KR970065709A - Process for the preparation of acarbiosyl transferases and methods of use for the conversion of acarbose homologues to acarbose and for the preparation of acarbose homologues - Google Patents

Process for the preparation of acarbiosyl transferases and methods of use for the conversion of acarbose homologues to acarbose and for the preparation of acarbose homologues Download PDF

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KR970065709A
KR970065709A KR1019970009945A KR19970009945A KR970065709A KR 970065709 A KR970065709 A KR 970065709A KR 1019970009945 A KR1019970009945 A KR 1019970009945A KR 19970009945 A KR19970009945 A KR 19970009945A KR 970065709 A KR970065709 A KR 970065709A
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acarbiosyl
acarbose
transferase
maltose
preparation
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KR1019970009945A
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안넬리제 크루에거
한스-게오르그 델베크
위르겐 게오르그 렌즈
베르너 쉬뢰더
헤르만 파페
클라우스 고에케
베아테 샤퍼
미카엘 헴커
볼프강 피에퍼스베르크
위르겐 디스틀러
안스가르 스트라트만
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볼프강 페트로빅키, 만프레트 조벨
바이엘 악티엔게젤샤프트
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Publication of KR970065709A publication Critical patent/KR970065709A/en

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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/10—Transferases (2.)
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    • C—CHEMISTRY; METALLURGY
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    • C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
    • C12N9/10—Transferases (2.)
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
    • C12P19/00—Preparation of compounds containing saccharide radicals
    • C12P19/26—Preparation of nitrogen-containing carbohydrates
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    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
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    • C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
    • C12N15/09—Recombinant DNA-technology
    • C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression

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Abstract

본 발명은 방선균류(actinomycetes), 주로 악티노플레인스종(Actinoplanessp.) SE 50/13 또는 SE 50/110 및 그의 변이종으로부터의 아카르비오실 트랜스퍼라제(AT), 이 효소의 단리, 정제 및 특성화 방법, 아카르비오실 트랜스퍼라제를 코딩하는 acbD유전자의 단리 및 특성화, 이종 숙주 생물체 내에서의 아카르비오실 트랜스퍼라제의 발현, 아카르보스 부수 성분의 아카르보스로의 전환 및 아카르보스 상동체의 제조를 위한 아카르비오실 트랜스퍼라제의 용도, 아카르비오실 트랜스퍼라제의 아카르보스 정제에서의 용도 및 아카르비오실 트랜스퍼라제의 불활성화에 의해 부수 성분의 형성이 감소되는 생산 변이체의 제조에 관한 것이다.The present invention relates to acarbiosyl transferases (ATs) from actinomycetes, mainly Actinoplanes sp. SE 50/13 or SE 50/110 and variants thereof, isolation, purification and characterization of this enzyme. Methods, isolation and characterization of acbD gene encoding acarbiosyl transferase, expression of acarbiosyl transferase in heterologous host organisms, conversion of acarbose minor components to acarbose and acarbose phase Use of acarbiosyl transferase for the preparation of fuselage, use in acarbose tablets of acarbiosyl transferase, and production variants in which formation of minor components is reduced by inactivation of acarbiosyl transferase. It is about manufacture.

본 발명의 아카르보스의 말토실 잔기를 치환하는 능력을 갖는 아카르비오실 트랜스퍼라제를 사용하여 발효 동안에 형성되는 아카르보스 유사 부수 성분의 당잔기를 말토스 잔기로 치환함으로써 아카르보스를 효과적으로 형성시킬 수 있다.Acarbose is effectively substituted by the maltose moiety of the acarbose like minor component formed during fermentation using an acarbioosyl transferase having the ability to substitute the maltosyl residue of the acarbose of the present invention. Can be formed.

Description

아카르비오실 트랜스퍼라제의 제조 방법, 및 아카르보스 상동체의 아카르보스로의 전환 및 아카르보스 상동체의 제조를 위한 그의 사용 방법Process for the preparation of acarbiosyl transferases and methods of use for the conversion of acarbose homologues to acarbose and for the preparation of acarbose homologues

본 내용은 요부공개 건이므로 전문내용을 수록하지 않았음As this is a public information case, the full text was not included.

제1도는 DNA 서열 분석을 위한 pAS5 및 pAS5/15.1의 플라스미드 유도체를 나타내는 도면, 제2도는 발현 벡터 pUWL201(6.40kb)를 나타내는 도면, 제3도는 아카르비오실 트랜스퍼라제의 아미노산 서열을 나타내는 도면, 제4도는 아카르비오실 트랜스퍼라제를 코딩하는 DNA의 서열을 나타내는 도면.1 shows a plasmid derivative of pAS5 and pAS5 / 15.1 for DNA sequence analysis, FIG. 2 shows the expression vector pUWL201 (6.40 kb), FIG. 3 shows the amino acid sequence of acarbiosyl transferase, 4 shows the sequence of DNA encoding acarbiosyl transferase.

Claims (10)

아카르비오실 트랜스퍼라제.Acarbiosyl transferase. 악티노플레인스 종 SE 50, SE 50/13 또는 SE 50/110 및 이들의 변이종에서 유래한 아카르비오실 트랜스퍼라제.Acarbiosyl transferases derived from actinoplanes species SE 50, SE 50/13 or SE 50/110 and variants thereof. 필수적으로 순수한 아카르비오실 트랜스퍼라제.Essentially pure acarbiosyl transferase. 제3도의 아미노산 서열을 갖는 아카르비오실 트랜스퍼라제 및 그의 기능성 유도체.Acarbiosyl transferase and functional derivative thereof having the amino acid sequence of FIG. 아카르비오실 트랜스퍼라제를 코딩하는 제4도의 DNA 서열 및 그의 기능성 유도체.DNA sequence of FIG. 4 encoding acarbiosyl transferase and functional derivatives thereof. 아카르비오실 트랜스퍼라제에 의해 촉매되는 하기 반응식에 의한 아카르보스 부수 성분의 아카르보스로의 전환 방법.A method for converting an acarbose minor component into acarbose according to the following scheme catalyzed by an acarbiosyl transferase. 아카르비오실-Gn+말토스→아카르보스+Gn Acarbiosyl-G n + maltose → acarbose + G n (상기 식에서 Gn은 글루코스, 디사카라이드 및 말토올리고사카라이드를 의미한다)(Wherein G n means glucose, disaccharide and maltooligosaccharide) 다른 슈도올리고사카라이드, 예를 들면 트레스타틴 또는 아밀로스타틴 또는 다른 글루코시다제 억제제의 아카르보스로의 전환에 있어서의 아카르비오실 트랜스퍼라제의 용도.Use of acarbiosyl transferase in the conversion of other pseudooligosaccharides such as trestatin or amylostatin or other glucosidase inhibitors to acarbose. 배양 상징액으로부터 아카르보스를 농축하고 이와 동시에 아카르비오실 트랜스퍼라제의 존재하에 아카르보스 상동체를 고분자량 덱스트린 또는 녹말과 반응시키고 저분자량 불순물을 투석 또는 역삼투에 의해 제거하거나,Concentrate the acarbose from the culture supernatant and at the same time react the acarbose homolog with high molecular weight dextrin or starch in the presence of acarbiosyl transferase and remove low molecular weight impurities by dialysis or reverse osmosis, 아카르비오실-(G)n+(G)m→ 아카르비오실+(G)m+(G)n Acarbiosyl- (G) n + (G) m → acarbiosyl + (G) m + (G) n (상기 식에서, G는 글루코스를 의미하고 m 및 n은 1 내지 20의 정수이고 각각의 경우에 m 및 n은 상이하다).(Wherein G means glucose and m and n are integers from 1 to 20 and in each case m and n are different). 또는 폴리사카라이드를 침전시킨 후 적합하게 고농도의 말토스의 존재하에 아카르비오실 트랜스퍼라제를 사용하여 말토스와 반응시켜 아카르보스를 방출하여Or after precipitating the polysaccharide, reacting with maltose using acarbiosyl transferase in the presence of a high concentration of maltose to release acarbose, 아카르비오실-Gm+말토스→아카르보스+Gm Acarbiosyl-G m + Maltose → Acarbos + G m 아카르보스 상동체를 아카르보스로 전환하기 위한 아카르비오실 트랜스퍼라제의 용도.Use of acarbiosyl transferase to convert acarbose homologues to acarbose. 이종 숙주 생물체 내의 아카르비오실 트랜스퍼라제의 재조합 제제.Recombinant preparation of acarbiosyl transferase in a heterologous host organism. 제5항에 따른 DNA를 함유하는 벡터.A vector containing the DNA according to claim 5. ※ 참고사항 : 최초출원 내용에 의하여 공개하는 것임.※ Note: The disclosure is based on the initial application.
KR1019970009945A 1996-03-22 1997-03-22 Process for the preparation of acarbiosyl transferases and methods of use for the conversion of acarbose homologues to acarbose and for the preparation of acarbose homologues KR970065709A (en)

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
DE19611252 1996-03-22
DE19611252.4 1996-03-22
DE19625269.5 1996-06-25
DE19625269A DE19625269A1 (en) 1996-03-22 1996-06-25 Acarviosyltransferase from Actinoplanes sp. SE 50/110 and mutants, processes for the preparation and use of acarviosyltransferase in the conversion of acarbose homologues into acarbose, for the production of acarbose homologues and for strain optimization

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AR (1) AR006364A1 (en)
DE (1) DE19625269A1 (en)
TW (1) TW363080B (en)
ZA (1) ZA972424B (en)

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ZA972424B (en) 1997-09-25
AR006364A1 (en) 1999-08-25
TW363080B (en) 1999-07-01

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