lys Pro Gly Ser Val Phe Asn Ile Gly Ser Leu
165 170 175
Gly Ala Glu Asn Ala Leu Ser Leu His Leu Pro Asn Lys Gln Glu Leu
180 185 190
Glu Leu Lys Tyr Gly Ser Leu Leu Lys Arg Tyr Phe Val Val Val Phe
195 200 205
His Pro Glu Thr Leu Ser Thr Gln Ser Val Asn Asp Gln Ile Asp Glu
210 215 220
Leu Leu Ser Ala Ile Ser Phe Phe Lys Asn Thr His Asp Phe Ile Phe
225 230 235 240
Ile Gly Ser Asn Ala Asp Thr Gly Ser Asp Ile Ile Gln Arg Lys Val
245 250 255
Lys Tyr Phe Cys Lys Glu Tyr Lys Phe Arg Tyr Leu Ile Ser Ile Arg
260 265 270
Ser Glu Asp Tyr Leu Ala Met Ile Lys Tyr Ser Cys Gly Leu Ile Gly
275 280 285
Asn Ser Ser Ser Gly Leu Ile Glu Val Pro Ser Leu Lys Val Ala Thr
290 295 300
Ile Asn Ile Gly Asp Arg Gln Lys Gly Arg Val Arg Gly Ala Ser Val
305 310 315 320
Ile Asp Val Pro Val Glu Lys Asn Ala Ile Val Arg Gly Ile Asn Ile
325 330 335
Ser Gln Asp Glu Lys Phe Ile Ser Val Val Gln Ser Ser Ser Asn Pro
340 345 350
Tyr Phe Lys Glu Asn Ala Leu Ile Asn Ala Val Arg Ile Ile Lys Asp
355 360 365
Phe Ile Lys Ser Lys Asn Lys Asp Tyr Lys Asp Phe Tyr Asp Ile Pro
370 375 380
Glu Cys Thr Thr Ser Tyr Asp
385 390
<210> 76
<211> 1176
<212> DNA
<213> Escherichia coli
<400> 76
atgaaaaaaa tattatacgt aactggatct agagctgaat atggaatagt tcggagactt 60
ttgacaatgc taagagaaac tccagaaata cagcttgatt tggcagttac aggaatgcat 120
tgtgataatg cgtatggaaa tacaatacat attatagaac aagataattt taatattatc 180
aaggttgtgg atataaatat caatacaact tcacatactc acattctcca ttcaatgagt 240
gtttgcctca attcgtttgg tgattttttt tcaaataaca catatgatgc ggttatggtt 300
ttaggcgata gatatgaaat attttcagtc gctatcgcag catcaatgca taatattcca 360
ttaattcata ttcatggtgg tgaaaagaca ttagctaatt atgatgagtt tattaggcat 420
tcaattacta aaatgagtaa actccatctt acttctacag aagagtataa aaaacgagta 480
attcaactag gtgaaaagcc tggtagtgtg tttaatattg gttctcttgg tgcagaaaat 540
gctctttcat tgcatttacc aaataagcag gagttggaac taaaatatgg ttcactgtta 600
aaacggtact ttgttgtagt attccatcct gaaacacttt ccacgcagtc ggttaatgat 660
caaatagatg agttattgtc agcgatttct ttttttaaaa atactcacga ctttattttt 720
attggcagta acgctgacac tggttctgat ataattcaga gaaaagtaaa atatttttgc 780
aaagagtata agttcagata tttgatttct attcgttcag aagattattt ggcaatgatt 840
aaatactctt gtgggctaat tgggaactcc tcctctggtt taattgaggt tccatcttta 900
aaagttgcaa caattaacat tggtgatagg cagaaaggcc gtgttcgtgg agccagtgta 960
atagatgtac ccgttgaaaa aaatgcaatc gtcagaggga taaatatatc tcaagatgaa 1020
aaatttatta gtgttgtaca gtcatctagt aatccttatt ttaaagaaaa tgctttaatt 1080
aatgctgtta gaattattaa ggattttatt aaatcaaaaa ataaagatta caaagatttt 1140
tatgacatcc cggaatgtac caccagttat gactag 1176
<210> 77
<211> 159
<212> PRT
<213> Saccharomyces cerevisiae
<400> 77
Met Ser Leu Pro Asp Gly Phe Tyr Ile Arg Arg Met Glu Glu Gly Asp
1 5 10 15
Leu Glu Gln Val Thr Glu Thr Leu Lys Val Leu Thr Thr Val Gly Thr
20 25 30
Ile Thr Pro Glu Ser Phe Ser Lys Leu Ile Lys Tyr Trp Asn Glu Ala
35 40 45
Thr Val Trp Asn Asp Asn Glu Asp Lys Lys Ile Met Gln Tyr Asn Pro
50 55 60
Met Val Ile Val Asp Lys Arg Thr Glu Thr Val Ala Ala Thr Gly Asn
65 70 75 80
Ile Ile Ile Glu Arg Lys Ile Ile His Glu Leu Gly Leu Cys Gly His
85 90 95
Ile Glu Asp Ile Ala Val Asn Ser Lys Tyr Gln Gly Gln Gly Leu Gly
100 105 110
Lys Leu Leu Ile Asp Gln Leu Val Thr Ile Gly Phe Asp Tyr Gly Cys
115 120 125
Tyr Lys Ile Ile Leu Asp Cys Asp Glu Lys Asn Val Lys Phe Tyr Glu
130 135 140
Lys Cys Gly Phe Ser Asn Ala Gly Val Glu Met Gln Ile Arg Lys
145 150 155
<210> 78
<211> 480
<212> DNA
<213> Saccharomyces cerevisiae
<400> 78
atgagcttac ccgatggatt ttatataagg cgaatggaag agggggattt ggaacaggtc 60
actgagacgc taaaggtttt gaccaccgtg ggcactatta cccccgaatc cttcagcaaa 120
ctcataaaat actggaatga agccacagta tggaatgata acgaagataa aaaaataatg 180
caatataacc ccatggtgat tgtggacaag cgcaccgaga cggttgccgc tacggggaat 240
atcatcatcg aaagaaagat cattcatgaa ctggggctat gtggccacat cgaggacatt 300
gcagtaaact ccaagtatca gggccaaggt ttgggcaagc tcttgattga tcaattggta 360
actatcggct ttgactacgg ttgttataag attattttag attgcgatga gaaaaatgtc 420
aaattctatg aaaaatgtgg gtttagcaac gcaggcgtgg aaatgcaaat tagaaaatag 480
<210> 79
<211> 188
<212> PRT
<213> Escherichia coli
<400> 79
Met Tyr Glu Arg Tyr Ala Gly Leu Ile Phe Asp Met Asp Gly Thr Ile
1 5 10 15
Leu Asp Thr Glu Pro Thr His Arg Lys Ala Trp Arg Glu Val Leu Gly
20 25 30
His Tyr Gly Leu Gln Tyr Asp Ile Gln Ala Met Ile Ala Leu Asn Gly
35 40 45
Ser Pro Thr Trp Arg Ile Ala Gln Ala Ile Ile Glu Leu Asn Gln Ala
50 55 60
Asp Leu Asp Pro His Ala Leu Ala Arg Glu Lys Thr Glu Ala Val Arg
65 70 75 80
Ser Met Leu Leu Asp Ser Val Glu Pro Leu Pro Leu Val Asp Val Val
85 90 95
Lys Ser Trp His Gly Arg Arg Pro Met Ala Val Gly Thr Gly Ser Glu
100 105 110
Ser Ala Ile Ala Glu Ala Leu Leu Ala His Leu Gly Leu Arg His Tyr
115 120 125
Phe Asp Ala Val Val Ala Ala Asp His Val Lys His His Lys Pro Ala
130 135 140
Pro Asp Thr Phe Leu Leu Cys Ala Gln Arg Met Gly Val Gln Pro Thr
145 150 155 160
Gln Cys Val Val Phe Glu Asp Ala Asp Phe Gly Ile Gln Ala Ala Arg
165 170 175
Ala Ala Gly Met Asp Ala Val Asp Val Arg Leu Leu
180 185
<210> 80
<211> 199
<212> PRT
<213> Escherichia coli
<400> 80
Met Leu Tyr Ile Phe Asp Leu Gly Asn Val Ile Val Asp Ile Asp Phe
1 5 10 15
Asn Arg Val Leu Gly Ala Trp Ser Asp Leu Thr Arg Ile Pro Leu Ala
20 25 30
Ser Leu Lys Lys Ser Phe His Met Gly Glu Ala Phe His Gln His Glu
35 40 45
Arg Gly Glu Ile Ser Asp Glu Ala Phe Ala Glu Ala Leu Cys His Glu
50 55 60
Met Ala Leu Pro Leu Ser Tyr Glu Gln Phe Ser His Gly Trp Gln Ala
65 70 75 80
Val Phe Val Ala Leu Arg Pro Glu Val Ile Ala Ile Met His Lys Leu
85 90 95
Arg Glu Gln Gly His Arg Val Val Val Leu Ser Asn Thr Asn Arg Leu
100 105 110
His Thr Thr Phe Trp Pro Glu Glu Tyr Pro Glu Ile Arg Asp Ala Ala
115 120 125
Asp His Ile Tyr Leu Ser Gln Asp Leu Gly Met Arg Lys Pro Glu Ala
130 135 140
Arg Ile Tyr Gln His Val Leu Gln Ala Glu Gly Phe Ser Pro Ser Asp
145 150 155 160
Thr Val Phe Phe Asp Asp Asn Ala Asp Asn Ile Glu Gly Ala Asn Gln
165 170 175
Leu Gly Ile Thr Ser Ile Leu Val Lys Asp Lys Thr Thr Ile Pro Asp
180 185 190
Tyr Phe Ala Lys Val Leu Cys
195
<210> 81
<211> 567
<212> DNA
<213> Escherichia coli
<400> 81
atgtacgagc gttatgcagg tttaattttt gatatggatg gcacaatcct ggatacggag 60
cctacgcacc gtaaagcgtg gcgcgaagta ttagggcact acggtcttca gtacgatatt 120
caggcgatga ttgcgcttaa tggatcgccc acctggcgta ttgctcaggc aattattgag 180
ctgaatcagg ccgatctcga cccgcatgcg ttagcgcgtg aaaaaacaga agcagtaaga 240
agtatgctgc tggatagcgt cgaaccgctt cctcttgttg atgtggtgaa aagttggcat 300
ggtcgtcgcc caatggctgt aggaacgggg agtgaaagcg ccatcgctga ggcattgctg 360
gcgcacctgg gattacgcca ttattttgac gccgtcgtcg ctgccgatca cgtcaaacac 420
cataaacccg cgccagacac atttttgttg tgcgcgcagc gtatgggcgt gcaaccgacg 480
cagtgtgtgg tctttgaaga tgccgatttc ggtattcagg cggcccgtgc agcaggcatg 540
gacgccgtgg atgttcgctt gctgtga 567
<210> 82
<211> 600
<212> DNA
<213> Escherichia coli
<400> 82
atgctctata tctttgattt aggtaatgtg attgtcgata tcgactttaa ccgtgtgctg 60
ggagcctgga gcgatttaac gcgtattccg ctggcatcgc ttaagaagag ttttcatatg 120
ggggaggcgt ttcatcagca tgagcgtggg gaaattagcg acgaagcgtt cgcagaggcg 180
ctgtgtcatg agatggctct accgctaagc tacgagcagt tctctcacgg ctggcaggcg 240
gtgtttgttg cgctgcgccc ggaagtgatc gccatcatgc ataaactgcg tgagcagggg 300
catcgcgtgg tggtgctttc caataccaac cgcctgcata ccaccttctg gccggaagaa 360
tacccggaaa ttcgtgatgc tgctgaccat atctatctgt cgcaagatct ggggatgcgc 420
aaacctgaag cacgaattta ccagcatgtt ttgcaggcgg aaggtttttc acccagcgat 480
acggtctttt tcgacgataa cgccgataat atagaaggag ccaatcagct gggcattacc 540
agtattctgg tgaaagataa aaccaccatc ccggactatt tcgcgaaggt gttatgctaa 600
<210> 83
<211> 421
<212> PRT
<213> Bacteroides ovatus
<400> 83
Met Asp Ser Lys Asn Asn Ile Gly His Ser Ala Asp Ile Ser Leu Thr
1 5 10 15
Ala Glu Leu Pro Ile Pro Ile Tyr Asn Gly Asn Thr Ile Met Asp Phe
20 25 30
Lys Lys Leu Ala Ser Leu Tyr Lys Asp Glu Leu Leu Asp Asn Val Leu
35 40 45
Pro Phe Trp Leu Glu His Ser Gln Asp His Glu Tyr Gly Gly Tyr Phe
50 55 60
Thr Cys Leu Asp Arg Glu Gly Lys Val Phe Asp Thr Asp Lys Phe Ile
65 70 75 80
Trp Leu Gln Ser Arg Glu Val Trp Met Phe Ser Met Leu Tyr Asn Lys
85 90 95
Val Glu Lys Arg Gln Glu Trp Leu Asp Cys Ala Ile Gln Gly Gly Glu
100 105 110
Phe Leu Lys Lys Tyr Gly His Asp Gly Asn Tyr Asn Trp Tyr Phe Ser
115 120 125
Leu Asp Arg Ser Gly Arg Pro Leu Val Glu Pro Tyr Asn Ile Phe Ser
130 135 140
Tyr Thr Phe Ala Thr Met Ala Phe Gly Gln Leu Ser Leu Thr Thr Gly
145 150 155 160
Asn Gln Glu Tyr Ala Asp Ile Ala Lys Lys Thr Phe Asp Ile Ile Leu
165 170 175
Ser Lys Val Asp Asn Pro Lys Gly Arg Trp Asn Lys Leu His Pro Gly
180 185 190
Thr Arg Asn Leu Lys Asn Phe Ala Leu Pro Met Ile Leu Cys Asn Leu
195 200 205
Ala Leu Glu Ile Glu His Leu Leu Asp Glu Thr Tyr Leu Arg Glu Thr
210 215 220
Met Asp Thr Cys Ile His Glu Val Met Glu Val Phe Tyr Arg Pro Glu
225 230 235 240
Leu Gly Gly Ile Ile Val Glu Asn Val Asp Ile Asp Gly Asn Leu Val
245 250 255
Asp Cys Phe Glu Gly Arg Gln Val Thr Pro Gly His Ala Ile Glu Ala
260 265 270
Met Trp Phe Ile Met Asp Leu Gly Lys Arg Leu Asn Arg Pro Glu Leu
275 280 285
Ile Glu Lys Ala Lys Glu Thr Thr Leu Thr Met Leu Asn Tyr Gly Trp
290 295 300
Asp Lys Gln Tyr Gly Gly Ile Tyr Tyr Phe Met Asp Arg Asn Gly Cys
305 310 315 320
Pro Pro Gln Gln Leu Glu Trp Asp Gln Lys Leu Trp Trp Val His Ile
325 330 335
Glu Thr Leu Ile Ser Leu Leu Lys Gly Tyr Gln Leu Thr Gly Asp Lys
340 345 350
Lys Cys Leu Glu Trp Phe Glu Lys Val His Asp Tyr Thr Trp Glu His
355 360 365
Phe Lys Asp Lys Glu Tyr Pro Glu Trp Tyr Gly Tyr Leu Asn Arg Arg
370 375 380
Gly Glu Val Leu Leu Pro Leu Lys Gly Gly Lys Trp Lys Gly Cys Phe
385 390 395 400
His Val Pro Arg Gly Leu Tyr Gln Cys Trp Lys Thr Leu Glu Glu Ile
405 410 415
Lys Asn Ile Val Ser
420
<210> 84
<211> 391
<212> PRT
<213> Synechocystis sp.
<400> 84
Met Ile Ala His Arg Arg Gln Glu Leu Ala Gln Gln Tyr Tyr Gln Ala
1 5 10 15
Leu His Gln Asp Val Leu Pro Phe Trp Glu Lys Tyr Ser Leu Asp Arg
20 25 30
Gln Gly Gly Gly Tyr Phe Thr Cys Leu Asp Arg Lys Gly Gln Val Phe
35 40 45
Asp Thr Asp Lys Phe Ile Trp Leu Gln Asn Arg Gln Val Trp Gln Phe
50 55 60
Ala Val Phe Tyr Asn Arg Leu Glu Pro Lys Pro Gln Trp Leu Glu Ile
65 70 75 80
Ala Arg His Gly Ala Asp Phe Leu Ala Arg His Gly Arg Asp Gln Asp
85 90 95
Gly Asn Trp Tyr Phe Ala Leu Asp Gln Glu Gly Lys Pro Leu Arg Gln
100 105 110
Pro Tyr Asn Val Phe Ser Asp Cys Phe Ala Ala Met Ala Phe Ser Gln
115 120 125
Tyr Ala Leu Ala Ser Gly Ala Gln Glu Ala Lys Ala Ile Ala Leu Gln
130 135 140
Ala Tyr Asn Asn Val Leu Arg Arg Gln His Asn Pro Lys Gly Gln Tyr
145 150 155 160
Glu Lys Ser Tyr Pro Gly Thr Arg Pro Leu Lys Ser Leu Ala Val Pro
165 170 175
Met Ile Leu Ala Asn Leu Thr Leu Glu Met Glu Trp Leu Leu Pro Pro
180 185 190
Thr Thr Val Glu Glu Val Leu Ala Gln Thr Val Arg Glu Val Met Thr
195 200 205
Asp Phe Leu Asp Pro Glu Ile Gly Leu Met Arg Glu Ala Val Thr Pro
210 215 220
Thr Gly Glu Phe Val Asp Ser Phe Glu Gly Arg Leu Leu Asn Pro Gly
225 230 235 240
His Gly Ile Glu Ala Met Trp Phe Met Met Asp Ile Ala Gln Arg Ser
245 250 255
Gly Asp Arg Gln Leu Gln Glu Gln Ala Ile Ala Val Val Leu Asn Thr
260 265 270
Leu Glu Tyr Ala Trp Asp Glu Glu Phe Gly Gly Ile Phe Tyr Phe Leu
275 280 285
Asp Arg Gln Gly His Pro Pro Gln Gln Leu Glu Trp Asp Gln Lys Leu
290 295 300
Trp Trp Val His Leu Glu Thr Leu Val Ala Leu Ala Lys Gly His Gln
305 310 315 320
Ala Thr Gly Gln Glu Lys Cys Trp Gln Trp Phe Glu Arg Val His Asp
325 330 335
Tyr Ala Trp Ser His Phe Ala Asp Pro Glu Tyr Gly Glu Trp Phe Gly
340 345 350
Tyr Leu Asn Arg Arg Gly Glu Val Leu Leu Asn Leu Lys Gly Gly Lys
355 360 365
Trp Lys Gly Cys Phe His Val Pro Arg Ala Leu Trp Leu Cys Ala Glu
370 375 380
Thr Leu Gln Leu Pro Val Ser
385 390
<210> 85
<211> 1266
<212> DNA
<213> Bacteroides ovatus
<400> 85
atggatagta agaataacat tggtcattca gcagacatct ctttaactgc tgaattaccc 60
ataccaatct ataatggaaa tacgattatg gatttcaaaa aactggcaag tctgtacaag 120
gatgagctcc tggacaacgt ccttcctttc tggcttgaac attcacaaga ccatgagtat 180
ggtggttact tcacctgtct ggaccgtgaa ggaaaagtat tcgatacgga taagtttatt 240
tggctgcaaa gtcgtgaggt atggatgttc tccatgcttt acaacaaagt ggagaaacgt 300
caggaatggc tagactgtgc cattcagggt ggcgaatttc taaaaaaata tggacatgac 360
ggcaattata actggtattt ttccctcgac cgttcgggta gaccattggt agaaccgtac 420
aatatattct cgtatacatt cgctaccatg gctttcggac agttgagcct tacaaccggt 480
aatcaggaat atgcggacat tgccaagaaa actttcgata taatcctttc caaagtggat 540
aatccgaaag ggagatggaa taagcttcat ccgggtaccc gtaatctgaa gaactttgcc 600
ttgccaatga tcctctgtaa cttggcactg gagatagagc atttattgga tgaaacgtat 660
ctgcgggaaa caatggatac ttgtatccat gaagtgatgg aagttttcta tcgtcctgaa 720
ctcggaggta tcattgttga aaacgtggac atagacggta atttggtcga ttgttttgaa 780
ggccgtcagg tgaccccggg acatgccatt gaagcgatgt ggtttatcat ggatctaggc 840
aagcgtctga atcgtccgga attgatagag aaagccaaag agactactct cacgatgctt 900
aattatggct gggacaagca atatggaggt atctactatt ttatggatcg taacggttgt 960
cctccccaac aattggagtg ggaccagaaa ctctggtggg tccatatcga aacgcttatt 1020
tccctgctga aaggctatca attgacggga gacaaaaaat gcttggaatg gtttgaaaag 1080
gtacatgact acacttggga gcatttcaag gataaagaat atcctgaatg gtatggctac 1140
ttgaaccgaa gaggcgaagt attgctacca ctcaaaggag gaaaatggaa aggatgcttc 1200
catgtgccaa gaggactgta tcagtgctgg aaaacattag aagaaataaa aaatatagta 1260
tcctaa 1266
<210> 86
<211> 1176
<212> DNA
<213> Synechocystis sp.
<400> 86
atgattgccc atcgccgtca ggagttagcc cagcaatatt accaggcttt acaccaggac 60
gtattgccct tttgggaaaa atattccctc gatcgccagg ggggcggtta ctttacctgc 120
ttagaccgta aaggccaggt ttttgacaca gataaattca tttggttaca aaaccgtcag 180
gtatggcagt ttgccgtttt ctacaaccgt ttggaaccaa aaccccaatg gttagaaatt 240
gcccgccatg gtgctgattt tttagctcgc cacggccgag atcaagacgg taattggtat 300
tttgctttgg atcaggaagg caaacccctg cgtcaaccct ataacgtttt ttccgattgc 360
ttcgccgcca tggcctttag tcaatatgcc ttagccagtg gggcgcagga agctaaagcc 420
attgccctgc aggcctacaa taacgtccta cgccgtcagc acaatcccaa aggtcaatac 480
gagaagtcct atccaggtac tagacccctc aaatccctgg cggtgccgat gattttagcc 540
aacctcaccc tggagatgga atggttatta ccgcctacta ccgtggaaga ggtgttggcc 600
caaaccgtca gagaagtgat gacggatttc ctcgacccag aaataggatt aatgcgggaa 660
gcggtgaccc ccacaggaga atttgttgat agttttgaag ggcggttgct caacccagga 720
cacggcattg aagccatgtg gttcatgatg gacattgccc aacgctccgg cgatcgccag 780
ttacaggagc aagccattgc agtggtgttg aacaccctgg aatatgcctg ggatgaagaa 840
tttggtggca tattttattt ccttgatcgc cagggccacc ctccccaaca actggaatgg 900
gaccaaaagc tctggtgggt acatttggaa accctggttg ccctagccaa gggccaccaa 960
gccactggcc aagaaaaatg ttggcaatgg tttgagcggg tccatgatta cgcctggagt 1020
catttcgccg atcctgagta tggggaatgg tttggctacc tgaatcgccg gggagaggtg 1080
ttactcaacc taaaaggggg gaaatggaaa gggtgcttcc acgtgccccg agctctgtgg 1140
ctctgtgcgg aaactctcca acttccggtt agttaa 1176
<210> 87
<211> 229
<212> PRT
<213> Escherichia coli
<400> 87
Met Ser Leu Leu Ala Gln Leu Asp Gln Lys Ile Ala Ala Asn Gly Gly
1 5 10 15
Leu Ile Val Ser Cys Gln Pro Val Pro Asp Ser Pro Leu Asp Lys Pro
20 25 30
Glu Ile Val Ala Ala Met Ala Leu Ala Ala Glu Gln Ala Gly Ala Val
35 40 45
Ala Ile Arg Ile Glu Gly Val Ala Asn Leu Gln Ala Thr Arg Ala Val
50 55 60
Val Ser Val Pro Ile Ile Gly Ile Val Lys Arg Asp Leu Glu Asp Ser
65 70 75 80
Pro Val Arg Ile Thr Ala Tyr Ile Glu Asp Val Asp Ala Leu Ala Gln
85 90 95
Ala Gly Ala Asp Ile Ile Ala Ile Asp Gly Thr Asp Arg Pro Arg Pro
100 105 110
Val Pro Val Glu Thr Leu Leu Ala Arg Ile His His His Gly Leu Leu
115 120 125
Ala Met Thr Asp Cys Ser Thr Pro Glu Asp Gly Leu Ala Cys Gln Lys
130 135 140
Leu Gly Ala Glu Ile Ile Gly Thr Thr Leu Ser Gly Tyr Thr Thr Pro
145 150 155 160
Glu Thr Pro Glu Glu Pro Asp Leu Ala Leu Val Lys Thr Leu Ser Asp
165 170 175
Ala Gly Cys Arg Val Ile Ala Glu Gly Arg Tyr Asn Thr Pro Ala Gln
180 185 190
Ala Ala Asp Ala Met Arg His Gly Ala Trp Ala Val Thr Val Gly Ser
195 200 205
Ala Ile Thr Arg Leu Glu His Ile Cys Gln Trp Tyr Asn Thr Ala Met
210 215 220
Lys Lys Ala Val Leu
225
<210> 88
<211> 690
<212> DNA
<213> Escherichia coli
<400> 88
atgtcgttac ttgcacaact ggatcaaaaa atcgctgcta acggtggcct gattgtctcc 60
tgccagccgg ttccggacag cccgctcgat aaacccgaaa tcgtcgccgc catggcatta 120
gcggcagaac aggcgggcgc ggttgccatt cgcattgaag gtgtggcaaa tctgcaagcc 180
acgcgtgcgg tggtgagcgt gccgattatt ggaattgtga aacgcgatct ggaggattct 240
ccggtacgca tcacggccta tattgaagat gttgatgcgc tggcgcaggc gggcgcggac 300
attatcgcca ttgacggcac cgaccgcccg cgtccggtgc ctgttgaaac gctgctggca 360
cgtattcacc atcacggttt actggcgatg accgactgct caacgccgga agacggcctg 420
gcatgccaaa agctgggagc cgaaattatt ggcactacgc tttctggcta taccacgcct 480
gaaacgccag aagagccgga tctggcgctg gtgaaaacgt tgagcgacgc cggatgtcgg 540
gtgattgccg aagggcgtta caacacgcct gctcaggcgg cggatgcgat gcgccacggc 600
gcgtgggcgg tgacggtcgg ttctgcaatc acgcgtcttg agcacatttg tcagtggtac 660
aacacagcga tgaaaaaggc ggtgctatga 690
<210> 89
<211> 346
<212> PRT
<213> Campylobacter jejuni
<400> 89
Met Lys Glu Ile Lys Ile Gln Asn Ile Ile Ile Ser Glu Glu Lys Ala
1 5 10 15
Pro Leu Val Val Pro Glu Ile Gly Ile Asn His Asn Gly Ser Leu Glu
20 25 30
Leu Ala Lys Ile Met Val Asp Ala Ala Phe Ser Ala Gly Ala Lys Ile
35 40 45
Ile Lys His Gln Thr His Ile Val Glu Asp Glu Met Ser Lys Ala Ala
50 55 60
Lys Lys Val Ile Pro Gly Asn Ala Lys Ile Ser Ile Tyr Glu Ile Met
65 70 75 80
Gln Lys Cys Ala Leu Asp Tyr Lys Asp Glu Leu Ala Leu Lys Glu Tyr
85 90 95
Thr Glu Lys Leu Gly Leu Val Tyr Leu Ser Thr Pro Phe Ser Arg Ala
100 105 110
Gly Ala Asn Arg Leu Glu Asp Met Gly Val Ser Ala Phe Lys Ile Gly
115 120 125
Ser Gly Glu Cys Asn Asn Tyr Pro Leu Ile Lys His Ile Ala Ala Phe
130 135 140
Lys Lys Pro Met Ile Val Ser Thr Gly Met Asn Ser Ile Glu Ser Ile
145 150 155 160
Lys Pro Thr Val Lys Ile Leu Leu Asp Asn Glu Ile Pro Phe Val Leu
165 170 175
Met His Thr Thr Asn Leu Tyr Pro Thr Pro His Asn Leu Val Arg Leu
180 185 190
Asn Ala Met Leu Glu Leu Lys Lys Glu Phe Ser Cys Met Val Gly Leu
195 200 205
Ser Asp His Thr Thr Asp Asn Leu Ala Cys Leu Gly Ala Val Val Leu
210 215 220
Gly Ala Cys Val Leu Glu Arg His Phe Thr Asp Ser Met His Arg Ser
225 230 235 240
Gly Pro Asp Ile Val Cys Ser Met Asp Thr Lys Ala Leu Lys Glu Leu
245 250 255
Ile Ile Gln Ser Glu Gln Met Ala Ile Ile Arg Gly Asn Asn Glu Ser
260 265 270
Lys Lys Ala Ala Lys Gln Glu Gln Val Thr Ile Asp Phe Ala Phe Ala
275 280 285
Ser Val Val Ser Ile Lys Asp Ile Lys Lys Gly Glu Val Leu Ser Met
290 295 300
Asp Asn Ile Trp Val Lys Arg Pro Gly Leu Gly Gly Ile Ser Ala Ala
305 310 315 320
Glu Phe Glu Asn Ile Leu Gly Lys Lys Ala Leu Arg Asp Ile Glu Asn
325 330 335
Asp Ala Gln Leu Ser Tyr Glu Asp Phe Ala
340 345
<210> 90
<211> 1041
<212> DNA
<213> Campylobacter jejuni
<400> 90
atgaaagaaa taaaaataca aaatataatc ataagtgaag aaaaagcacc cttagtcgtg 60
cctgaaatag gcattaatca taatggcagt ttagaactag ctaaaattat ggtagatgca 120
gcctttagcg caggtgctaa gattataaag catcaaaccc acatcgttga agatgagatg 180
agtaaggccg ctaaaaaagt aattcctggt aatgcaaaaa taagcattta tgagattatg 240
caaaaatgtg ctttagatta taaagatgag ctagcactta aagaatacac agaaaaatta 300
ggtcttgttt atcttagcac acctttttct cgtgcaggtg caaaccgctt agaagatatg 360
ggagttagtg cttttaagat tggttcaggt gagtgtaata attatccgct tattaaacac 420
atagcagcct ttaaaaagcc tatgatagtt agcacaggaa tgaatagtat tgaaagtata 480
aaaccaactg taaaaatctt attagacaat gaaattccct ttgttttaat gcactcgacc 540
aatctttacc caaccccgca taatcttgta agattaaacg ctatgcttga attaaaaaaa 600
gaattttctt gcatggtagg cttaagcgac cacacaacag ataatcttgc gtgtttaggt 660
gcggttgcac ttggtgcttg tgtgcttgaa agacatttta ctgatagtat gcatagaagt 720
ggccctgata tagtttgttc tatggataca aaggctttaa aagagctaat tatccaaagt 780
gagcaaatgg ctataatgaa aggaaataat gaaagcaaaa aagcagctaa gcaagaacaa 840
gttacaattg attttgcctt tgcaagcgta gttagcatta aagatattaa aaaaggcgaa 900
gttttatcta tggacaatat ctgggttaaa agacctggac ttggtggaat tagtgcggct 960
gaatttgaaa atattttagg caaaaaagca ttaagagata tagaaaatga tactcagtta 1020
agctatgagg attttgcgtg a 1041
<210> 91
<211> 221
<212> PRT
<213> Escherichia coli
<400> 91
Met Ser Leu Ala Ile Ile Pro Ala Arg Gly Gly Ser Lys Gly Ile Lys
1 5 10 15
Asn Lys Asn Leu Val Leu Leu Asn Asn Lys Pro Leu Ile Tyr Tyr Thr
20 25 30
Ile Lys Ala Ala Leu Asn Ala Lys Ser Ile Ser Lys Val Val Val Ser
35 40 45
Ser Asp Ser Asp Glu Ile Leu Asn Tyr Ala Lys Ser Gln Asn Val Asp
50 55 60
Ile Leu Lys Arg Pro Ile Ser Leu Ala Gln Asp Asp Thr Thr Ser Asp
65 70 75 80
Lys Val Leu Leu His Ala Leu Lys Phe Tyr Lys Asp Tyr Glu Asp Val
85 90 95
Val Phe Leu Gln Pro Thr Ser Pro Leu Arg Thr Asn Ile His Ile Asn
100 105 110
Glu Ala Phe Asn Leu Tyr Lys Asn Ser Asn Ala Asn Ala Leu Ile Ser
115 120 125
Val Ser Glu Cys Asp Asn Lys Ile Leu Lys Ala Phe Val Cys Asn Asp
130 135 140
Cys Gly Asp Leu Ala Gly Ile Cys Asn Asp Glu Tyr Pro Phe Met Pro
145 150 155 160
Arg Gln Lys Leu Pro Lys Thr Tyr Met Ser Asn Gly Ala Ile Tyr Ile
165 170 175
Leu Lys Ile Lys Glu Phe Leu Asn Asn Pro Ser Phe Leu Gln Ser Lys
180 185 190
Thr Lys His Phe Leu Met Asp Glu Ser Ser Ser Leu Asp Ile Asp Cys
195 200 205
Leu Glu Asp Leu Lys Lys Val Glu Gln Ile Trp Lys Lys
210 215 220
<210> 92
<211> 666
<212> DNA
<213> Escherichia coli
<400> 92
atgagcctgg ccattatccc ggcacgtggc ggttctaaag gcatcaaaaa caaaaacctg 60
gttctgctga acaataaacc gctgatttat tacaccatca aagcggccct gaacgccaaa 120
agtattagca aagtggttgt gagctctgat tctgatgaaa tcctgaacta cgcaaaaagt 180
cagaacgttg atatcctgaa acgtccgatc agtctggcac aggatgatac cacgagcgat 240
aaagtgctgc tgcatgcgct gaaattctac aaagattacg aagatgttgt gttcctgcag 300
ccgaccagcc cgctgcgtac gaatattcac atcaacgaag cgttcaacct gtacaaaaac 360
agcaacgcaa acgcgctgat ttctgttagt gaatgcgata acaaaatcct gaaagcgttt 420
gtgtgcaatg attgtggcga tctggccggt atttgtaacg atgaataccc gttcatgccg 480
cgccagaaac tgccgaaaac ctatatgagc aatggtgcca tctacatcct gaaaatcaaa 540
gaattcctga acaacccgag cttcctgcag tctaaaacga aacatttcct gatggatgaa 600
agtagctctc tggatattga ttgcctggaa gatctgaaaa aagtggaaca gatctggaaa 660
aaataa 666
<210> 93
<211> 417
<212> PRT
<213> Escherichia coli
<400> 93
Met Tyr Tyr Leu Lys Asn Thr Asn Phe Trp Met Phe Gly Leu Phe Phe
1 5 10 15
Phe Phe Tyr Phe Phe Ile Met Gly Ala Tyr Phe Pro Phe Phe Pro Ile
20 25 30
Trp Leu His Asp Ile Asn His Ile Ser Lys Ser Asp Thr Gly Ile Ile
35 40 45
Phe Ala Ala Ile Ser Leu Phe Ser Leu Leu Phe Gln Pro Leu Phe Gly
50 55 60
Leu Leu Ser Asp Lys Leu Gly Leu Arg Lys Tyr Leu Leu Trp Ile Ile
65 70 75 80
Thr Gly Met Leu Val Met Phe Ala Pro Phe Phe Ile Phe Ile Phe Gly
85 90 95
Pro Leu Leu Gln Tyr Asn Ile Leu Val Gly Ser Ile Val Gly Gly Ile
100 105 110
Tyr Leu Gly Phe Cys Phe Asn Ala Gly Ala Pro Ala Val Glu Ala Phe
115 120 125
Ile Glu Lys Val Ser Arg Arg Ser Asn Phe Glu Phe Gly Arg Ala Arg
130 135 140
Met Phe Gly Cys Val Gly Trp Ala Leu Cys Ala Ser Ile Val Gly Ile
145 150 155 160
Met Phe Thr Ile Asn Asn Gln Phe Val Phe Trp Leu Gly Ser Gly Cys
165 170 175
Ala Leu Ile Leu Ala Val Leu Leu Phe Phe Ala Lys Thr Asp Ala Pro
180 185 190
Ser Ser Ala Thr Val Ala Asn Ala Val Gly Ala Asn His Ser Ala Phe
195 200 205
Ser Leu Lys Leu Ala Leu Glu Leu Phe Arg Gln Pro Lys Leu Trp Phe
210 215 220
Leu Ser Leu Tyr Val Ile Gly Val Ser Cys Thr Tyr Asp Val Phe Asp
225 230 235 240
Gln Gln Phe Ala Asn Phe Phe Thr Ser Phe Phe Ala Thr Gly Glu Gln
245 250 255
Gly Thr Arg Val Phe Gly Tyr Val Thr Thr Met Gly Glu Leu Leu Asn
260 265 270
Ala Ser Ile Met Phe Phe Ala Pro Leu Ile Ile Asn Arg Ile Gly Gly
275 280 285
Lys Asn Ala Leu Leu Leu Ala Gly Thr Ile Met Ser Val Arg Ile Ile
290 295 300
Gly Ser Ser Phe Ala Thr Ser Ala Leu Glu Val Val Ile Leu Lys Thr
305 310 315 320
Leu His Met Phe Glu Val Pro Phe Leu Leu Val Gly Cys Phe Lys Tyr
325 330 335
Ile Thr Ser Gln Phe Glu Val Arg Phe Ser Ala Thr Ile Tyr Leu Val
340 345 350
Cys Phe Cys Phe Phe Lys Gln Leu Ala Met Ile Phe Met Ser Val Leu
355 360 365
Ala Gly Asn Met Tyr Glu Ser Ile Gly Phe Gln Gly Ala Tyr Leu Val
370 375 380
Leu Gly Leu Val Ala Leu Gly Phe Thr Leu Ile Ser Val Phe Thr Leu
385 390 395 400
Ser Gly Pro Gly Pro Leu Ser Leu Leu Arg Arg Gln Val Asn Glu Val
405 410 415
Ala
<210> 94
<211> 1254
<212> DNA
<213> Escherichia coli
<400> 94
atgtactatt taaaaaacac aaacttttgg atgttcggtt tattcttttt cttttacttt 60
tttatcatgg gagcctactt cccgtttttc ccgatttggc tacatgacat caaccatatc 120
agcaaaagtg atacgggtat tatttttgcc gctatttctc tgttctcgct attattccaa 180
ccgctgtttg gtctgctttc tgacaaactc gggctgcgca aatacctgct gtggattatt 240
accggcatgt tagtgatgtt tgcgccgttc tttattttta tcttcgggcc actgttacaa 300
tacaacattt tagtaggatc gattgttggt ggtatttatc taggcttttg ttttaacgcc 360
ggtgcgccag cagtagaggc atttattgag aaagtcagcc gtcgcagtaa tttcgaattt 420
ggtcgcgcgc ggatgtttgg ctgtgttggc tgggcgctgt gtgcctcgat tgtcggcatc 480
atgttcacca tcaataatca gtttgttttc tggctgggct ctggctgtgc actcatcctc 540
gccgttttac tctttttcgc caaaacggat gcgccctctt ctgccacggt tgccaatgcg 600
gtaggtgcca accattcggc atttagcctt aagctggcac tggaactgtt cagacagcca 660
aaactgtggt ttttgtcact gtatgttatt ggcgtttcct gcacctacga tgtttttgac 720
caacagtttg ctaatttctt tacttcgttc tttgctaccg gtgaacaggg tacgcgggta 780
tttggctacg taacgacaat gggcgaatta cttaacgcct cgattatgtt ctttgcgcca 840
ctgatcatta atcgcatcgg tgggaaaaac gccctgctgc tggctggcac tattatgtct 900
gtacgtatta ttggctcatc gttcgccacc tcagcgctgg aagtggttat tctgaaaacg 960
ctgcatatgt ttgaagtacc gttcctgctg gtgggctgct ttaaatatat taccagccag 1020
tttgaagtgc gtttttcagc gacgatttat ctggtctgtt tctgcttctt taagcaactg 1080
gcgatgattt ttatgtctgt actggcgggc aatatgtatg aaagcatcgg tttccagggc 1140
gcttatctgg tgctgggtct ggtggcgctg ggcttcacct taatttccgt gttcacgctt 1200
agcggccccg gtccgctttc tctactgcgt cgtcaggtga atgaagtcgc ttaa 1254
<210> 95
<211> 1024
<212> PRT
<213> Escherichia coli
<400> 95
Met Thr Met Ile Thr Asp Ser Leu Ala Val Val Leu Gln Arg Arg Asp
1 5 10 15
Trp Glu Asn Pro Gly Val Thr Gln Leu Asn Arg Leu Ala Ala His Pro
20 25 30
Pro Phe Ala Ser Trp Arg Asn Ser Glu Glu Ala Arg Thr Asp Arg Pro
35 40 45
Ser Gln Gln Leu Arg Ser Leu Asn Gly Glu Trp Arg Phe Ala Trp Phe
50 55 60
Pro Ala Pro Glu Ala Val Pro Glu Ser Trp Leu Glu Cys Asp Leu Pro
65 70 75 80
Glu Ala Asp Thr Val Val Val Pro Ser Asn Trp Gln Met His Gly Tyr
85 90 95
Asp Ala Pro Ile Tyr Thr Asn Val Thr Tyr Pro Ile Thr Val Asn Pro
100 105 110
Pro Phe Val Pro Thr Glu Asn Pro Thr Gly Cys Tyr Ser Leu Thr Phe
115 120 125
Asn Val Asp Glu Ser Trp Leu Gln Glu Gly Gln Thr Arg Ile Ile Phe
130 135 140
Asp Gly Val Asn Ser Ala Phe His Leu Trp Cys Asn Gly Arg Trp Val
145 150 155 160
Gly Tyr Gly Gln Asp Ser Arg Leu Pro Ser Glu Phe Asp Leu Ser Ala
165 170 175
Phe Leu Arg Ala Gly Glu Asn Arg Leu Ala Val Met Val Leu Arg Trp
180 185 190
Ser Asp Gly Ser Tyr Leu Glu Asp Gln Asp Met Trp Arg Met Ser Gly
195 200 205
Ile Phe Arg Asp Val Ser Leu Leu His Lys Pro Thr Thr Gln Ile Ser
210 215 220
Asp Phe His Val Ala Thr Arg Phe Asn Asp Asp Phe Ser Arg Ala Val
225 230 235 240
Leu Glu Ala Glu Val Gln Met Cys Gly Glu Leu Arg Asp Tyr Leu Arg
245 250 255
Val Thr Val Ser Leu Trp Gln Gly Glu Thr Gln Val Ala Ser Gly Thr
260 265 270
Ala Pro Phe Gly Gly Glu Ile Ile Asp Glu Arg Gly Gly Tyr Ala Asp
275 280 285
Arg Val Thr Leu Arg Leu Asn Val Glu Asn Pro Lys Leu Trp Ser Ala
290 295 300
Glu Ile Pro Asn Leu Tyr Arg Ala Val Val Glu Leu His Thr Ala Asp
305 310 315 320
Gly Thr Leu Ile Glu Ala Glu Ala Cys Asp Val Gly Phe Arg Glu Val
325 330 335
Arg Ile Glu Asn Gly Leu Leu Leu Leu Asn Gly Lys Pro Leu Leu Ile
340 345 350
Arg Gly Val Asn Arg His Glu His His Pro Leu His Gly Gln Val Met
355 360 365
Asp Glu Gln Thr Met Val Gln Asp Ile Leu Leu Met Lys Gln Asn Asn
370 375 380
Phe Asn Ala Val Arg Cys Ser His Tyr Pro Asn His Pro Leu Trp Tyr
385 390 395 400
Thr Leu Cys Asp Arg Tyr Gly Leu Tyr Val Val Asp Glu Ala Asn Ile
405 410 415
Glu Thr His Gly Met Val Pro Met Asn Arg Leu Thr Asp Asp Pro Arg
420 425 430
Trp Leu Pro Ala Met Ser Glu Arg Val Thr Arg Met Val Gln Arg Asp
435 440 445
Arg Asn His Pro Ser Val Ile Ile Trp Ser Leu Gly Asn Glu Ser Gly
450 455 460
His Gly Ala Asn His Asp Ala Leu Tyr Arg Trp Ile Lys Ser Val Asp
465 470 475 480
Pro Ser Arg Pro Val Gln Tyr Glu Gly Gly Gly Ala Asp Thr Thr Ala
485 490 495
Thr Asp Ile Ile Cys Pro Met Tyr Ala Arg Val Asp Glu Asp Gln Pro
500 505 510
Phe Pro Ala Val Pro Lys Trp Ser Ile Lys Lys Trp Leu Ser Leu Pro
515 520 525
Gly Glu Thr Arg Pro Leu Ile Leu Cys Glu Tyr Ala His Ala Met Gly
530 535 540
Asn Ser Leu Gly Gly Phe Ala Lys Tyr Trp Gln Ala Phe Arg Gln Tyr
545 550 555 560
Pro Arg Leu Gln Gly Gly Phe Val Trp Asp Trp Val Asp Gln Ser Leu
565 570 575
Ile Lys Tyr Asp Glu Asn Gly Asn Pro Trp Ser Ala Tyr Gly Gly Asp
580 585 590
Phe Gly Asp Thr Pro Asn Asp Arg Gln Phe Cys Met Asn Gly Leu Val
595 600 605
Phe Ala Asp Arg Thr Pro His Pro Ala Leu Thr Glu Ala Lys His Gln
610 615 620
Gln Gln Phe Phe Gln Phe Arg Leu Ser Gly Gln Thr Ile Glu Val Thr
625 630 635 640
Ser Glu Tyr Leu Phe Arg His Ser Asp Asn Glu Leu Leu His Trp Met
645 650 655
Val Ala Leu Asp Gly Lys Pro Leu Ala Ser Gly Glu Val Pro Leu Asp
660 665 670
Val Ala Pro Gln Gly Lys Gln Leu Ile Glu Leu Pro Glu Leu Pro Gln
675 680 685
Pro Glu Ser Ala Gly Gln Leu Trp Leu Thr Val Arg Val Val Gln Pro
690 695 700
Asn Ala Thr Ala Trp Ser Glu Ala Gly His Ile Ser Ala Trp Gln Gln
705 710 715 720
Trp Arg Leu Ala Glu Asn Leu Ser Val Thr Leu Pro Ala Ala Ser His
725 730 735
Ala Ile Pro His Leu Thr Thr Ser Glu Met Asp Phe Cys Ile Glu Leu
740 745 750
Gly Asn Lys Arg Trp Gln Phe Asn Arg Gln Ser Gly Phe Leu Ser Gln
755 760 765
Met Trp Ile Gly Asp Lys Lys Gln Leu Leu Thr Pro Leu Arg Asp Gln
770 775 780
Phe Thr Arg Ala Pro Leu Asp Asn Asp Ile Gly Val Ser Glu Ala Thr
785 790 795 800
Arg Ile Asp Pro Asn Ala Trp Val Glu Arg Trp Lys Ala Ala Gly His
805 810 815
Tyr Gln Ala Glu Ala Ala Leu Leu Gln Cys Thr Ala Asp Thr Leu Ala
820 825 830
Asp Ala Val Leu Ile Thr Thr Ala His Ala Trp Gln His Gln Gly Lys
835 840 845
Thr Leu Phe Ile Ser Arg Lys Thr Tyr Arg Ile Asp Gly Ser Gly Gln
850 855 860
Met Ala Ile Thr Val Asp Val Glu Val Ala Ser Asp Thr Pro His Pro
865 870 875 880
Ala Arg Ile Gly Leu Asn Cys Gln Leu Ala Gln Val Ala Glu Arg Val
885 890 895
Asn Trp Leu Gly Leu Gly Pro Gln Glu Asn Tyr Pro Asp Arg Leu Thr
900 905 910
Ala Ala Cys Phe Asp Arg Trp Asp Leu Pro Leu Ser Asp Met Tyr Thr
915 920 925
Pro Tyr Val Phe Pro Ser Glu Asn Gly Leu Arg Cys Gly Thr Arg Glu
930 935 940
Leu Asn Tyr Gly Pro His Gln Trp Arg Gly Asp Phe Gln Phe Asn Ile
945 950 955 960
Ser Arg Tyr Ser Gln Gln Gln Leu Met Glu Thr Ser His Arg His Leu
965 970 975
Leu His Ala Glu Glu Gly Thr Trp Leu Asn Ile Asp Gly Phe His Met
980 985 990
Gly Ile Gly Gly Asp Asp Ser Trp Ser Pro Ser Val Ser Ala Glu Phe
995 1000 1005
Gln Leu Ser Ala Gly Arg Tyr His Tyr Gln Leu Val Trp Cys Gln
1010 1015 1020
Lys
<210> 96
<211> 3075
<212> DNA
<213> Escherichia coli
<400> 96
atgaccatga ttacggattc actggccgtc gttttacaac gtcgtgactg ggaaaaccct 60
ggcgttaccc aacttaatcg ccttgcagca catccccctt tcgccagctg gcgtaatagc 120
gaagaggccc gcaccgatcg cccttcccaa cagttgcgca gcctgaatgg cgaatggcgc 180
tttgcctggt ttccggcacc agaagcggtg ccggaaagct ggctggagtg cgatcttcct 240
gaggccgata ctgtcgtcgt cccctcaaac tggcagatgc acggttacga tgcgcccatc 300
tacaccaacg tgacctatcc cattacggtc aatccgccgt ttgttcccac ggagaatccg 360
acgggttgtt actcgctcac atttaatgtt gatgaaagct ggctacagga aggccagacg 420
cgaattattt ttgatggcgt taactcggcg tttcatctgt ggtgcaacgg gcgctgggtc 480
ggttacggcc aggacagtcg tttgccgtct gaatttgacc tgagcgcatt tttacgcgcc 540
ggagaaaacc gcctcgcggt gatggtgctg cgctggagtg acggcagtta tctggaagat 600
caggatatgt ggcggatgag cggcattttc cgtgacgtct cgttgctgca taaaccgact 660
acacaaatca gcgatttcca tgttgccact cgctttaatg atgatttcag ccgcgctgta 720
ctggaggctg aagttcagat gtgcggcgag ttgcgtgact acctacgggt aacagtttct 780
ttatggcagg gtgaaacgca ggtcgccagc ggcaccgcgc ctttcggcgg tgaaattatc 840
gatgagcgtg gtggttatgc cgatcgcgtc acactacgtc tgaacgtcga aaacccgaaa 900
ctgtggagcg ccgaaatccc gaatctctat cgtgcggtgg ttgaactgca caccgccgac 960
ggcacgctga ttgaagcaga agcctgcgat gtcggtttcc gcgaggtgcg gattgaaaat 1020
ggtctgctgc tgctgaacgg caagccgttg ctgattcgag gcgttaaccg tcacgagcat 1080
catcctctgc atggtcaggt catggatgag cagacgatgg tgcaggatat cctgctgatg 1140
aagcagaaca actttaacgc cgtgcgctgt tcgcattatc cgaaccatcc gctgtggtac 1200
acgctgtgcg accgctacgg cctgtatgtg gtggatgaag ccaatattga aacccacggc 1260
atggtgccaa tgaatcgtct gaccgatgat ccgcgctggc taccggcgat gagcgaacgc 1320
gtaacgcgaa tggtgcagcg cgatcgtaat cacccgagtg tgatcatctg gtcgctgggg 1380
aatgaatcag gccacggcgc taatcacgac gcgctgtatc gctggatcaa atctgtcgat 1440
ccttcccgcc cggtgcagta tgaaggcggc ggagccgaca ccacggccac cgatattatt 1500
tgcccgatgt acgcgcgcgt ggatgaagac cagcccttcc cggctgtgcc gaaatggtcc 1560
atcaaaaaat ggctttcgct acctggagag acgcgcccgc tgatcctttg cgaatacgcc 1620
cacgcgatgg gtaacagtct tggcggtttc gctaaatact ggcaggcgtt tcgtcagtat 1680
ccccgtttac agggcggctt cgtctgggac tgggtggatc agtcgctgat taaatatgat 1740
gaaaacggca acccgtggtc ggcttacggc ggtgattttg gcgatacgcc gaacgatcgc 1800
cagttctgta tgaacggtct ggtctttgcc gaccgcacgc cgcatccagc gctgacggaa 1860
gcaaaacacc agcagcagtt tttccagttc cgtttatccg ggcaaaccat cgaagtgacc 1920
agcgaatacc tgttccgtca tagcgataac gagctcctgc actggatggt ggcgctggat 1980
ggtaagccgc tggcaagcgg tgaagtgcct ctggatgtcg ctccacaagg taaacagttg 2040
attgaactgc ctgaactacc gcagccggag agcgccgggc aactctggct cacagtacgc 2100
gtagtgcaac cgaacgcgac cgcatggtca gaagccggac acatcagcgc ctggcagcag 2160
tggcgtctgg ctgaaaacct cagcgtgaca ctccccgccg cgtcccacgc catcccgcat 2220
ctgaccacca gcgaaatgga tttttgcatc gagctgggta ataagcgttg gcaatttaac 2280
cgccagtcag gctttctttc acagatgtgg attggcgata aaaaacaact gctgacgccg 2340
ctgcgcgatc agttcacccg tgcaccgctg gataacgaca ttggcgtaag tgaagcgacc 2400
cgcattgacc ctaacgcctg ggtcgaacgc tggaaggcgg cgggccatta ccaggccgaa 2460
gcagcgttgt tgcagtgcac ggcagataca cttgctgatg cggtgctgat tacgaccgct 2520
cacgcgtggc agcatcaggg gaaaacctta tttatcagcc ggaaaaccta ccggattgat 2580
ggtagtggtc aaatggcgat taccgttgat gttgaagtgg cgagcgatac accgcatccg 2640
gcgcggattg gcctgaactg ccagctggcg caggtagcag agcgggtaaa ctggctcgga 2700
ttagggccgc aagaaaacta tcccgaccgc cttactgccg cctgttttga ccgctgggat 2760
ctgccattgt cagacatgta taccccgtac gtcttcccga gcgaaaacgg tctgcgctgc 2820
gggacgcgcg aattgaatta tggcccacac cagtggcgcg gcgacttcca gttcaacatc 2880
agccgctaca gtcaacagca actgatggaa accagccatc gccatctgct gcacgcggaa 2940
gaaggcacat ggctgaatat cgacggtttc catatgggga ttggtggcga cgactcctgg 3000
agcccgtcag tatcggcgga attccagctg agcgccggtc gctaccatta ccagttggtc 3060
tggtgtcaaa aataa 3075
<210> 97
<211> 3123
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 97
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgatcgct 180
caccgtcgtc aggaactggc tcaacagtat tatcaggctc tgcaccaaga tgtgctgccg 240
ttctgggaaa agtattcgct ggatcgtcaa ggcggtggct attttacctg cctggaccgc 300
aagggtcagg tttttgatac ggacaagttc atttggctgc aaaaccgtca agtgtggcaa 360
tttgcggttt tctacaatcg cctggaaccg aaaccgcagt ggctggaaat cgctcgtcat 420
ggtgcggatt ttctggcacg tcacggtcgt gatcaggacg gtaactggta tttcgccctg 480
gatcaggaag gcaaaccgct gcgccaaccg tacaatgtgt tttccgactg tttcgcggcg 540
atggcgttta gccagtatgc actggcttct ggtgctcaag aagcgaaggc cattgcactg 600
caagcgtata acaatgttct gcgtcgccag cataacccga aaggtcaata tgaaaagagt 660
tacccgggta cccgtccgct gaaatccctg gcagtgccga tgatcctggc taatctgacg 720
ctggaaatgg aatggctgct gccgccgacc acggtcgaag aagtgctggc ccagaccgtt 780
cgtgaagtca tgacggattt tctggacccg gaaattggcc tgatgcgcga agcagttacc 840
ccgacgggtg aatttgtcga ttcattcgaa ggccgcctgc tgaacccggg tcatggcatt 900
gaagcgatgt ggtttatgat ggatattgcc cagcgttcgg gtgaccgcca gctgcaagaa 960
caggctattg cggtggttct gaataccctg gaatatgcat gggatgaaga atttggtggc 1020
atcttttact tcctggaccg tcaaggtcac ccgccgcagc aactggaatg ggatcagaaa 1080
ctgtggtggg tccatctgga aaccctggtg gccctggcaa aaggtcacca ggcgacgggc 1140
caagaaaagt gctggcagtg gtttgaacgc gtgcatgatt atgcatggag ccactttgct 1200
gacccggaat atggtgaatg gttcggctac ctgaaccgtc gcggtgaagt gctgctgaat 1260
ctgaaaggtg gcaaatggaa gggctgcttc cacgttccgc gtgcgctgtg gctgtgtgcc 1320
gaaaccctgc aactgccggt ctcttaataa tcgaaggaga tacaacatga gcttacccga 1380
tggattttat ataaggcgaa tggaagaggg ggatttggaa caggtcactg agacgctaaa 1440
ggttttgacc accgtgggca ctattacccc cgaatccttc agcaaactca taaaatactg 1500
gaatgaagcc acagtatgga atgataacga agataaaaaa ataatgcaat ataaccccat 1560
ggtgattgtg gacaagcgca ccgagacggt tgccgctacg gggaatatca tcatcgaaag 1620
aaagatcatt catgaactgg ggctatgtgg ccacatcgag gacattgcag taaactccaa 1680
gtatcagggc caaggtttgg gcaagctctt gattgatcaa ttggtaacta tcggctttga 1740
ctacggttgt tataagatta ttttagattg cgatgagaaa aatgtcaaat tctatgaaaa 1800
atgtgggttt agcaacgcag gcgtggaaat gcaaattaga aaatagaata actagcataa 1860
acccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaacca atttgcctgg 1920
cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag 1980
cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa 2040
aacgaaaggc tcagtcgaaa gactgggcct ttcgggatcc aggccggcct gttaagacgg 2100
ccagtgaatt cgagctcggt acctaccgtt cgtataatgt atgctatacg aagttatcga 2160
gctctagaga atgatcccct cattaggcca cacgttcaag tgcagcgcac accgtggaaa 2220
cggatgaagg cacgaaccca gttgacataa gcctgttcgg ttcgtaaact gtaatgcaag 2280
tagcgtatgc gctcacgcaa ctggtccaga accttgaccg aacgcagcgg tggtaacggc 2340
gcagtggcgg ttttcatggc ttgttatgac tgtttttttg tacagtctat gcctcgggca 2400
tccaagcagc aagcgcgtta cgccgtgggt cgatgtttga tgttatggag cagcaacgat 2460
gttacgcagc agcaacgatg ttacgcagca gggcagtcgc cctaaaacaa agttaggtgg 2520
ctcaagtatg ggcatcattc gcacatgtag gctcggccct gaccaagtca aatccatgcg 2580
ggctgctctt gatcttttcg gtcgtgagtt cggagacgta gccacctact cccaacatca 2640
gccggactcc gattacctcg ggaacttgct ccgtagtaag acattcatcg cgcttgctgc 2700
cttcgaccaa gaagcggttg ttggcgctct cgcggcttac gttctgccca ggtttgagca 2760
gccgcgtagt gagatctata tctatgatct cgcagtctcc ggcgagcacc ggaggcaggg 2820
cattgccacc gcgctcatca atctcctcaa gcatgaggcc aacgcgcttg gtgcttatgt 2880
gatctacgtg caagcagatt acggtgacga tcccgcagtg gctctctata caaagttggg 2940
catacgggaa gaagtgatgc actttgatat cgacccaagt accgccacct aacaattcgt 3000
tcaagccgag atcgtagaat ttcgacgacc tgcagccaag cataacttcg tataatgtat 3060
gctatacgaa cggtaggatc ctctagagtc gacctgcagg catgagatgt gtataagaga 3120
cag 3123
<210> 98
<211> 2965
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 98
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgaaagaa 180
atcaaaatcc agaacatcat catcagcgaa gaaaaagcgc cgctggttgt gccggaaatc 240
ggcattaacc ataatggtag tctggaactg gcaaaaatca tggtggatgc ggcctttagc 300
gccggtgcaa aaatcattaa acatcagacc cacattgtgg aagatgaaat gtctaaagca 360
gcgaaaaaag ttatcccggg caacgcgaaa atcagtatct acgaaatcat gcagaaatgc 420
gcgctggatt acaaagatga actggccctg aaagaatata ccgaaaaact gggtctggtg 480
tacctgtcta ccccgtttag tcgtgcgggt gcaaaccgtc tggaagatat gggtgttagt 540
gcgttcaaaa tcggcagcgg tgaatgtaac aattatccgc tgatcaaaca tattgccgca 600
tttaaaaaac cgatgattgt tagcaccggc atgaatagca tcgaatctat taaaccgacg 660
gtgaaaatcc tgctggataa cgaaattccg tttgttctga tgcataccac gaatctgtac 720
ccgaccccgc acaacctggt gcgtctgaat gccatgctgg aactgaaaaa agaattctct 780
tgcatggttg gtctgagtga tcacaccacg gataatctgg catgcctggg tgcagtggtt 840
ctgggtgcgt gtgtgctgga acgtcatttc accgatagca tgcaccgctc tggtccggat 900
attgtttgta gtatggatac gaaagcactg aaagaactga tcattcagag cgaacagatg 960
gcgatcattc gcggcaacaa tgaatctaaa aaagcggcca aacaggaaca ggtgaccatc 1020
gattttgcat tcgcgagtgt ggttagcatc aaagatatca aaaaaggcga agtgctgagc 1080
atggataata tttgggttaa acgtccgggt ctgggcggta tctctgcagc ggaatttgaa 1140
aacattctgg gcaaaaaagc actgcgcgat attgaaaatg atgcgcagct gtcttatgaa 1200
gatttcgcct aataaatcga tactagcata accccttggg gcctctaaac gcgtcgacac 1260
gcaaaaaggc catccgtcag gatggccttc tgcttaattt gatgcctggc agtttatggc 1320
gggcgtcctg cccgccaccc tccgggccgt tgcttcgcaa cgttcaaatc cgctcccggc 1380
ggatttgtcc tactcaggag agcgttcacc gacaaacaac agataaaacg aaaggcccag 1440
tctttcgact gagcctttcg ttttatttga tgcctggcag ttccctactc tcgcatgggg 1500
agaccccaca ctaccatccg gtatcgataa gcttgatggc gaaaggggga tgtgctgcaa 1560
ggcgattaag ttgggtaacg ccagggtttt cccagtcacg acgttgtaaa acgacggcca 1620
gtgaattcga gctcggtacc taccgttcgt ataatgtatg ctatacgaag ttatcgagct 1680
ctagagaatg atcccctccc tcacgctgcc gcaagcactc agggcgcaag ggctgctaaa 1740
ggaagcggaa cacgtagaaa gccagtccgc agaaacggtg ctgaccccgg atgaatgtca 1800
gctactgggc tatctggaca agggaaaacg caagcgcaaa gagaaagcag gtagcttgca 1860
gtgggcttac atggcgatag ctagactggg cggttttatg gacagcaagc gaaccggaat 1920
tgccagctgg ggcgccctct ggtaaggttg ggaagccctg caaagtaaac tggatggctt 1980
tcttgccgcc aaggatctga tggcgcaggg gatcaagatc tgatcaagag acaggatgag 2040
gatcgtttcg catgattgaa caagatggat tgcacgcagg ttctccggcc gcttgggtgg 2100
agaggctatt cggctatgac tgggcacaac agacaatcgg ctgctctgat gccgccgtgt 2160
tccggctgtc agcgcagggg cgcccggttc tttttgtcaa gaccgacctg tccggtgccc 2220
tgaatgaact gcaggacgag gcagcgcggc tatcgtggct ggccacgacg ggcgttcctt 2280
gcgcagctgt gctcgacgtt gtcactgaag cgggaaggga ctggctgcta ttgggcgaag 2340
tgccggggca ggatctcctg tcatctcacc ttgctcctgc cgagaaagta tccatcatgg 2400
ctgatgcaat gcggcggctg catacgcttg atccggctac ctgcccattc gaccaccaag 2460
cgaaacatcg catcgagcga gcacgtactc ggatggaagc cggtcttgtc gatcaggatg 2520
atctggacga agagcatcag gggctcgcgc cagccgaact gttcgccagg ctcaaggcgc 2580
gcatgcccga cggcgaggat ctcgtcgtga cccatggcga tgcctgcttg ccgaatatca 2640
tggtggaaaa tggccgcttt tctggattca tcgactgtgg ccggctgggt gtggcggacc 2700
gctatcagga catagcgttg gctacccgtg atattgctga agagcttggc ggcgaatggg 2760
ctgaccgctt cctcgtgctt tacggtatcg ccgctcccga ttcgcagcgc atcgccttct 2820
atcgccttct tgacgagttc ttctgagcgg gactctggga atttcgacga cctgcagcca 2880
agcataactt cgtataatgt atgctatacg aacggtagga tcctctagag tcgacctgca 2940
ggcatgagat gtgtataaga gacag 2965
<210> 99
<211> 3904
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 99
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttaa ctttaagaag gagatataca aatgatcgct 180
caccgtcgtc aggaactggc tcaacagtat tatcaggctc tgcaccaaga tgtgctgccg 240
ttctgggaaa agtattcgct ggatcgtcaa ggcggtggct attttacctg cctggaccgc 300
aagggtcagg tttttgatac ggacaagttc atttggctgc aaaaccgtca agtgtggcaa 360
tttgcggttt tctacaatcg cctggaaccg aaaccgcagt ggctggaaat cgctcgtcat 420
ggtgcggatt ttctggcacg tcacggtcgt gatcaggacg gtaactggta tttcgccctg 480
gatcaggaag gcaaaccgct gcgccaaccg tacaatgtgt tttccgactg tttcgcggcg 540
atggcgttta gccagtatgc actggcttct ggtgctcaag aagcgaaggc cattgcactg 600
caagcgtata acaatgttct gcgtcgccag cataacccga aaggtcaata tgaaaagagt 660
tacccgggta cccgtccgct gaaatccctg gcagtgccga tgatcctggc taatctgacg 720
ctggaaatgg aatggctgct gccgccgacc acggtcgaag aagtgctggc ccagaccgtt 780
cgtgaagtca tgacggattt tctggacccg gaaattggcc tgatgcgcga agcagttacc 840
ccgacgggtg aatttgtcga ttcattcgaa ggccgcctgc tgaacccggg tcatggcatt 900
gaagcgatgt ggtttatgat ggatattgcc cagcgttcgg gtgaccgcca gctgcaagaa 960
caggctattg cggtggttct gaataccctg gaatatgcat gggatgaaga atttggtggc 1020
atcttttact tcctggaccg tcaaggtcac ccgccgcagc aactggaatg ggatcagaaa 1080
ctgtggtggg tccatctgga aaccctggtg gccctggcaa aaggtcacca ggcgacgggc 1140
caagaaaagt gctggcagtg gtttgaacgc gtgcatgatt atgcatggag ccactttgct 1200
gacccggaat atggtgaatg gttcggctac ctgaaccgtc gcggtgaagt gctgctgaat 1260
ctgaaaggtg gcaaatggaa gggctgcttc cacgttccgc gtgcgctgtg gctgtgtgcc 1320
gaaaccctgc aactgccggt ctcttaattt cgtcgacaca caggaaacat attaaaaatt 1380
aaaacctgca ggagtttaaa cgcggccgcg atatcgttgt aaaacgacgg ccagtgcaag 1440
aatcataaaa aatttatttg ctttcaggaa aatttttctg tataatagat tcataaattt 1500
gagagaggag tttttgtgag cggataacaa ttccccatct tagtatatta gttaagtata 1560
aatacacaag gagatataca tatgaaagaa atcaaaatcc agaacatcat catcagcgaa 1620
gaaaaagcgc cgctggttgt gccggaaatc ggcattaacc ataatggtag tctggaactg 1680
gcaaaaatca tggtggatgc ggcctttagc gccggtgcaa aaatcattaa acatcagacc 1740
cacattgtgg aagatgaaat gtctaaagca gcgaaaaaag ttatcccggg caacgcgaaa 1800
atcagtatct acgaaatcat gcagaaatgc gcgctggatt acaaagatga actggccctg 1860
aaagaatata ccgaaaaact gggtctggtg tacctgtcta ccccgtttag tcgtgcgggt 1920
gcaaaccgtc tggaagatat gggtgttagt gcgttcaaaa tcggcagcgg tgaatgtaac 1980
aattatccgc tgatcaaaca tattgccgca tttaaaaaac cgatgattgt tagcaccggc 2040
atgaatagca tcgaatctat taaaccgacg gtgaaaatcc tgctggataa cgaaattccg 2100
tttgttctga tgcataccac gaatctgtac ccgaccccgc acaacctggt gcgtctgaat 2160
gccatgctgg aactgaaaaa agaattctct tgcatggttg gtctgagtga tcacaccacg 2220
gataatctgg catgcctggg tgcagtggtt ctgggtgcgt gtgtgctgga acgtcatttc 2280
accgatagca tgcaccgctc tggtccggat attgtttgta gtatggatac gaaagcactg 2340
aaagaactga tcattcagag cgaacagatg gcgatcattc gcggcaacaa tgaatctaaa 2400
aaagcggcca aacaggaaca ggtgaccatc gattttgcat tcgcgagtgt ggttagcatc 2460
aaagatatca aaaaaggcga agtgctgagc atggataata tttgggttaa acgtccgggt 2520
ctgggcggta tctctgcagc ggaatttgaa aacattctgg gcaaaaaagc actgcgcgat 2580
attgaaaatg atgcgcagct gtcttatgaa gatttcgcct aaaataacta gcataacccc 2640
ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa accaatttgc ctggcggcag 2700
tagcgcggtg gtcccacctg accccatgcc gaactcagaa gtgaaacgcc gtagcgccga 2760
tggtagtgtg gggtctcccc atgcgagagt agggaactgc caggcatcaa ataaaacgaa 2820
aggctcagtc gaaagactgg gcctttcggg atccaggccg gcctgttaac gaattaatct 2880
tccgcggcgg tatcgataag cttgatatcg aattccgaag ttcctattct ctagaaagta 2940
taggaacttc aggtctgaag aggagtttac gtccagccaa gctagcttgg ctgcaggtcg 3000
tcgaaattct accgggtagg ggaggcgctt ttcccaaggc agtctggagc atgcgcttta 3060
gcagccccgc tgggcacttg gcgctacaca agtggcctct ggcctcgcac acattccaca 3120
tccaccggta ggcgccaacc ggctccgttc tttggtggcc ccttcgcgcc accttctact 3180
cctcccctag tcaggaagtt cccccccgcc ccgcagctcg cgtcgtgcag gacgtgacaa 3240
atggaagtag cacgtctcac tagtctcgtg cagatggaca gcaccgctga gcaatggaag 3300
cgggtaggcc tttggggcag cggccaatag cagctttgct ccttcgcttt ctgggctcag 3360
gggcgggctc agggggcggg gcgggcgccc gaaggtcctc cggaggcccg gcattctgca 3420
cgcttcaaaa gcgcacgtct gccgcgctgt tctcctcttc ctcatctccg ggcctttcga 3480
cctgcagcct gttgacaatt aatcatcggc atagtatatc ggcatagtat aatacgacaa 3540
ggtgaggaac taaaccatgg gtcaaagtag cgatgaagcc aacgctcccg ttgcagggca 3600
gtttgcgctt cccctgagtg ccacctttgg cttaggggat cgcgtacgca agaaatctgg 3660
tgccgcttgg cagggtcaag tcgtcggttg gtattgcaca aaactcactc ctgaaggcta 3720
tgcggtcgag tccgaatccc acccaggctc agtgcaaatt tatcctgtgg ctgcacttga 3780
acgtgtggcc taatgagggg atcaattctc tagagctcgc tgatcagaag ttcctattct 3840
ctagaaagta taggaacttc gatggcgcct catccctgaa gccaaagatg tgtataagag 3900
acag 3904
<210> 100
<211> 3793
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 100
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttgg cgtcgagaag gagatagaaa atgtgcggta 180
tcgttggtgc tatcgcacag cgtgatgtag cgaaaatcct cctggaaggt ctgcgtcgtc 240
tcgaataccg tggttacgac tctgccggtc tggcagtagt ggatgcagaa ggtcacatga 300
ctcgtctgcg tcgtctgggt aaagtgcaga tgctcgcgca ggcggcggaa gaacacccac 360
tccacggtgg tacgggtatc gcacacactc gttgggcaac ccacggtgaa ccgtctgagg 420
tcaacgcaca cccgcatgtt agcgagcaca tcgtagtcgt tcacaacggt atcatcgaga 480
accacgaacc actccgtgag gaactcaaag cccgtggtta caccttcgta agcgaaaccg 540
acacggaagt tatcgcccac ctcgttaact gggaactcaa acagggtggt actctgcgtg 600
aagcagttct gcgtgccatt ccacagctgc gtggtgcata cggtaccgtg atcatggact 660
ctcgtcatcc ggataccctg ctcgccgcac gttctggttc tccactcgtt atcggtctgg 720
gtatgggtga gaacttcatc gcctctgatc agctggccct gctcccagtt acccgtcgct 780
tcatcttcct ggaagagggt gacatcgccg aaatcacccg tcgttccgtt aacatcttcg 840
acaaaacggg tgcggaagtt aaacgtcagg acatcgagtc taacctgcag tatgacgctg 900
gtgacaaagg catctaccgt cactacatgc agaaagagat ctacgaacag ccgaacgcga 960
tcaaaaacac cctgaccggt cgtatctctc acggtcaggt tgacctgtct gagctgggtc 1020
caaacgcgga cgaactcctg tccaaagtcg agcacatcca gatcctggct tgtggtacct 1080
cttacaactc cggtatggtt tctcgttact ggttcgaatc tctggcaggt atcccatgcg 1140
acgttgaaat cgcctccgaa ttccgttatc gtaaatctgc ggtacgtcgt aactccctca 1200
tgatcaccct gtctcagtct ggtgaaaccg ctgatactct ggcaggtctg cgtctcagca 1260
aagaactggg ttacctgggt tctctggcca tctgcaacgt tccgggttct agcctggttc 1320
gtgagtctgt gctggctctg atgaccaacg cgggtacgga gatcggtgtt gcctctacca 1380
aagcgttcac tacccagctc actgtcctgc tgatgctggt tgccaaactg tctcgtctca 1440
aaggcctcga cgctagcatc gaacacgaca tcgtacacgg tctgcaggcc ctcccatctc 1500
gtatcgagca gatgctgccg caggacaaac gtatcgaagc actggcagaa gacttcagcg 1560
acaaacacca cgcgctgttt ctgggtcgtg gtgaccagta cccaattgcg ctggaaggtg 1620
ccctgaaact gaaagagatc agctacatcc atgcagaggc atacgcagcg ggtgagctga 1680
aacatggtcc actggccctg atcgacgcag atatgccggt tattgtggtt gctccgaaca 1740
acggcctgct ggagaaactg aaatccaaca tcgaggaagt acgtgcgcgt ggtggtcagc 1800
tgtacgtgtt tgctgaccag gacgcgggtt tcgtttccag cgacaacatg cacatcatcg 1860
aaatgccgca tgttgaagag gtaatcgcgc caatcttcta caccgtaccg ctgcagctgc 1920
tggcgtacca tgtagccctg atcaaaggta cggacgttga ccagccgcgt aacctggcga 1980
aatccgtgac cgtggaataa cgaaggagat agaaccatga gcttacccga tggattttat 2040
ataaggcgaa tggaagaggg ggatttggaa caggtcactg agacgctaaa ggttttgacc 2100
accgtgggca ctattacccc cgaatccttc agcaaactca taaaatactg gaatgaagcc 2160
acagtatgga atgataacga agataaaaaa ataatgcaat ataaccccat ggtgattgtg 2220
gacaagcgca ccgagacggt tgccgctacg gggaatatca tcatcgaaag aaagatcatt 2280
catgaactgg ggctatgtgg ccacatcgag gacattgcag taaactccaa gtatcagggc 2340
caaggtttgg gcaagctctt gattgatcaa ttggtaacta tcggctttga ctacggttgt 2400
tataagatta ttttagattg cgatgagaaa aatgtcaaat tctatgaaaa atgtgggttt 2460
agcaacgcag gcgtggaaat gcaaattaga aaatagcatc cgtatcggaa acactagcat 2520
aaccccttgg ggcctctaaa cgggtcttga ggggtttttt gctgaaacca atttgcctgg 2580
cggcagtagc gcggtggtcc cacctgaccc catgccgaac tcagaagtga aacgccgtag 2640
cgccgatggt agtgtggggt ctccccatgc gagagtaggg aactgccagg catcaaataa 2700
aacgaaaggc tcagtcgaaa gactgggcct ttcgcttcca caactttgta taataaagtt 2760
gtccccacgg ccagtgaatt cgagctcggt acctaccgtt cgtataatgt atgctatacg 2820
aagttatcga gctctagaga atgatcccct cattaggcca cacgttcaag tgcagcgcac 2880
accgtggaaa cggatgaagg cacgaaccca gttgacataa gcctgttcgg ttcgtaaact 2940
gtaatgcaag tagcgtatgc gctcacgcaa ctggtccaga accttgaccg aacgcagcgg 3000
tggtaacggc gcagtggcgg ttttcatggc ttgttatgac tgtttttttg tacagtctat 3060
gcctcgggca tccaagcagc aagcgcgtta cgccgtgggt cgatgtttga tgttatggag 3120
cagcaacgat gttacgcagc agcaacgatg ttacgcagca gggcagtcgc cctaaaacaa 3180
agttaggtgg ctcaagtatg ggcatcattc gcacatgtag gctcggccct gaccaagtca 3240
aatccatgcg ggctgctctt gatcttttcg gtcgtgagtt cggagacgta gccacctact 3300
cccaacatca gccggactcc gattacctcg ggaacttgct ccgtagtaag acattcatcg 3360
cgcttgctgc cttcgaccaa gaagcggttg ttggcgctct cgcggcttac gttctgccca 3420
ggtttgagca gccgcgtagt gagatctata tctatgatct cgcagtctcc ggcgagcacc 3480
ggaggcaggg cattgccacc gcgctcatca atctcctcaa gcatgaggcc aacgcgcttg 3540
gtgcttatgt gatctacgtg caagcagatt acggtgacga tcccgcagtg gctctctata 3600
caaagttggg catacgggaa gaagtgatgc actttgatat cgacccaagt accgccacct 3660
aacaattcgt tcaagccgag atcgtagaat ttcgacgacc tgcagccaag cataacttcg 3720
tataatgtat gctatacgaa cggtaggatc ctctagagtc gacctgcagg catgagatgt 3780
gtataagaga cag 3793
<210> 101
<211> 3847
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 101
ctgtctctta tacacatctc cggccagatg attaattcct aatttttgtt gacactctat 60
cattgataga gttattttac cactccctat cagtgataga gaaaagtgaa atgaatagtt 120
cgacaaaaat ctagaaataa ttttgtttgg cgtcgagaag gagatagaac catgtccaac 180
aatggctcgt caccgctggt gctttggtat aaccaactcg gcatgaatga tgtagacagg 240
gttgggggca aaaatgcctc cctgggtgaa atgattacta acctttccgg aatgggtgtt 300
tccgttccga atggtttcgc cacaaccgcc gacgcgttta accagtttct ggaccaaagc 360
ggcgtaaacc agcgcattta tgaactgctg gataaaacgg atattgacga tgttactcag 420
cttgcgaaag cgggcgcgca aatccgccag tggattatcg acactccctt ccagcctgag 480
ctggaaaacg ccatcagcga agcctatgca cagctttctg ccgatgacga aaacgcctct 540
tttgcggtgc gctcctccgc caccgcagaa gatatgccgg acgcttcttt tgccggtcag 600
caggaaacct tcctcaacgt tcagggtttt gacgccgttc tcgtggcagt gaaacatgta 660
tttgcttctc tgtttaacga tcgcgccatc tcttatcgtg tgcaccaggg ttacgatcac 720
cgtggtgtgg cgctctccgc cggtgttcaa cggatggtgc gctctgacct cgcatcatct 780
ggcgtgatgt tctccattga taccgaatcc ggctttgacc aggtggtgtt tatcacttcc 840
gcatggggcc ttggtgagat ggtcgtgcag ggtgcggtta acccggatga gttttacgtg 900
cataaaccga cactggcggc gaatcgcccg gctatcgtgc gccgcaccat ggggtcgaaa 960
aaaatccgca tggtttacgc gccgacccag gagcacggca agcaggttaa aatcgaagac 1020
gtaccgcagg aacagcgtga catcttctcg ctgaccaacg aagaagtgca ggaactggca 1080
aaacaggccg tacaaattga gaaacactac ggtcgcccga tggatattga gtgggcgaaa 1140
gatggccaca ccggtaaact gttcattgtg caggcgcgtc cggaaaccgt gcgctcacgc 1200
ggtcaggtca tggagcgtta tacgctgcat tcacagggta agattatcgc cgaaggccgt 1260
gctatcggtc atcgcatcgg tgcgggtccg gtgaaagtca tccatgatat cagcgaaatg 1320
aaccgcatcg aacctggtga cgtgctggtc actgacatga ccgacccgga ctgggaaccg 1380
atcatgaaga aagcatctgc catcgtcacc aaccgtggcg gtcgtacctg tcacgcggcg 1440
atcatcgctc gtgaactggg cattccggcg gtagtgggct gtggtgatgc aacagaacgg 1500
atgaaagacg gtgagaacgt cactgtttct tgtgccgaag gtgataccgg ttacgtctat 1560
gcggagttgc tggaatttag cgtgaaaagc tccagcgtag aaacgatgcc ggatctgccg 1620
ttgaaagtga tgatgaacgt cggtaacccg gaccgagctt tcgacttcgc ctgtctgccg 1680
aacgaaggcg tgggacttgc gcgtctggaa tttatcatca accgtatgat tggcgtccac 1740
ccacgcgcac tgcttgagtt tgacgatcag gaaccgcagt tgcaaaacga aatccgcgag 1800
atgatgaaag gttttgattc tccgcgtgaa ttttacgttg gtcgtctgac tgaagggatc 1860
gcgacgctgg gtgccgcgtt ttatccgaag cgcgtcattg tccgtctctc tgattttaaa 1920
tcgaacgaat atgccaacct ggtcggtggt gagcgttacg agccagatga agagaacccg 1980
atgctcggct tccgtggcgc gggacgctat atttccgaca gcttccgcga ctgtttcgcg 2040
ctggagtgcg aagcagtgaa acgtgtgcgc aacgacatgg ggctgaccaa cgttgagatc 2100
atgatcccgt tcgtgcgaac cgtagatcag gcgaaagcgg tggttgagga actggcgcgt 2160
caggggctga aacgtggtga gaacgggctg aaaatcatca tgatgtgtga aatcccgtcc 2220
aacgccttgc tggccgagca gttcctcgaa tatttcgacg gcttctcaat tggctcaaac 2280
gacatgacgc agctggcgct cggtctggat cgtgactccg gcgtggtgtc tgaactgttc 2340
gatgagcgca acgatgcggt gaaagcactg ctgtcgatgg cgattcgtgc cgcgaagaaa 2400
cagggcaaat atgtcgggat ttgcggtcag ggtccgtccg accacgaaga ctttgccgca 2460
tggttgatgg aagaggggat cgatagcctg tctctgaacc cggacaccgt ggtgcaaacc 2520
tggttaagcc tggctgaact gaagaaataa catccgtatc ggaaacacta gcataacccc 2580
ttggggcctc taaacgggtc ttgaggggtt ttttgctgaa accaatttgc ctggcggcag 2640
tagcgcggtg gtcccacctg accccatgcc gaactcagaa gtgaaacgcc gtagcgccga 2700
tggtagtgtg gggtctcccc atgcgagagt agggaactgc caggcatcaa ataaaacgaa 2760
aggctcagtc gaaagactgg gcctttcgct tccacaactt tgtataataa agttgtcccc 2820
acggccagtg aattcgagct cggtacctac cgttcgtata atgtatgcta tacgaagtta 2880
tcgagctcta gagaatgatc ccctcattag gccacacgtt caagtgcagc gcacaccgtg 2940
gaaacggatg aaggcacgaa cccagttgac ataagcctgt tcggttcgta aactgtaatg 3000
caagtagcgt atgcgctcac gcaactggtc cagaaccttg accgaacgca gcggtggtaa 3060
cggcgcagtg gcggttttca tggcttgtta tgactgtttt tttgtacagt ctatgcctcg 3120
ggcatccaag cagcaagcgc gttacgccgt gggtcgatgt ttgatgttat ggagcagcaa 3180
cgatgttacg cagcagcaac gatgttacgc agcagggcag tcgccctaaa acaaagttag 3240
gtggctcaag tatgggcatc attcgcacat gtaggctcgg ccctgaccaa gtcaaatcca 3300
tgcgggctgc tcttgatctt ttcggtcgtg agttcggaga cgtagccacc tactcccaac 3360
atcagccgga ctccgattac ctcgggaact tgctccgtag taagacattc atcgcgcttg 3420
ctgccttcga ccaagaagcg gttgttggcg ctctcgcggc ttacgttctg cccaggtttg 3480
agcagccgcg tagtgagatc tatatctatg atctcgcagt ctccggcgag caccggaggc 3540
agggcattgc caccgcgctc atcaatctcc tcaagcatga ggccaacgcg cttggtgctt 3600
atgtgatcta cgtgcaagca gattacggtg acgatcccgc agtggctctc tatacaaagt 3660
tgggcatacg ggaagaagtg atgcactttg atatcgaccc aagtaccgcc acctaacaat 3720
tcgttcaagc cgagatcgta gaatttcgac gacctgcagc caagcataac ttcgtataat 3780
gtatgctata cgaacggtag gatcctctag agtcgacctg caggcatgag atgtgtataa 3840
gagacag 3847
<210> 102
<211> 5554
<212> DNA
<213> Artificial Sequence
<220>
<223> plasmid
<400> 102
catcgattta ttatgacaac ttgacggcta catcattcac tttttcttca caaccggcac 60
ggaactcgct cgggctggcc ccggtgcatt ttttaaatac ccgcgagaaa tagagttgat 120
cgtcaaaacc aacattgcga ccgacggtgg cgataggcat ccgggtggtg ctcaaaagca 180
gcttcgcctg gctgatacgt tggtcctcgc gccagcttaa gacgctaatc cctaactgct 240
ggcggaaaag atgtgacaga cgcgacggcg acaagcaaac atgctgtgcg acgctggcga 300
tatcaaaatt gctgtctgcc aggtgatcgc tgatgtactg acaagcctcg cgtacccgat 360
tatccatcgg tggatggagc gactcgttaa tcgcttccat gcgccgcagt aacaattgct 420
caagcagatt tatcgccagc agctccgaat agcgcccttc cccttgcccg gcgttaatga 480
tttgcccaaa caggtcgctg aaatgcggct ggtgcgcttc atccgggcga aagaaccccg 540
tattggcaaa tattgacggc cagttaagcc attcatgcca gtaggcgcgc ggacgaaagt 600
aaacccactg gtgataccat tcgcgagcct ccggatgacg accgtagtga tgaatctctc 660
ctggcgggaa cagcaaaata tcacccggtc ggcaaacaaa ttctcgtccc tgatttttca 720
ccaccccctg accgcgaatg gtgagattga gaatataacc tttcattccc agcggtcggt 780
cgataaaaaa atcgagataa ccgttggcct caatcggcgt taaacccgcc accagatggg 840
cattaaacga gtatcccggc agcaggggat cattttgcgc ttcagccata cttttcatac 900
tcccgccatt cagagaagaa accaattgtc catattgcat cagacattgc cgtcactgcg 960
tcttttactg gctcttctcg ctaaccaaac cggtaacccc gcttattaaa agcattctgt 1020
aacaaagcgg gaccaaagcc atgacaaaaa cgcgtaacaa aagtgtctat aatcacggca 1080
gaaaagtcca cattgattat ttgcacggcg tcacactttg ctatgccata gcatttttat 1140
ccataagatt agcggatcct acctgacgct ttttatcgca actctctact gtttctccat 1200
acccgttttt ttgggaattc gagctctaag gaggttataa aaaatgtcta atctgctgac 1260
ggtccaccaa aacctgccgg ctctgccggt cgatgctacc tctgatgaag ttcgcaaaaa 1320
cctgatggat atgtttcgtg atcgccaggc attcagcgaa catacctgga aaatgctgct 1380
gtccgtgtgc cgttcatggg cggcctggtg taaactgaac aatcgcaaat ggtttccggc 1440
ggaaccggaa gatgtccgtg actatctgct gtacctgcag gcccgcggtc tggcagttaa 1500
aacgatccag caacatctgg gccaactgaa tatgctgcac cgtcgctccg gtctgccgcg 1560
tccgagcgat tctaatgcgg tgtcactggt tatgcgtcgc attcgtaaag aaaacgtgga 1620
tgcaggcgaa cgcgctaaac aggcactggc ttttgaacgt accgatttcg accaagttcg 1680
ctcgctgatg gaaaacagcg atcgttgcca ggacatccgc aatctggcgt tcctgggtat 1740
tgcctataac accctgctgc gcattgcaga aatcgctcgt attcgcgtga aagatatcag 1800
ccgtacggac ggcggtcgca tgctgattca catcggccgt accaaaacgc tggtctctac 1860
cgcaggcgtg gaaaaagctc tgagtctggg tgtgacgaaa ctggttgaac gctggattag 1920
tgtctccggc gtggcggatg acccgaacaa ttacctgttt tgtcgtgttc gcaaaaatgg 1980
tgtcgcagct ccgtcagcca cctcgcagct gagcacgcgt gcactggaag gcatcttcga 2040
agctacccat cgcctgattt atggcgccaa agatgactcg ggtcaacgtt acctggcgtg 2100
gtctggtcac agtgcacgtg ttggtgccgc acgtgatatg gcccgtgccg gtgtttccat 2160
cccggaaatt atgcaggcag gcggttggac caacgttaat atcgtcatga actatattcg 2220
caatctggac tcggaaacgg gtgctatggt tcgcctgctg gaagacggtg actaatgagt 2280
gccggagttc atcgaaaaaa tggacgaggc actggctgaa attggttttg tatttgggga 2340
gcaatggcga tgacgcatcc tcacgataat atccgggtag gcgcaatcac tttcgtctac 2400
tccgttacaa agcgaggctg ggtatttccc ggcctttctg ttatccgaaa tccactgaaa 2460
gcacagcggc tggctgagga gataaataat aaacgagggg ctgtatgcac aaagcatctt 2520
ctgttgagtt aagaacgagt atcgagatgg cacatagcct tgctcaaatt ggaatcaggt 2580
ttgtgccaat accagtagaa acagacgaag aatccatggg tatggacagt tttccctttg 2640
atatgtaacg gtgaacagtt gttctacttt tgtttgttag tcttgatgct tcactgatag 2700
atacaagagc cataagaacc tcagatcctt ccgtatttag ccagtatgtt ctctagtgtg 2760
gttcgttgtt tttgcgtgag ccatgagaac gaaccattga gatcatactt actttgcatg 2820
tcactcaaaa attttgcctc aaaactggtg agctgaattt ttgcagttaa agcatcgtgt 2880
agtgtttttc ttagtccgtt acgtaggtag gaatctgatg taatggttgt tggtattttg 2940
tcaccattca tttttatctg gttgttctca agttcggtta cgagatccat ttgtctatct 3000
agttcaactt ggaaaatcaa cgtatcagtc gggcggcctc gcttatcaac caccaatttc 3060
atattgctgt aagtgtttaa atctttactt attggtttca aaacccattg gttaagcctt 3120
ttaaactcat ggtagttatt ttcaagcatt aacatgaact taaattcatc aaggctaatc 3180
tctatatttg ccttgtgagt tttcttttgt gttagttctt ttaataacca ctcataaatc 3240
ctcatagagt atttgttttc aaaagactta acatgttcca gattatattt tatgaatttt 3300
tttaactgga aaagataagg caatatctct tcactaaaaa ctaattctaa tttttcgctt 3360
gagaacttgg catagtttgt ccactggaaa atctcaaagc ctttaaccaa aggattcctg 3420
atttccacag ttctcgtcat cagctctctg gttgctttag ctaatacacc ataagcattt 3480
tccctactga tgttcatcat ctgagcgtat tggttataag tgaacgatac cgtccgttct 3540
ttccttgtag ggttttcaat cgtggggttg agtagtgcca cacagcataa aattagcttg 3600
gtttcatgct ccgttaagtc atagcgacta atcgctagtt catttgcttt gaaaacaact 3660
aattcagaca tacatctcaa ttggtctagg tgattttaat cactatacca attgagatgg 3720
gctagtcaat gataattact agtccttttc ctttgagttg tgggtatctg taaattctgc 3780
tagacctttg ctggaaaact tgtaaattct gctagaccct ctgtaaattc cgctagacct 3840
ttgtgtgttt tttttgttta tattcaagtg gttataattt atagaataaa gaaagaataa 3900
aaaaagataa aaagaataga tcccagccct gtgtataact cactacttta gtcagttccg 3960
cagtattaca aaaggatgtc gcaaacgctg tttgctcctc tacaaaacag accttaaaac 4020
cctaaaggct taagtagcac cctcgcaagc tcggttgcgg ccgcaatcgg gcaaatcgct 4080
gaatattcct tttgtctccg accatcaggc acctgagtcg ctgtcttttt cgtgacattc 4140
agttcgctgc gctcacggct ctggcagtga atgggggtaa atggcactac aggcgccttt 4200
tatggattca tgcaaggaaa ctacccataa tacaagaaaa gcccgtcacg ggcttctcag 4260
ggcgttttat ggcgggtctg ctatgtggtg ctatctgact ttttgctgtt cagcagttcc 4320
tgccctctga ttttccagtc tgaccacttc ggattatccc gtgacaggtc attcagactg 4380
gctaatgcac ccagtaaggc agcggtatca tcaacggggt ctgacgctca gtggaacgaa 4440
aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa ggatcttcac ctagatcctt 4500
ttaaattaaa aatgaagttt taaatcaatc taaagtatat atgagtaaac ttggtctgac 4560
agttaccaat gcttaatcag tgaggcacct atctcagcga tctgtctatt tcgttcatcc 4620
atagttgcct gactccccgt cgtgtagata actacgatac gggagggctt accatctggc 4680
cccagtgctg caatgatacc gcgagaccca cgctcaccgg ctccagattt atcagcaata 4740
aaccagccag ccggaagggc cgagcgcaga agtggtcctg caactttatc cgcctccatc 4800
cagtctatta attgttgccg ggaagctaga gtaagtagtt cgccagttaa tagtttgcgc 4860
aacgttgttg ccattgctac aggcatcgtg gtgtcacgct cgtcgtttgg tatggcttca 4920
ttcagctccg gttcccaacg atcaaggcga gttacatgat cccccatgtt gtgcaaaaaa 4980
gcggttagct ccttcggtcc tccgatcgtt gtcagaagta agttggccgc agtgttatca 5040
ctcatggtta tggcagcact gcataattct cttactgtca tgccatccgt aagatgcttt 5100
tctgtgactg gtgagtactc aaccaagtca ttctgagaat agtgtatgcg gcgaccgagt 5160
tgctcttgcc cggcgtcaat acgggataat accgcgccac atagcagaac tttaaaagtg 5220
ctcatcattg gaaaacgttc ttcggggcga aaactctcaa ggatcttacc gctgttgaga 5280
tccagttcga tgtaacccac tcgtgcaccc aactgatctt cagcatcttt tactttcacc 5340
agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg caaaaaaggg aataagggcg 5400
acacggaaat gttgaatact catactcttc ctttttcaat attattgaag catttatcag 5460
ggttattgtc tcatgagcgg atacatattt gaatgtattt agaaaaataa acaaataggg 5520
gttccgcgca catttccccg aaaagtgcca cctg 5554
<210> 103
<211> 3415
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 103
ccggccagat gattaattcc taatttttgt tgacactcta tcattgatag agttatttta 60
ccactcccta tcagtgatag agaaaagtga aatgaatagt tcgacaaaaa tctagaaata 120
attttgttta actttaagaa ggagatatac aaatgaacaa cgacaactcc acgaccacca 180
acaataacgc tattgaaatc tatgtggatc gtgcgaccct gccgacgatc cagcaaatga 240
ccaaaattgt tagccagaaa acgtctaaca aaaaactgat ctcatggtcg cgctacccga 300
ttaccgataa aagcctgctg aagaaaatta acgcggaatt tttcaaagaa caatttgaac 360
tgacggaaag cctgaaaaac atcatcctgt ctgaaaacat cgataacctg atcattcatg 420
gcaataccct gtggagtatt gatgtggttg acattatcaa agaagtcaac ctgctgggca 480
aaaatattcc gatcgaactg cacttttatg atgacggttc cgccgaatac gttcgtatct 540
acgaatttag taaactgccg gaatccgaac agaaatacaa aaccagcctg tctaaaaaca 600
acatcaaatt ctcaatcgat ggcaccgact cgttcaaaaa cacgatcgaa aacatctacg 660
gtttcagcca actgtatccg accacgtacc acatgctgcg tgcagatatc ttcgacacca 720
cgctgaaaat taacccgctg cgcgaactgc tgtcaaacaa catcaaacag atgaaatggg 780
attacttcaa agacttcaac tacaaacaaa aagatatctt ttactcactg accaacttca 840
acccgaaaga aatccaggaa gacttcaaca aaaactcgaa caaaaacttc atcttcatcg 900
gcagtaactc cgcgaccgcc acggcagaag aacaaatcaa tattatcagc gaagcgaaga 960
aagaaaacag cagcattatc accaattcaa tttcggatta tgacctgttt ttcaaaggtc 1020
atccgtctgc cacgtttaac gaacagatta tcaatgcaca cgatatgatc gaaatcaaca 1080
acaaaatccc gttcgaagct ctgatcatga ccggcattct gccggatgcc gttggcggta 1140
tgggtagttc cgtctttttc agtatcccga aagaagtcaa aaacaaattc gtgttctata 1200
aaagtggtac ggatatcgaa aataactccc tgattcaggt gatgctgaaa ctgaatctga 1260
ttaaccgcga taatattaaa ctgatctctg acatttaatt tcgtcgacac acaggaaaca 1320
tattaaaaat taaaacctgc aggagtttaa acgcggccgc gatatcgttg taaaacgacg 1380
gccagtgcaa gaatcataaa aaatttattt gctttcagga aaatttttct gtataataga 1440
ttcataaatt tgagagagga gtttttgtga gcggataaca attccccatc ttagtatatt 1500
agttaagtat aaatacacaa ggagatatac atatgagcct ggccattatc ccggcacgtg 1560
gcggttctaa aggcatcaaa aacaaaaacc tggttctgct gaacaataaa ccgctgattt 1620
attacaccat caaagcggcc ctgaacgcca aaagtattag caaagtggtt gtgagctctg 1680
attctgatga aatcctgaac tacgcaaaaa gtcagaacgt tgatatcctg aaacgtccga 1740
tcagtctggc acaggatgat accacgagcg ataaagtgct gctgcatgcg ctgaaattct 1800
acaaagatta cgaagatgtt gtgttcctgc agccgaccag cccgctgcgt acgaatattc 1860
acatcaacga agcgttcaac ctgtacaaaa acagcaacgc aaacgcgctg atttctgtta 1920
gtgaatgcga taacaaaatc ctgaaagcgt ttgtgtgcaa tgattgtggc gatctggccg 1980
gtatttgtaa cgatgaatac ccgttcatgc cgcgccagaa actgccgaaa acctatatga 2040
gcaatggtgc catctacatc ctgaaaatca aagaattcct gaacaacccg agcttcctgc 2100
agtctaaaac gaaacatttc ctgatggatg aaagtagctc tctggatatt gattgcctgg 2160
aagatctgaa aaaagtggaa cagatctgga aaaaataaaa tactgaaacc aatttgcctg 2220
gcggcagtag cgcggtggtc ccacctgacc ccatgccgaa ctcagaagtg aaacgccgta 2280
gcgccgatgg tagtgtgggg tctccccatg cgagagtagg gaactgccag gcatcaaata 2340
aaacgaaagg ctcagtcgaa agactgggcc tttcgcttcc acaactttgt ataataaagt 2400
tgtccccacg gccagtgaat tcgagctcgg tacctaccgt tcgtataatg tatgctatac 2460
gaagttatcg agctctagag aatgatcccc tcattaggcc acacgttcaa gtgcagcgca 2520
caccgtggaa acggatgaag gcacgaaccc agttgacata agcctgttcg gttcgtaaac 2580
tgtaatgcaa gtagcgtatg cgctcacgca actggtccag aaccttgacc gaacgcagcg 2640
gtggtaacgg cgcagtggcg gttttcatgg cttgttatga ctgttttttt gtacagtcta 2700
tgcctcgggc atccaagcag caagcgcgtt acgccgtggg tcgatgtttg atgttatgga 2760
gcagcaacga tgttacgcag cagcaacgat gttacgcagc agggcagtcg ccctaaaaca 2820
aagttaggtg gctcaagtat gggcatcatt cgcacatgta ggctcggccc tgaccaagtc 2880
aaatccatgc gggctgctct tgatcttttc ggtcgtgagt tcggagacgt agccacctac 2940
tcccaacatc agccggactc cgattacctc gggaacttgc tccgtagtaa gacattcatc 3000
gcgcttgctg ccttcgacca agaagcggtt gttggcgctc tcgcggctta cgttctgccc 3060
aggtttgagc agccgcgtag tgagatctat atctatgatc tcgcagtctc cggcgagcac 3120
cggaggcagg gcattgccac cgcgctcatc aatctcctca agcatgaggc caacgcgctt 3180
ggtgcttatg tgatctacgt gcaagcagat tacggtgacg atcccgcagt ggctctctat 3240
acaaagttgg gcatacggga agaagtgatg cactttgata tcgacccaag taccgccacc 3300
taacaattcg ttcaagccga gatcgtagaa tttcgacgac ctgcagccaa gcataacttc 3360
gtataatgta tgctatacga acggtaggat cctctagagt cgacctgcag gcatg 3415
<210> 104
<211> 3763
<212> DNA
<213> Artificial Sequence
<220>
<223> Expression cassette
<400> 104
ccggccagat gattaattcc taatttttgt tgacactcta tcattgatag agttatttta 60
ccactcccta tcagtgatag agaaaagtga aatgaatagt tcgacaaaaa tctagaaata 120
attttgttta actttaagaa ggagatatac aaatgtgtaa cgataatcaa aatacggtcg 180
atgttgttgt gagcaccgtt aacgataacg tcatcgaaaa caacacgtac caagttaaac 240
cgatcgatac cccgaccacg tttgacagtt actcctggat tcagacgtgc ggcaccccga 300
tcctgaaaga tgacgaaaaa tattcactgt cgtttgattt cgtcgccccg gaactggatc 360
aggacgaaaa attctgtttc gaatttaccg gcgatgttga cggtaaacgt tatgtcacgc 420
agaccaacct gacggtggtt gcaccgaccc tggaagttta cgtcgatcat gctagtctgc 480
cgtccctgca gcaactgatg aaaatcatcc agcagaaaaa cgaatactca cagaatgaac 540
gtttcatttc gtggggccgc atcggtctga cggaagataa cgcggaaaaa ctgaatgccc 600
atatttatcc gctggcaggc aacaatacct cacaggaact ggtggatgca gtgatcgatt 660
acgctgactc gaaaaaccgt ctgaatctgg aactgaacac gaataccgcg cacagctttc 720
cgaacctggc cccgattctg cgcattatca gctctaaaag caacatcctg atctctaaca 780
tcaacctgta cgatgacggc agtgctgaat atgtgaacct gtacaattgg aaagataccg 840
aagacaaatc cgtgaaactg agcgattctt tcctggttct gaaagactac tttaacggta 900
ttagttccga aaaaccgagc ggcatctatg gtcgctacaa ctggcatcaa ctgtataata 960
cgtcttatta cttcctgcgt aaagattacc tgaccgttga accgcagctg cacgacctgc 1020
gcgaatatct gggcggtagt ctgaaacaaa tgtcctggga tggcttttca cagctgtcga 1080
aaggtgacaa agaactgttc ctgaacattg tcggctttga tcaggaaaaa ctgcagcaag 1140
aataccagca atcagaactg ccgaatttcg tgtttacggg caccacgacc tgggcaggcg 1200
gtgaaaccaa agaatattac gctcagcaac aggtgaacgt cgtgaacaat gcgattaatg 1260
aaaccagccc gtattacctg ggccgtgaac atgacctgtt tttcaaaggt cacccgcgcg 1320
gcggtattat caatgatatt atcctgggca gtttcaacaa tatgattgac atcccggcca 1380
aagtgtcctt tgaagttctg atgatgacgg gtatgctgcc ggataccgtg ggcggtattg 1440
cgtcatcgct gtattttagc atcccggccg aaaaagtctc tttcattgtg tttaccagct 1500
ctgatacgat caccgatcgt gaagacgcgc tgaaatctcc gctggtgcag gttatgatga 1560
ccctgggcat tgttaaagaa aaagatgtgc tgttctggtc ggatctgccg gattgttcct 1620
cgggtgtttg tattgctcag tattaatttc gtcgacacac aggaaacata ttaaaaatta 1680
aaacctgcag gagtttaaac gcggccgcga tatcgttgta aaacgacggc cagtgcaaga 1740
atcataaaaa atttatttgc tttcaggaaa atttttctgt ataatagatt cataaatttg 1800
agagaggagt ttttgtgagc ggataacaat tccccatctt agtatattag ttaagtataa 1860
atacacaagg agatatacat atgagcctgg ccattatccc ggcacgtggc ggttctaaag 1920
gcatcaaaaa caaaaacctg gttctgctga acaataaacc gctgatttat tacaccatca 1980
aagcggccct gaacgccaaa agtattagca aagtggttgt gagctctgat tctgatgaaa 2040
tcctgaacta cgcaaaaagt cagaacgttg atatcctgaa acgtccgatc agtctggcac 2100
aggatgatac cacgagcgat aaagtgctgc tgcatgcgct gaaattctac aaagattacg 2160
aagatgttgt gttcctgcag ccgaccagcc cgctgcgtac gaatattcac atcaacgaag 2220
cgttcaacct gtacaaaaac agcaacgcaa acgcgctgat ttctgttagt gaatgcgata 2280
acaaaatcct gaaagcgttt gtgtgcaatg attgtggcga tctggccggt atttgtaacg 2340
atgaataccc gttcatgccg cgccagaaac tgccgaaaac ctatatgagc aatggtgcca 2400
tctacatcct gaaaatcaaa gaattcctga acaacccgag cttcctgcag tctaaaacga 2460
aacatttcct gatggatgaa agtagctctc tggatattga ttgcctggaa gatctgaaaa 2520
aagtggaaca gatctggaaa aaataaaata ctgaaaccaa tttgcctggc ggcagtagcg 2580
cggtggtccc acctgacccc atgccgaact cagaagtgaa acgccgtagc gccgatggta 2640
gtgtggggtc tccccatgcg agagtaggga actgccaggc atcaaataaa acgaaaggct 2700
cagtcgaaag actgggcctt tcgcttccac aactttgtat aataaagttg tccccacggc 2760
cagtgaattc gagctcggta cctaccgttc gtataatgta tgctatacga agttatcgag 2820
ctctagagaa tgatcccctc attaggccac acgttcaagt gcagcgcaca ccgtggaaac 2880
ggatgaaggc acgaacccag ttgacataag cctgttcggt tcgtaaactg taatgcaagt 2940
agcgtatgcg ctcacgcaac tggtccagaa ccttgaccga acgcagcggt ggtaacggcg 3000
cagtggcggt tttcatggct tgttatgact gtttttttgt acagtctatg cctcgggcat 3060
ccaagcagca agcgcgttac gccgtgggtc gatgtttgat gttatggagc agcaacgatg 3120
ttacgcagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa gttaggtggc 3180
tcaagtatgg gcatcattcg cacatgtagg ctcggccctg accaagtcaa atccatgcgg 3240
gctgctcttg atcttttcgg tcgtgagttc ggagacgtag ccacctactc ccaacatcag 3300
ccggactccg attacctcgg gaacttgctc cgtagtaaga cattcatcgc gcttgctgcc 3360
ttcgaccaag aagcggttgt tggcgctctc gcggcttacg ttctgcccag gtttgagcag 3420
ccgcgtagtg agatctatat ctatgatctc gcagtctccg gcgagcaccg gaggcagggc 3480
attgccaccg cgctcatcaa tctcctcaag catgaggcca acgcgcttgg tgcttatgtg 3540
atctacgtgc aagcagatta cggtgacgat cccgcagtgg ctctctatac aaagttgggc 3600
atacgggaag aagtgatgca ctttgatatc gacccaagta ccgccaccta acaattcgtt 3660
caagccgaga tcgtagaatt tcgacgacct gcagccaagc ataacttcgt ataatgtatg 3720
ctatacgaac ggtaggatcc tctagagtcg acctgcaggc atg 3763
Claims (15)
- 적어도 하나의 N-아세틸뉴라민산 모이어티(moiety)를 포함하는 사카라이드(saccharide)의 발효 생산 방법으로서, 상기 방법이
a) (i) 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함하는 시알산 생합성 경로(sialic acid biosynthesis pathway);
(ii) 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제; 및
(iii) 이종유래(heterologous) 시알릴트랜스퍼라제
를 포함하는 적어도 하나의 유전자 조작된 미생물 세포를 제공하는 단계;
b) 적어도 하나의 유전자 조작된 미생물 세포를 발효 브로쓰(fermentation broth)에서 그리고 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 상기 사카라이드의 생산을 위해 허용되는 조건 하에 배양하는 단계; 및 임의로
c) 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 상기 사카라이드를 회수하는 단계를 포함하는, 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 사카라이드의 발효 생산 방법. - 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 사카라이드의 발효 생산을 위한 유전자 조작된 미생물 세포로서, 여기서 유전자 조작된 미생물 세포가
(i) 글루코사민-6-포스페이트 N-아세틸트랜스퍼라제를 포함하는 합성 시알산 생합성 경로;
(ii) 시티딘 5'-모노포스포-(CMP)-N-아세틸뉴라민산 신테타제; 및
(iii) 이종유래 시알릴트랜스퍼라제
를 포함하는, 유전자 조작된 미생물 세포. - 제1항 또는 제2항에 있어서, 시알산 생합성 경로가 a) N-아세틸글루코사민-6-포스페이트 포스파타제 및 N-아세틸글루코사민 2-에피머라제; 및/또는 b) N-아세틸글루코사민-6-포스페이트 에피머라제 및 N-아세틸만노사민-6-포스페이트 포스파타제를 추가로 포함하는 것인, 제1항에 따른 방법 또는 제2항에 따른 유전자 조작된 미생물 세포.
- 제1항 내지 제3항 중 어느 한 항에 있어서,
유전자 조작된 미생물 세포가,
i) 서열번호: 91로 표시된 바와 같은 폴리펩티드를 코딩하는(encoding) 뉴클레오티드 서열;
ii) 서열번호: 92로 표시된 바와 같은 뉴클레오티드 서열;
iii) 서열번호: 91로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열과 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 유사성(sequence similarity)을 갖는 뉴클레오티드 서열;
iv) 서열번호: 92로 표시된 바와 같은 뉴클레오티드 서열 중 하나와 적어도 80%, 90%, 95%, 96%, 97%, 98%, 99% 또는 99% 초과의 서열 유사성을 갖는 뉴클레오티드 서열;
v) i., ii., iii. 및 iv.의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi) i., ii., iii. 및 iv 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하는 것인, 제1항 내지 제3항 중 어느 한 항에 따른 방법 또는 제2항 또는 제3항에 따른 유전자 조작된 미생물 세포. - 제1항 내지 제4항 중 어느 한 항에 있어서, 유전자 조작된 미생물 세포가,
I. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드;
II. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 아미노산 서열 중 어느 하나와 적어도 80%의 서열 유사성을 갖는 아미노산 서열을 포함하거나 이로 이루어진 폴리펩티드; 및
III. I. 및 II.의 폴리펩티드 중 어느 하나의 단편
으로 이루어진 군으로부터 바람직하게 선택된 이종유래 시알릴트랜스퍼라제를 함유하는 것인, 제1항 내지 제4항 중 어느 한 항에 따른 방법 또는 제2항 내지 제4항 중 어느 한 항에 따른 유전자 조작된 미생물 세포. - 제1항 내지 제5항 중 어느 한 항에 있어서, 유전자 조작된 미생물 세포가,
i. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열;
ii. 서열번호: 34 내지 66 중 어느 하나로 표시된 바와 같은 뉴클레오티드 서열;
iii. 서열번호: 1 내지 33 중 어느 하나로 표시된 바와 같은 폴리펩티드를 코딩하는 뉴클레오티드 서열 중 하나와 뉴클레오티드 서열 중 하나와 적어도 80%의 서열 유사성을 갖는 뉴클레오티드 서열;
iv. 서열번호: 34 내지 66으로 표시된 뉴클레오티드 서열 중 어느 하나와 적어도 80%의 서열 유사성을 갖는 뉴클레오티드 서열;
v. i., ii., iii. 및 iv의 뉴클레오티드 서열 중 어느 하나에 상보적인 뉴클레오티드 서열; 및
vi. i., ii., iii., iv. 및 v.의 뉴클레오티드 서열 중 어느 하나의 단편
으로 이루어진 군으로부터 선택된 뉴클레오티드 서열을 포함하고 발현하는 핵산 분자를 함유하는 것인, 제1항 내지 제5항 중 어느 한 항에 따른 방법 또는 제2항 내지 제5항 중 어느 한 항에 따른 유전자 조작된 미생물 세포. - 수용체 분자(acceptor molecule)가 N-아세틸글루코사민, 갈락토스, N-아세틸갈락토사민, 락토스, 락툴로스, N-아세틸락토사민, 락토-N-비오스, 락툴로스, 멜리비오스, 라피노스, 락토-N-트리오스 II, 2'-푸코실락토스, 3-푸코실락토스, 3'-시알릴락토스, 6'-시알릴락토스, 3'-시알릴-N-아세틸락토사민, 6'-시알릴-N-아세틸락토사민, 3'-갈락토실락토스, 6'-갈락토실락토스, 락토-N-테트라오스, 락토-N-네오테트라오스, 2'3-디푸코실락토스, 3-푸코실-3'-시알릴락토스, 3-푸코실-6'-시알릴락토스, 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c, 락토-N-푸코펜타오스 I, 락토-N-푸코펜타오스 II, 락토-N-푸코펜타오스 III, 락토-N-푸코펜타오스 V, 락토-N-네오푸코펜타오스 I 및 락토-N-네오푸코펜타오스 V로 이루어진 군으로부터 선택된 것인, 제1항 내지 제6항 중 어느 한 항에 따른 방법 또는 제2항 내지 제6항 중 어느 한 항에 따른 유전자 조작된 미생물 세포.
- 제1항 내지 제7항 중 어느 한 항에 있어서, 발효 브로쓰가 적어도 하나의 탄소 공급원을 함유하며, 적어도 하나의 탄소 공급원이 바람직하게는 글루코스, 프럭토스, 수크로스, 글리세롤 및 그의 조합으로 이루어진 군으로부터 선택된 것인 방법.
- 제1항 내지 제8항 중 어느 한 항에 있어서, 적어도 하나의 유전자 조작된 미생물 세포가 글루코사민, N-아세틸글루코사민, N-아세틸만노사민 및 N-아세틸뉴라민산으로 이루어진 군으로부터 선택된 하나 이상의 부재 하에 및/또는 그의 첨가 없이 배양된 것인 방법.
- 제1항 내지 제9항 중 어느 한 항에 있어서, 적어도 하나의 N-아세틸뉴라민산 모이어티를 포함하는 사카라이드가 3'-시알릴갈락토스, 6'-시알릴갈락토스, 3'-시알릴-N-아세틸락토사민, 6'-시알릴-N-아세틸락토사민, 3'-시알릴락토스, 6'-시알릴락토스, 시알릴락토-N-테트라오스 a, 시알릴락토-N-테트라오스 b, 시알릴락토-N-테트라오스 c, 푸코실-시알릴락토-N-테트라오스 a, 푸코실-시알릴락토-N-테트라오스 b, 푸코실-시알릴락토-N-테트라오스 c, 디시알릴락토-N-테트라오스, 푸코실디시알릴락토-N-테트라오스 I, 푸코실디시알릴락토-N-테트라오스 II로 이루어진 군으로부터 선택된 것인 방법.
- 전체 세포 발효 공정에서 시알릴화 사카라이드의 생산을 위한, 제2항 내지 제7항 중 어느 한 항에 따른 유전자 조작된 미생물 세포의 용도.
- 제1항 내지 제10항 중 어느 한 항에 따른 방법에 의해 또는 제2항 내지 제7항 중 어느 한 항에 따른 유전자 조작된 미생물 세포의 사용에 의해 생산된 시알릴화 사카라이드.
- 영양 조성물의 제조를 위한, 제12항에 따른 시알릴화 사카라이드의 용도
- 제1항 내지 제9항 중 어느 한 항에 따른 방법에 의해 또는 제2항 내지 제7항 중 어느 한 항에 따른 유전자 조작된 미생물 세포에 의해 생산된, 적어도 하나의 시알릴화 사카라이드, 바람직하게는 적어도 하나의 시알릴화 올리고사카라이드를 함유하는 영양 조성물.
- 제14항에 있어서, 의약, 제약, 제제, 유아용 유동식(infant formula) 및 식이 보충제(dietary supplement)로 이루어진 군으로부터 선택된 것인 영양 조성물.
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PCT/EP2019/063669 WO2019228993A1 (en) | 2018-05-28 | 2019-05-27 | Fermentative production of sialylated saccharides |
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EP3848471A1 (en) * | 2020-01-10 | 2021-07-14 | Chr. Hansen HMO GmbH | Sequential fermentative production of oligosaccharides |
CN111411065B (zh) * | 2020-03-30 | 2022-07-05 | 江南大学 | 一种基于人工双碳源的产n-乙酰神经氨酸的重组菌 |
CN116323930A (zh) | 2020-08-10 | 2023-06-23 | 因比奥斯公司 | 唾液酸化双糖及/或寡糖的细胞生产 |
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JP2021525522A (ja) | 2021-09-27 |
US20210198709A1 (en) | 2021-07-01 |
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AU2019278599A1 (en) | 2020-12-17 |
AU2019278599B2 (en) | 2023-11-09 |
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PH12020552047A1 (en) | 2021-06-28 |
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