KR102579533B1 - 아세틸콜린 수용체 저해 펩타이드 및 이의 용도 - Google Patents
아세틸콜린 수용체 저해 펩타이드 및 이의 용도 Download PDFInfo
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- KR102579533B1 KR102579533B1 KR1020220150342A KR20220150342A KR102579533B1 KR 102579533 B1 KR102579533 B1 KR 102579533B1 KR 1020220150342 A KR1020220150342 A KR 1020220150342A KR 20220150342 A KR20220150342 A KR 20220150342A KR 102579533 B1 KR102579533 B1 KR 102579533B1
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- KR
- South Korea
- Prior art keywords
- peptide
- acetylcholine receptor
- espep
- acid
- binding
- Prior art date
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Abstract
본 발명은 아세틸콜린 수용체 저해 펩타이드 및 이의 용도에 관한 것으로, 랜덤 펩타이드 재조합 파지를 이용하여 아세틸콜린 수용체와 결합력이 높은 파지를 스크리닝하고, 파지의 DNA를 통해 아세틸콜린 수용체 결합 펩타이드를 선별하였다. 선별된 펩타이드를 이용하여 아세틸콜린 수용체와의 결합력 및 아세틸콜린 수용체 작용 억제 효과를 확인하였고, 펩타이드의 변형을 통해 아세틸콜린 수용체와의 결합에 중요한 펩타이드 부위 및 서열을 확인함으로써, 본 발명의 아세틸콜린 수용체 저해 펩타이드를 이용하여 주름 개선용 화장료 조성물, 아세틸콜린 수용체 관련 질환의 예방 또는 치료를 위한 의약품 및 개선용 건강기능식품을 개발할 수 있을 것으로 기대된다.
Description
본 발명은 아세틸콜린 수용체 저해 펩타이드 및 이의 용도에 관한 것이다.
아세틸콜린(acetylcholine)은 동물의 신경조직에 존재하는 화학물질로 신경의 말단에서 분비되며, 신경의 자극을 근육에 전달하는 역할을 한다. 신경말단으로부터 분비되는 전달물질로는 운동신경과 부교감신경에서는 아세틸콜린이, 교감신경에서는 에피네프린(아드레날린)이 알려져 있다. 아세틸콜린이 분비되면 혈압강하, 심장박동억제, 장관수축, 골격근 수축 등의 생리작용을 나타낸다. 근육이 수축하려면 신경이 근육에 수축하라는 명령을 내리고 이에 따라 근육이 수축을 하게 되는데, 그 명령은 신경과 근육이 서로 만나는 부분(신경-근 접합부)에서 신경이 아세틸콜린을 분비하고 이 물질이 근육의 아세틸콜린 수용체에 접합하여 근육은 수축을 할 수 있게 된다(Vincent, A., 1985; Lindstrom, J.M., et al., 1976). 대퇴부 골격근을 지배하는 말초의 아세틸콜린 수용체를 차단하면 근육운동의 마비가 일어나며, 호흡 또는 심장 운동을 담당하는 평활근 및 심장근의 아세틸콜린 수용체를 차단하면 호흡, 심장 운동 마비가 일어난다.
아세틸콜린 수용체는 무스카린성 아세틸콜린 수용체(muscarinic acetylcholine receptor, mAchR)와 니코틴성 아세틸콜린 수용체(nicotinic acetylcholine receptor, nAchR)로 분류된다. 무스카린성 아세틸콜린 수용체는 무스카린(muscarine)으로 활성화시킬 수 있는 G 단백결합 수용체(G protein-coupled receptor)이며 아형(subtype)에 따라 서로 다른 신호전달기전을 활성화시킨다. 무스카린성 아세틸콜린 수용체는 중추신경계(central nervous system)와 말초 장기(peripheral organ)를 포함하여 온 몸에 분포하고 있으며, 주로 부교감신경계(parasympathetic nervous system)의 신경절 이후 섬유에서 분비되는 아세틸콜린의 생리적 작용을 매개하는 역할을 한다.
니코틴성 아세틸콜린 수용체는 니코틴(nicotine)에 의한 약리작용을 모방하는 수용체로, 신경전달물질에 의해서 조작되는 이온 채널(ion channel)이다. 이온 채널의 개방 및 폐쇄를 통해서 나트륨, 칼륨, 칼슘 이온 등을 비선택적으로 통과시키는 비선택적 양이온 통로로서, 신경세포와 근육세포 간의 전기적 신호전달(electronic signaling)을 조절한다. 니코틴성 아세틸콜린 수용체는 발현 부위에 따라 근육형(muscle type)과 신경형(neuronal type)으로 나눌 수 있다. 근육형 니코틴성 아세틸콜린 수용체는 운동신경(motor neuron)과 골격근(skeletal muscle)이 만나는 신경근육접합부(neuromuscular junction)에 발현되어 있으며, 운동 신경에서 분비된 아세틸콜린이 골격근 세포막의 종판전압(end plate potential, EPP)을 유발하는데 기여한다. 한편, 신경형 니코틴성 아세틸콜린 수용체는 자율신경계(autonomic nervous system, ANS)의 말초 신경절(ganglia)에 발현되어 있어 신경절 이전 섬유(preganglionic fiber)에서 분비된 아세틸콜린이 신경절 이후 섬유(postganglionic fiber)를 흥분시키는데 기여한다.
아세틸콜린의 활성을 방해 혹은 저해하거나 행동을 모방하는 약물이 매우 유용하게 사용되고 있다. 아세틸콜린 수용체 작용제는 중증 근무력증과 알츠하이머 병을 치료하는데 이용된다. 중증 근무력증의 경우, 몸에서 니코틴성 아세틸콜린 수용체에 대한 항체를 생산하여 정상적인 아세틸콜린 신호 전달을 저해하여 생기는 자가면역질환으로, 아세틸콜린에스터레이스(acetylcholine esterase, AChE) 저해제를 이용하여 신경과 근육 사이의 시냅스 간극에서 아세틸콜린이 불활성화되기 전에 각 수용체와 상호작용할 수 있는 시간을 늘려주어 중증 근무력증을 치료할 수 있다.
또한, 아세틸콜린의 분비를 방해하면 근육의 수축을 억제하여 근육이 마비가 되면서 주름살이 펴지는데, 이를 이용한 것이 보톡스(botox)이다. 보톡스는 운동신경 말단에서 근육의 수축에 필수적인 물질인 아세틸콜린이 분비되는 과정을 막는다. 그 결과 근육이 움직일 수 없게 되고, 근육에 의해 유발되던 주름이 없어지게 된다. 보톡스에 의한 근-이완 효과는 3~6주 후에 점차 사라지기 때문에 반복투여를 필요로 한다.
이에 본 발명자들은, 아세틸콜린 수용체 결합 펩타이드를 연구하는 과정에서, 아세틸콜린 수용체와 결합력이 높은 펩타이드를 스크리닝하여 확보하였고, 이러한 펩타이드가 아세틸콜린 수용체와 결합하여 아세틸콜린 결합을 방지함으로써 아세틸콜린 수용체 작용을 억제한다는 것을 확인함으로써 본 발명을 완성할 수 있었다.
또한 상기 스크리닝된 펩타이드가 아세틸콜린 수용체와 결합하여 그 작용을 억제하는 데에 있어서 중요한 특정 아미노산의 일정한 배치순서를 확인하고, 일정한 식으로 표현되는 상기 펩타이드의 라이브러리가 전체적으로 아세틸콜린 수용체와 결합하여 그 작용을 억제하는 것을 확인하고 본 발명을 완성하였다.
종래선행기술인 한국등록특허 제1216008호에는 바이오패닝을 이용하여 선별한, 아세틸콜린 수용체에 결합하는 펩타이드가 기재되어 있으나, 본 발명의 아미노산 서열을 포함하는 펩타이드 및 라이브러리는 기재되어 있지 않다. 또한, 한국공개특허 제2018-0028748호에는 아세틸콜린을 포함하는 신경전달물질 방출 조절 펩타이드 및 이의 주름 개선 효과가 기재되어 있으나, 본 발명의 아미노산 서열을 포함하는 펩타이드의 아세틸콜린 수용체 결합력 및 이로 인한 아세틸콜린 수용체 작용 억제 효과는 기재되어 있지 않다. 한국공개특허 제2014-0139010호에는 경피 투과 촉진을 위한 펩타이드가 기재되어 있으나, 본 발명의 아세틸콜린 수용체 결합을 통한 작용 억제 펩타이드와는 그 구성이 차이가 있다.
Lindstrom, J.M., et al., Antibody to acetylcholine receptor in myasthenia gravis: Prevalence, chlincal correlates, and diagnostic value, Neurology, 26, 1054-1059, 1976.
Vincent, A., et al., Acetylcholine receptor antibody as a diagnosis test for myasthenia gravis: results in 153 validated cases and 2967 diagnostic assays, J. Neurol. Neurolsurg. Psychiatry, 48, 1246-1252, 1985.
본 발명의 목적은 아세틸콜린 수용체 저해 펩타이드 및 이를 포함하는 조성물을 제공하는 데 있다.
본 발명은 하기 화학식 1로 표현되는 아미노산 서열을 포함하는 아세틸콜린 수용체 결합 펩타이드에 관한 것이다.
[화학식 1]
X
L
-(K 또는 R)-X-(K 또는 R)-X
M
-(K 또는 R)-X
N
상기 XL, XM, XN은 각각 독립적으로 1~8개, X는 1개의 임의의 아미노산으로 이루어진 서열일 수 있으며, 바람직하게는 XL , XM, XN은 각각 독립적으로 1~4개, X는 1개의 임의의 아미노산일 수 있다.
본 발명은 하기 화학식 2로 표현되는 아미노산 서열을 포함하는 아세틸콜린 수용체 결합 펩타이드에 관한 것이다.
[화학식 2]
(K 또는 R)-X-(K 또는 R)-(K 또는 R)-X
M-1
-(K 또는 R)
상기 XM-1은 1~3개, X는 1개의 임의의 아미노산으로 이루어진 서열일 수 있으며, 바람직하게는 XM-1은 3개, X는 1개의 임의의 아미노산일 수 있다.
상기 펩타이드는 서열번호 1 내지 200 (표 10 참조)으로 이루어진 군에서 선택되는 중 어느 하나의 서열에 의한 펩타이드일 수 있다.
본 발명은 하기 화학식 3 또는 3-1 로 표현되는 아미노산 서열을 포함하는 아세틸콜린 수용체 결합 펩타이드에 관한 것이다.
[화학식 3]
X
L
-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-X
M-1
-(K 또는 R)
상기 XL, XM-1은 각각 독립적으로 1~3개, X는 1개의 임의의 아미노산으로 이루어진 서열일 수 있으며, 바람직하게는 상기 XL, XM-1은 각각 독립적으로 3개, X는 1개의 임의의 아미노산일 수 있다.
[화학식 3-1]
(K 또는 R)-X-(K 또는 R)-(K 또는 R)-
X
M
-1
-(K 또는 R)-
X
N
상기 XM -1, XN은 각각 독립적으로 1~3개, X는 1개의 임의의 아미노산으로 이루어진 서열일 수 있으며, 바람직하게는 상기 XM -1, XN은 각각 독립적으로 3개, X는 1개의 임의의 아미노산일 수 있다.
상기 펩타이드는 서열번호 201 내지 400 (표 11 참조)으로 이루어진 군에서 선택되는 중 어느 하나의 서열에 의한 펩타이드일 수 있다.
본 발명은 하기 화학식 4로 표현되는 아미노산 서열을 포함하는 아세틸콜린 수용체 결합 펩타이드에 관한 것이다.
[화학식 4]
X
L
-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-X
M-1
-(K 또는 R)-X
N
상기 XL, XM-1, XN 은 각각 독립적으로 1~3개, X는 1개의 임의의 아미노산일 수 있으며, 바람직하게는 상기 XL, XM-1, XN은 각각 독립적으로 3개, X는 1개의 임의의 아미노산일 수 있다.
상기 펩타이드는 서열번호 401 내지 600 (표 12 참조)으로 이루어진 군에서 선택되는 중 어느 하나의 서열에 의한 펩타이드인 것을 특징으로 하는 아세틸콜린 수용체 결합 펩타이드일 수 있다.
상기 펩타이드는 8~28개의 아미노산 서열로 이루어진 것일 수 있다. 바람직하게는 8~16개이며, 더 바람직하게는 8~14개이며, 더욱 바람직하게는 11~16개이며, 가장 바람직하게는 14개이다.
상기 아세틸콜린 수용체 결합 펩타이드는 상기 화학식 1로 표현되는 아미노산 서열을 중심으로 하여, 1, 3 및 8번째 아미노산인 K 또는 R 이 아세틸콜린 수용체 결합에 있어서 중요한 부위일 수 있다.
또한 상기 아세틸콜린 수용체 결합 펩타이드는 상기 화학식 2로 표현되는 아미노산 서열을 중심으로 하여, 1, 3, 4 및 8번째 아미노산인 K 또는 R 이 아세틸콜린 수용체 결합에 있어서 중요한 부위일 수 있다.
상기 아세틸콜린 수용체 결합 펩타이드의 아미노산 서열은 본 발명의 선행특허인 대한민국 등록특허 제10-1971092호에 기재된 아미노산 서열을 제외한다. 상기 대한민국 등록특허 제10-1971092호에 기재된 아미노산 서열은 WTWKGRKSLLR을 포함한다.
상기 아세틸콜린 수용체 결합 펩타이드의 아미노산 서열은 대한민국 공개특허 10-2014-0139010호에 기재된 아미노산 서열을 제외한다. 상기 대한민국 공개특허 10-2014-0139010호에 기재된 아미노산 서열은 (gly)n1-(arg)n2, (gly)p-RGRDDRRQRRR-(gly)q, (gly)p-YGRKKRRQRRR-(gly)q, (gly)p-RKKRRQRRR-(gly)q 상기 아래첨자 p 및 q는 각각 독립적으로 0 내지 20의 정수이고, 상기 아래첨자 n1은 독립적으로 1 내지 8의 정수이고, 상기 아래첨자 n2는 독립적으로 7 내지 17의 홀수인 서열 펩타이드이다.
본 발명은 상기 펩타이드의 N-말단 또는 C-말단이 변형(modification)된 것을 특징으로 하는 아세틸콜린 수용체 결합 펩타이드에 관한 것이다.
상기 N-말단 또는 C-말단이 변형(modification)된 것은 팔미토일화(palmitoylation), 아세틸화(acetylation), 아미드화(amidation), 포르밀화(formylation) 또는 페길화(PEGylation)되거나, 또는 2-메르캅토아세트산(2-mercaptoacetic acid), 3-메르캅토프로피오닉산(3-mercaptopropionic acid), 6-메르캅토헥사노익산(6-mercaptohexanoic acid), 피로글루탐산(pyroglutamic acid), 숙신이미드산(succinimide acid), 시스트라민(cystramine), 시스테아민(cysteamine), 메틸에스테르(methyl ester), 에틸에스테르(ethyl ester), 벤질에스테르(benzyl ester) 및 지방산, 예를 들면, 미리스트산(myristic acid), 스테아르산(stearic acid), 팔미트산(palmitic acid), 콜레스테롤(cholesterole), 6-아미노 헥사노익산(6-amino hexanoic acid) 및 8-아미노 옥타노익산(8-amino octanoic acid)로 이루어진 군으로부터 선택된 1 이상의 결합에 의한 것을 특징으로 하는 아세틸콜린 수용체 결합 펩타이드일 수 있다.
본 발명은 아세틸콜린 수용체 결합 펩타이드에 관한 것이다.
본 발명에 있어서, "펩타이드(peptide)"란 아미드 결합 또는 펩타이드 결합으로 연결된 2개 이상의 아미노산으로 이루어진 폴리머를 의미한다.
본 발명에 있어서, "아미노산(amino acid)"이란 펩타이드 분야에서 합성 펩타이드를 제조하는데 사용되는 천연 아미노산 및 기타 아미노산, 예컨대, 천연 아미노산, 비천연 아미노산, 염기서열로 암호화되지 않은 아미노산 등의 L- 및 D- 이성질체를 모두 포함한다.
상기 천연 아미노산은 알라닌(alanine, Ala, A), 시스테인(cystenin, Cys, C), 아스파트산(Aspartic acid, Asp, D), 글루탐산(glutamic acid, Glu, E), 페닐알라닌(phenylalanine, Phe, F), 글리신(glycine, Gly, G), 히스티딘(histidine, His, H), 이소류신(isoleucine, Ile, I), 리신(lysine, Lys, K), 류신(leucine, Leu, L), 메티오닌(methionine, Met, M), 아스파라긴(asparagine, Asn, N), 프롤린(proline, Pro, P), 글루타민(glutamine, Gln, Q), 아르기닌(arginine, Arg, R), 세린(serine, Ser, S), 트레오닌(threonine, Thr, T), 발린(Valine, Val, V), 트립토판(tryptophan, Trp, W), 및 티로신(tyrosine, Tyr, Y)일 수 있다.
상기 기타 아미노산은 2-아미노아디픽산(2-aminoadipic acid, 2-aminohexanedioic acid), α-아스파라긴(α-asparagine), 2-아미노부타노익산(2-aminobutanoic acid), 2-아미노카프릭산(2-aminocapric acid, 2-aminodecanoic acid), α-글루타민(α-glutamine), α-아미노이소부티릭산(α-aminoisobutyric acid, α-methylalanine), 2-아미노피멜릭산(2-aminopimelic acid, 2-aminohepanedioic acid), γ-아미노-β-히드록시벤젠펜타노익산(γ-amino-β-hydroxybenzenepentanoic acid), 2-아미노수베릭산(2-aminosuberic acid, 2-aminooctanedioic acid), 2-카르복시아제티딘(2-carboxyazetidine), β-알라닌(β-alanine), β-아스파트산(β-aspartic acid), 3,6-디아미노헥사노익산(3,6-diaminohexanoic acid, β-lysine), 부타노익산(butanoic acid), 4-아미노-3-히드록시부타노익산(4-amino-3-hydroxybutanoic acid), γ-아미노-β-히드록시시클로헥산펜타노익산(γ-amino-β-hydroxycyclohexanepentanoic acid), 3-시클로헥실알라닌(3-cyclohexylalanine), N5-아미노카르보닐오르니틴(N5-aminocarbonylornithine), 3-설포알라닌(3-sulfoalanine), 2,4-디아미노프로파노익산(2,3-diaminopropanoic acid), 2,7-디아미노수베릭산(2,7-diaminosuberic acid, 2,7-diaminooctanedioic acid), S-에틸티오시스테인(S-ethylthiocysteine), γ-글루탐산(γ-glutamic acid), γ-카르복시글루탐산(γ-carboxylglutamic acid), 히드록시아세트산(hydroxyacetic acid, glycolic acid), 피로글루탐산(pyroglutamic acid), 호모아르기닌(homogrginine), 호모시스테인(homocysteine), 호모히스티딘(homohistidine), 2-히드록시이소발레릭산(2-hydroxyisovaleric acid), 호모세린(homoserine), 2-히드록시펜타노익산(2-hydroxypentanoic acid), 5-히드록시리신(5-hhydroxylysine), 4-히드록시프롤린(4-hydroxyproline), 이소발린(isovaline), 2-히드록시프로파노익산(2-hydroxypropanoic acid, lactic acid), 메르캅토아세트산(mercaptoacetic acid), 메르캅토부타노익산(mercaptobutanoic acid), 3-히드록시-4-메틸프로롤린(3-hydroxy-4-methylproline), 메르캅토프로파노익산(mercaptopropanoic acid), 3-나프틸알라닌(3-naphthylalanine), 노르류신(norleucine), 노르티로신(nortyrosine), 노르발린(norvaline), 2-카르복시옥타히드로인돌(2-carboxyoctahydroindole), 오르니틴(ornithine), 페니실라민(penicillamine, 3-mercaptovaline), 2-페닐글리신(2-phenylglycine), 2-카르복시피페리딘(2-carboxypiperidine), 사르코신(sarcosine, N-methylglycine), 1-아미노-1-카르복시시클로펜탄(1-amino-1-carboxycyclopentane), 스타틴(statin, 4-amino-3-hydroxy-6-methylheptanoic acid), 3-티에닐알라린(3-thienylalanine), 3-카르복시이소퀴놀린(3-carboxyisoquinoline), 3-메틸발린(3-methylvaline), ε-N-트리메틸리신(ε-N-trimethyllysine), 3-티아졸릴알라닌(3-thiazolylalanine), α-아미노-2,4-디옥소피리미딘프로파노익산(α-amino-2,4-dioxopyrimidinepropanoic acid) 등일 수 있다.
본 발명의 펩타이드는 당 분야에 널리 공지된 방법을 통해 획득할 수 있다. 구체적으로는 유전자 재조합과 단백질 발현 시스템을 이용하여 제조하거나 펩타이드 합성과 같은 화학적 합성 방법을 통하여 시험관 내에서 합성하는 방법 및 무세포 단백질 합성법 등으로 제조될 수 있다. 보다 구체적으로는 당 분야에서 잘 알려진 방법, 예를 들면, 자동 펩타이드 합성기로 합성할 수 있으며, 유전자 조작 기술로 생산할 수 있으나, 이에 제한되는 것은 아니다. 예를 들어, 유전자 조작을 통하여 융합 파트너와 본 발명의 펩타이드로 이루어진 융합 단백질을 암호화 하는 유전자를 제조하고, 제조된 유전자를 숙주 미생물에 형질전환시킨 후, 숙주 미생물에서 융합 단백질 형태로 발현 시키고, 단백질 분해 효소 또는 화합물을 이용하여 융합 단백질로부터 본 발명의 펩타이드를 절단, 분리하여 원하는 펩타이드를 생산할 수 있다.
본 발명의 펩타이드는 염 형태로 존재할 수도 있다. 본 발명에 사용 가능한 염의 형태는 화합물의 최종 분리 및 정제 동안 또는 아미노기를 적절한 산과 반응시키는 것에 의해 만들어지는 것일 수 있다. 예를 들면, 산 부가염으로 아세테이트, 아디페이트, 알기네이트, 시트레이트, 아스파테이트, 벤조에이트, 벤젠설포네이트, 바이설페이트, 부티레이트, 캄포레이트, 캄포설포네이트, 디글루코네이트, 글리세로포스페이트, 헤미설페이트, 헵타노에이트, 헥사노에이트, 포르메이트, 푸마레이트, 하이드로 클로라이드, 하이드로브로마이드, 하이드로요오다이드, 2-하이드록시에탄설포네이트, 락테이트, 말레에이트, 메시틸렌설포네이트, 메탄설포네이트, 나프틸렌설포네이트, 니코티네이트, 2-나프탈렌설포네이트, 옥살레이트, 파모에이트, 펙티네이트, 퍼설페이트, 3-페닐프로피오네이트, 피크레이트, 피발레이트, 프로피오네이트, 숙시네이트, 타르트레이트, 트리클로로아테이트, 트리플루오로아세테이트, 포스페이트, 글루타메이트, 바이카보네이트, 파라-톨루엔설포네이트 및 운데카노에이트일 수 있으나, 이에 한정되는 것은 아니다. 또한, 산 부가염을 형성하기 위해 사용될 수 있는 산의 예로는 염산, 브롬화수소산, 황산 및 인산과 같은 무기산 및 옥살산, 말레산, 숙신산 및 시트르산과 같은 유기산일 수 있으나, 이에 한정하는 것은 아니다.
상기 펩타이드는 표적화 서열, 태그(tag), 표지된 잔기, 반감기 또는 펩타이드의 안정성을 증가시키기 위한 목적으로 고안된 아미노산 서열 추가, 표적화, 효능 증가 또는 안정성을 증가시키기 위한 항체 또는 항체절편, 인간혈청알부민(HSA) 등과의 결합 또는 펩타이드의 N-말단 또는 C-말단을 변형(modification)시킬 수 있다.
상기 "항체"는 항원성 부위에 대해서 지시되는 특이적인 단백질 분자를 의미한다. 바람직하게는 상기 항체는 특정 단백질 또는 이의 면역원성 단편에 대해 특이적으로 결합하는 항체를 의미하며, 단클론 항체(monoclonal antibody, mAb), 다클론 항체(polyclonal antibody, pAb) 및 재조합 항체를 모두 포함할 수 있다. 상기 항체를 생성하는 것은 당업계에 널리 공지된 기술을 이용하여 용이하게 제조할 수 있다.
상기 항체는 2개의 전체 길이 경쇄(light chain) 및 2개의 전체 길이 중쇄(heavy chain)를 가지는 완전한 형태뿐만 아니라, 항체 분자의 기능적인 단편을 포함할 수 있다. 항체 분자의 기능적 단편이란 적어도 항원 결합기능을 보유하고 있는 단편을 뜻하며, Fab, F(ab'), F(ab')2, F(ab)2, Fv 등이 있다.
상기 펩타이드는 특정 조직으로 전달 또는 안정성 확보를 위해 나노입자, 마이크로입자, 금속입자, 세라믹입자, 하이드로겔 등에 내포 또는 고정화할 수 있으나, 이에 한정되는 것은 아니다.
상기 나노입자, 마이크로입자, 금속입자, 세라믹입자, 하이드로겔 등은 생체적합적이며, 독성이 없는 것일 수 있다.
본 발명에 있어서, "아세틸콜린 수용체(acetylcholine receptor, AchR)"는 신경의 말단에서 분비되는 아세틸콜린이 결합하는 수용체로써, 아세틸콜린에 의한 신경의 자극을 전달하는 통로 역할을 한다. 예를 들면, 근육의 수축이 필요한 경우, 신경이 근육에 수축하라는 명령을 내리면 신경과 근육이 만나는 부분에서 신경이 아세틸콜린을 분비하고, 분비된 아세틸콜린이 근육의 아세틸콜린 수용체에 결합하면 근육이 수축된다.
본 발명의 아세틸콜린 수용체는 무스카린성 아세틸콜린 수용체와 니코틴성 아세틸콜린 수용체로 분류되며, 본 발명의 아세틸콜린 수용체는 니코틴성 아세틸콜린 수용체가 바람직하다.
상기 아세틸콜린 수용체 결합 펩타이드는 아세틸콜린 수용체에 결합하여 아세틸콜린이 수용체에 결합하는 것을 방지하여 아세틸콜린 수용체 작용을 억제할 수 있다. 바람직하게는 근육의 수축을 억제하여 주름개선, 비정상적인 근육 수축 억제 효과를 나타내며, 수술시 근육의 이완을 촉진하여 수술의 편의성을 확보할 수 있다.
또한, 본 발명은 상기 아세틸콜린 수용체 결합 펩타이드를 암호화 하는 폴리뉴클레오티드(polynucleotide)를 제공한다. 상기 아세틸콜린 수용체에 대하여 결합 활성을 나타낼 수 있는 펩타이드를 암호화 할 수 있는 한, 상기 폴리뉴클레오티드를 이루고 있는 염기 서열과 상동성을 나타내는 염기서열을 포함하는 폴리뉴클레오티드 역시 본 발명에서 제공하는 폴리뉴클레오티드의 범주에 포함될 수 있다. 바람직하게는 80% 이상의 상동성을 나타내는 염기서열을 포함하는 폴리뉴클레오티드이고, 더 바람직하게는 90% 이상의 상동성을 나타내는 염기서열을 포함하는 폴리뉴클레오티드이고, 가장 바람직하게는 95% 이상의 상동성을 나타내는 염기서열을 포함하는 폴리뉴클레오티드이다.
본 발명은 또한, 상기 아세틸콜린 수용체 결합 펩타이드를 포함하는 주름 개선용 화장료 조성물을 제공한다.
상기 아세틸콜린 수용체 결합 펩타이드는 아세틸콜린 수용체 작용을 억제하여 근육의 수축이 일어나는 것을 방지하여 주름을 개선할 수 있다.
상기 화장료 조성물은 상기 아세틸콜린 수용체 결합 펩타이드 및 화장 분야에서 통상적으로 사용되는 보조제, 예컨대 친수성 또는 친유성 겔화제, 친수성 또는 친유성 활성제, 보존제, 항산화제, 용매, 방향제, 충전제, 차단제, 안료, 흡취제 또는 염료를 함유할 수 있다.
상기 보조제의 양은 당해 분야에서 통상적으로 사용되는 양이며, 어떠한 경우라도 보조제 및 그 비율은 본 발명에 따른 화장료 조성물의 바람직한 성질에 악영향을 미치지 않도록 선택될 수 있다.
상기 주름 개선용 화장료 조성물은 추가 첨가제를 포함하여 제조할 수 있다.
상기 첨가제는 보습제(moisturizer), 기능성 원료, 점증제(thickener), 유연제, 유화제(emulsifier), 계면활성제 및 pH 조절제 등일 수 있다.
상기 보습제로는 글리세린, 프로필렌글리콜, 부틸렌글리콜, 히알루론산(hyaluronic acid) 및 세라마이드(ceramide) 성분 등일 수 있으나, 이에 한정하는 것은 아니다.
상기 점증제로는 폴리머, 잔탄검, 구아검일 수 있으나, 이에 한정하는 것은 아니다.
상기 유연제로는 미네랄 오일, 쉐아버터(shea butter), 파라핀일 수 있으나, 이에 한정하는 것은 아니다.
상기 유화제로는 디메치콘, 비즈왁스 등 일 수 있다.
상기 주름 개선용 화장료 조성물은 주름 개선 효과가 있는 원료와 혼합하여 사용될 수 있다.
상기 주름 개선 효과가 있는 원료는 비타민 A, 비타민 A 유도체(레티닐팔미테이트, 레티닐아세테이트 등), 아데노신, 폴리에톡실레이티드레틴아마이드일 수 있으나, 이에 한정하는 것은 아니다.
상기 화장료 조성물은 로션, 스킨소프너, 스킨토너, 아스트리젠트, 크림, 파운데이션, 에센스, 팩, 마스크팩, 비누, 바디클렌져, 클렌징폼, 바디오일 및 바디로션으로 이루어지 군에서 선택되는 하나 이상의 제형으로 이루어질 수 있으나, 이에 제한되지 않는다.
상기 화장료 조성물은 매일 사용할 수 있으며, 또한, 정해지지 않은 기간 동안에도 사용할 수 있고, 바람직하게는 사용자의 연령, 피부상태 또는 피부타입에 따라 사용량, 사용횟수 및 기간을 조절할 수 있다.
또한, 본 발명은 상기 아세틸콜린 수용체 결합 펩타이드를 포함하는 아세틸콜린 수용체 과잉활성 관련 질환의 예방 또는 치료용 약학 조성물을 제공한다.
상기 약학 조성물은 아세틸콜린 수용체와 결합하여 아세틸콜린 수용체 활성화를 억제함으로써 상기 아세틸콜린 수용체 과잉활성 관련 질환을 예방 또는 치료할 수 있다.
상기 아세틸콜린 수용체 과잉활성 관련 질환은 근육이 비정상적으로 과하게 수축하는 현상이 있는 질환으로, 경부근긴장이상증(cervical dystonia), 사지 긴장 이상증(limb dystonia), 체부 근육긴장이상증(truncal dystonia), 안검경련(blepharospasm, 얼굴떨림), 경직(spasticity), 반측안면경련(hemifacial spasm), 사시(strabismus), 안구떨림(nystagmus), 틱(tics), 만성 통증(chronic pain), 만성 편두통(chronic migraine), 신경인성 방광(neurogenic bladder), 배뇨근-괄약근 협동장애(detrusor-sphincter dyssynergia), 식도이완 불능증(achalasia cardia), 다한증(hyperhidrosis), 타액 분비 과다(sialorrhea), 소아 뇌성마비, 뇌졸중 후 근육강직, 요통, 전립선 비대증, 요실금, 성대 결절 및 교정, 치질, 치열 등일 수 있다.
또한, 상기 약학 조성물은 수술시 근육의 이완을 촉진하여 수술의 편의성을 확보하는데 사용될 수 있으며, 니코틴 중독에 의한 질환의 치료제 또는 보조제, 주름 제거, 사각턱 또는 종아리 교정을 위해 사용될 수 있으나, 이에 한정되는 것은 아니다.
상기 약학 조성물은 상기 아세틸콜린 수용체 결합 펩타이드 및 약학적으로 허용 가능한 부형제를 포함할 수 있다.
상기 약학 조성물은 각각 통상의 방법에 따라 산제, 과립제, 정제, 캡슐제, 현탁액, 에멀젼, 시럽, 에어로졸 등의 경구형 제형, 외용제, 좌제 및 멸균 주사용액의 형태로 제형화하여 사용될 수 있다. 상기 약학 조성물에 포함될 수 있는 담체, 부형제 및 희석제로는 락토즈, 덱스트로즈, 수크로스, 솔비톨, 만니톨, 자일리톨, 에리스리톨, 말티톨, 전분, 아카시아 고무, 알지네이트, 젤라틴, 칼슘 포스페이트, 칼슘 실리케이트, 셀룰로스, 메틸 셀룰로스, 미정질 셀룰로스, 폴리비닐 피롤리돈, 물, 메틸히드록시벤조에이트, 프로필히드록시벤조에이트, 탈크, 마그네슘 스테아레이트 및 광물유를 들 수 있다. 제제화 할 경우에는 보통 사용하는 충진제, 증량제, 결합제, 습윤제, 붕해제, 계면활성제 등의 희석제 또는 부형제를 사용하여 조제된다. 경구투여를 위한 고형제제에는 정제, 환제, 산제, 과립제, 캡슐제 등이 포함되며, 이러한 고형제제는 상기 아세틸콜린 수용체 결합 펩타이드에 적어도 하나 이상의 부형제 예를 들면, 전분, 탄산칼슘, 수크로스 또는 락토즈, 젤라틴 등을 섞어 조제된다. 또한 단순한 부형제 이외에 마그네슘 스테아레이트, 탈크 같은 윤활제들도 사용된다. 경구를 위한 액상 제제로는 현탁제, 내용액제, 유제, 시럽제 등이 해당되는데 흔히 사용되는 단순 희석제인 물, 리퀴드 파라핀 이외에 여러 가지 부형제, 예를 들면 습윤제, 감미제, 방향제, 보존제 등이 포함될 수 있다. 비경구 투여를 위한 제제에는 멸균된 수용액, 비수성용제, 현탁제, 유제, 동결건조제제, 좌제가 포함된다. 비수성용제, 현탁제로는 프로필렌글리콜, 폴리에틸렌 글리콜, 올리브 오일과 같은 식물성 기름, 에틸올레이트와 같은 주사 가능한 에스테르 등이 사용될 수 있다. 좌제의 기제로는 위텝솔(witepsol), 마크로골, 트윈(tween) 61, 카카오지, 라우린지, 글리세로젤라틴 등이 사용될 수 있다.
또한, 제형에 특별한 제한을 두지는 않으나, 연고제, 로션제, 스프레이제, 패취제, 크림제, 겔제 및 젤 중에서 선택되는 1종의 제형을 갖는 피부 외용제로 사용될 수 있다. 경피 흡수를 증가시키는 제제, 예를 들면 비한정적인 의미로 특히 디메틸설폭사이드, 디메틸아세트아마이드, 디메틸포름아마이드, 계면활성제, 알코올, 아세톤, 프로필렌글리콜 또는 폴리에틸렌글리콜을 포함할 수 있다. 도포 빈도는 치료받을 대상의 연령, 성별, 체중, 치료할 특정 질환 또는 병리 상태, 질환 또는 병리 상태의 심각도, 투여경로 및 처방자의 판단에 따라 상당히 변할 수 있으며, 도포 빈도는 매달 내지 1일 10회, 바람직하게는 매주 내지 1일 4회, 더 바람직하게는 1주당 3회 내지 1일 3회, 보다 더 바람직하게는 1일 1회 또는 2회 적용될 수 있다.
본 발명의 약학 조성물은 쥐, 가축, 인간 등의 포유동물에 다양한 경로로 투여될 수 있다. 투여의 모든 방식은 예상될 수 있는데, 예를 들면, 경구, 직장 또는 정맥, 근육, 피하, 피부, 자궁 내 경막 또는 뇌혈관 내 주사에 의해 투여될 수 있다. 바람직하게는 피부 투여이다.
본 발명은 또한, 상기 아세틸콜린 수용체 결합 펩타이드를 포함하는 아세틸콜린 수용체 과잉활성 관련 질환 개선용 건강기능식품 조성물에 관한 것이다.
상기 건강기능식품 조성물은 상기 아세틸콜린 수용체 결합 펩타이드 및 식품학적으로 허용 가능한 식품 보조 첨가제를 포함할 수 있다.
본 발명의 건강기능식품 조성물은 정제, 캡슐제, 환제 또는 액제 등의 형태를 포함하며, 본 발명의 나노캡슐을 첨가할 수 있는 식품으로는, 예를 들어, 각종 식품류, 음료, 껌, 차, 비타민 복합제, 건강기능성식품류 등이 있다.
본 발명의 또 다른 하나의 양태는, 상기 아세틸콜린 수용체 결합 펩타이드를 포함하는 의료기기용 조성물을 제공한다.
상기 의료기기용 조성물은 필러일 수 있으나, 이에 한정되는 것은 아니다.
본 발명에 있어서 "필러(filler)"란, 피부 조직을 보충할 수 있는 물질로, 얼굴의 탱탱함 복원, 윤곽 개선, 주름 완화를 위해 주입되어 채우는 것을 목적으로 한다.
상기 의료기기용 조성물은 아세틸콜린 수용체 결합 펩타이드에 의해 근육의 수축을 억제하여 주름을 완화할 수 있고, 윤곽 개선 효과도 나타낼 수 있으며, 상기 아세틸콜린 수용체 결합 펩타이드가 고정된 생체적합적인 마이크로입자, 나노입자, 하이드로겔 등을 주입함으로 조직을 채워줄 수 있다.
본 발명은 아세틸콜린 수용체 저해 펩타이드 및 이의 용도에 관한 것으로, 랜덤 펩타이드 재조합 파지를 이용하여 아세틸콜린 수용체와 결합력이 높은 파지를 스크리닝하고, 파지의 DNA를 통해 아세틸콜린 수용체 결합 펩타이드를 선별하였다. 선별된 펩타이드를 이용하여 아세틸콜린 수용체와의 결합력 및 아세틸콜린 수용체 작용 억제 효과를 확인하였고, 펩타이드의 변형을 통해 아세틸콜린 수용체와의 결합에 중요한 펩타이드 부위 및 서열을 확인하였다.
또한 상기 확인된 펩타이드의 아미노산 서열을 중요한 펩타이드 부위 및 서열을 중심으로 일정한 식으로 표현하고, 상기 펩타이드의 라이브러리를 제작하였으며, 제작된 라이브러리가 전체적으로 아세틸콜린 수용체와 결합하여 그 작용을 억제하는 것을 확인하였다.
이를 통해, 본 발명의 아세틸콜린 수용체 저해 펩타이드를 이용하여 주름 개선용 화장료 조성물, 아세틸콜린 수용체 관련 질환의 예방 또는 치료를 의한 의약품 및 개선용 건강기능식품을 개발할 수 있을 것으로 기대된다.
도 1은 스크리닝하여 선별한 펩타이드(에스펩-1~11)의 아세틸콜린 수용체 결합력을 분석한 결과를 보여주고 있다.
도 2는 TE671 세포에서 선별한 펩타이드의 아세틸콜린 수용체 작용 억제 효과를 확인한 결과로, (A~C)는 Synake와 에스펩-1, 에스펩-2, 에스펩-4, 에스펩-10펩타이드의 농도에 따른 아세틸콜린에 의한 아세틸콜린 수용체 작용 억제 효과를, (D)는 니코틴에 의한 아세틸콜린 수용체 작용 억제 효과를 확인한 결과를 보여주고 있다.
도 3은 Synake, 에스펩-1, 에스펩-2 펩타이드의 TE671 세포에 대한 세포 독성 결과를 보여주고 있다. 100 ㅅM 고농도에서도 모두 세포독성이 없음을 확인하였다.
도 4는 에스펩-2의 농도에 따른 아세틸콜린 수용체와의 결합력을 확인한 결과를 보여주고 있다.
도 5는 에스펩-2의 아미노산 서열을 기반으로 양 말단이 절단된 형태의 펩타이드를 이용하여 펩타이드 크기에 따른 아세틸콜린 수용체 결합에 중요한 서열 부위를 확인한 결과로, (A)는 N-말단이 절단된 형태의 펩타이드(에스펩-2-ND1~ND5)를, (B)는 C-말단이 절단된 형태의 펩타이드(에스펩-2-CD1~CD5)를 이용하여 확인한 결과를 보여주고 있다.
도 6은 (A) 에스펩-2-ND3 및 (B)에스펩-2-ND5의 아세틸콜린 수용체 결합능을 비교한 결과를 보여주고 있다. 에스펩-2-ND3 (8mer) 까지는 결합력이 별 차이가 없으나 에스펩-2-ND5(6mer)는 에스펩-2(11mer) 대비 48배 떨어짐을 확인하였다.
도 7은 (A)에스펩-1-ND3 및 (B)에스펩-2-ND3의 아미노산 서열을 기반으로 알라닌-스캐닝 방법을 이용하여 아세틸콜린 수용체 결합에 중요한 부위를 확인한 결과이다.
도 8은 Palmitoyl가 결합되어 있는 에스펩-2와 에스펩-2의 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 팔미테이트가 결합되면서 결합력이 향상됨을 확인 할 수 있다.
도 9는 Palmitoyl-에스펩-2가 마이셀을 형성함을 확인하였다. Pal-에스펩-2가 마이셀을 이루면서 avidity 효과에 의해 결합력이 향상되었다.
도 10은 Palmitoyl-에스펩-2를 농도별로 처리하였을 때 AchR 작용 억제 효과를 에스펩-2, Synake, 분가로톡신(Bungarotoxin)과 비교한 결과이다. Pal-에스펩-2의 결합력이 향상되면서 저해능도 에스펩-2 대비 10배 정도 향상되었다.
도 11은 Palmitoyl가 결합된 에스펩-2의 TE671 세포에 대한 독성 결과를 보여주고 있다. 10 ㅅM 고농도에서도 세포독성이 없음을 확인하였다.
도 12는 에스펩-2-ND3를 야생으로 하고, 이의 아미노산 서열 중 K는 R로, R은 K 또는 N으로 치환한 펩타이드의 표면 플라스몬 공명 분석 방법과 동일한 방법으로 AchR과의 결합력을 확인한 것이다. K나 R로 치환했을 때는 활성이 유지되나 N으로 치환시키면 활성이 현저하게 떨어짐을 확인하였다.
도 13은 아세틸콜린 수용체에 대하여 최적화 펩타이드 라이브러리인 8mer L1[(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R), X는 1개의 임의의 아미노산으로 이루어진 서열], 11mer L2[XXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R), X는 1개의 임의의 아미노산으로 이루어진 서열], 14mer L3[XXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R)XXX, X는 1개의 임의의 아미노산으로 이루어진 서열], 18mer L4[XXXXX(K 또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R)XXXXX, X는 1개의 임의의 아미노산으로 이루어진 서열]을 제작한 후 바이오 패닝 단계에서 스크리닝한 파지의 아세틸콜린 수용체에 대한 결합력 및 특이도를 확인한 결과를 보여주고 있다. 박스 친 부분이 첫 번째 라이브러리로써 L1(8mer), L2(11mer), L3(14mer) 라이브러리 모두 첫 번째 라이브러리 자체가 이미 아세틸콜린 수용체에 대한 결합력과 특이성이 높음을 확인할 수 있었으나 펩타이드 크기가 제일 큰 L4(18mer) 라이브러리 자체는 아세틸콜린수용체에 대한 결합력과 특이성이 없음을 확인할 수 있다.
도 14는 최적화 펩타이드 L1(8mer) 라이브러리의 바이오패닝 4,5단계에서의 아세틸콜린 수용체에 특이적인 펩타이드 파지를 스크리닝하여 스트렙트아비딘 흡광도 대비 아세틸콜린 수용체 흡광도 비율을 분석한 결과를 보여주고 있다. 컨트롤(con)로 에스펩-2를 같이 실험하였으며 모든 펩타이드들이 에스펩-2보다 결합력과 특이성이 좋음을 확인할 수 있다.
도 15는 최적화 펩타이드 L2(11mer) 라이브러리의 바이오패닝 4,5단계에서의 아세틸콜린 수용체에 특이적인 펩타이드 파지를 스크리닝하여 스트렙트아비딘 흡광도 대비 아세틸콜린 수용체 흡광도 비율을 분석한 결과를 보여주고 있다. 컨트롤(con)로 에스펩-2를 같이 하였으며 모든 펩타이드들이 에스펩-2보다 결합력과 특이성이 좋음을 확인할 수 있다.
도 16은 최적화 펩타이드 L3(14mer) 라이브러리의 바이오패닝 4,5단계에서의 아세틸콜린 수용체에 특이적인 펩타이드 파지를 스크리닝하여 스트렙트아비딘 흡광도 대비 아세틸콜린 수용체 흡광도 비율을 분석한 결과를 보여주고 있다. 컨트롤(con)로 에스펩-2를 같이 하였으며 모든 펩타이드들이 에스펩-2보다 결합력과 특이성이 좋음을 확인할 수 있다.
도 17은 최적화 펩타이드 L1(8mer)라이브러리에서 특이적으로 우수한 펩타이드를 40개 선별하여 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 결합력이 높음을 확인하였다.
도 18은 최적화 펩타이드 L2(11mer)라이브러리에서 특이적으로 우수한 펩타이드를 40개 선별하여 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 결합력이 높음을 확인하였다.
도 19는 최적화 펩타이드 L3(14mer)라이브러리에서 특이적으로 우수한 펩타이드를 40개 선별하여 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 결합력이 높음을 확인하였다.
도 20은 선별한 최적화 펩타이드 L1(8mer) 40개 중 높은 순으로 20개 펩타이드에 대한 아세틸콜린 수용체 억제효과를 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 저해능이 좋음을 확인하였다.
도 21은 선별한 최적화 펩타이드 L2(11mer) 40개 중 높은 순으로 20개 펩타이드에 대한 아세틸콜린 수용체 억제효과를 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 저해능이 좋음을 확인하였다.
도 22는 선별한 최적화 펩타이드 L3(14mer) 40개 중 높은 순으로 20개 펩타이드에 대한 아세틸콜린 수용체 억제효과를 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 저해능이 좋음을 확인하였다.
도 23은 최적화 펩타이드 L1(8mer), L2(11mer), L3(14mer) 중 가장 친화력이 높은 대표 펩타이드에 대한 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다.
도 24는 에스펩-2와 선별된 대표 펩타이드의 아세틸콜린 수용체 억제효과를 농도별로 비교한 결과를 보여주고 있다.
도 25는 에스펩-2와 선별된 대표 펩타이드의 아세틸콜린 수용체 억제효과 배수를 비교한 결과를 보여주고 있다. L3-37 펩타이드가 에스펩-2보다 32배 억제능이 향상되었음을 확인하였다.
도 26은 대표 펩타이드인 L3-37 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 27은 대표 펩타이드인 L3-28 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 28은 대표 펩타이드인 L3-27 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 29는 대표 펩타이드인 L2-110 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 30은 Myristic-L3-37과 Stearic-L3-37의 마이셀을 형성한 결과를 보여주고 있다.
도 31은 Palmitoyl-L3-37, Palmitoyl-L3-28, Palmitoyl-L3-27, Palmitoyl-L2-110을 농도별로 처리하였을 때 AchR 작용 억제 효과를 분가로톡신(Bungarotoxin)과 비교한 결과이다.
도 32는 Palmitoy-L3-37의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 33는 Palmitoy-L3-28의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 34는 Palmitoy-L3-27의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 35는 Palmitoy-L2-110의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 36은 Palmitoyl-L3-37 펩타이드를 CatWalk 장비를 이용하여 동물 효능을 평가한 결과를 보여주고 있다.
도 37는 Palmitoyl-L3-37 펩타이드를 농도별로 동물 효능 평가를 하기 위해 DAS 분석법을 이용한 결과를 보여주고 있다.
도 38는 Palmitoyl-L3-37 펩타이드의 동물 효능을 DAS 분석법으로 평가한 뒤 회복 되는 결과를 보여주고 있다.
도 39은 Palmitoyl-L3-37 펩타이드의 동물 급성 독성 평가 결과를 보여주고 있다.
도 2는 TE671 세포에서 선별한 펩타이드의 아세틸콜린 수용체 작용 억제 효과를 확인한 결과로, (A~C)는 Synake와 에스펩-1, 에스펩-2, 에스펩-4, 에스펩-10펩타이드의 농도에 따른 아세틸콜린에 의한 아세틸콜린 수용체 작용 억제 효과를, (D)는 니코틴에 의한 아세틸콜린 수용체 작용 억제 효과를 확인한 결과를 보여주고 있다.
도 3은 Synake, 에스펩-1, 에스펩-2 펩타이드의 TE671 세포에 대한 세포 독성 결과를 보여주고 있다. 100 ㅅM 고농도에서도 모두 세포독성이 없음을 확인하였다.
도 4는 에스펩-2의 농도에 따른 아세틸콜린 수용체와의 결합력을 확인한 결과를 보여주고 있다.
도 5는 에스펩-2의 아미노산 서열을 기반으로 양 말단이 절단된 형태의 펩타이드를 이용하여 펩타이드 크기에 따른 아세틸콜린 수용체 결합에 중요한 서열 부위를 확인한 결과로, (A)는 N-말단이 절단된 형태의 펩타이드(에스펩-2-ND1~ND5)를, (B)는 C-말단이 절단된 형태의 펩타이드(에스펩-2-CD1~CD5)를 이용하여 확인한 결과를 보여주고 있다.
도 6은 (A) 에스펩-2-ND3 및 (B)에스펩-2-ND5의 아세틸콜린 수용체 결합능을 비교한 결과를 보여주고 있다. 에스펩-2-ND3 (8mer) 까지는 결합력이 별 차이가 없으나 에스펩-2-ND5(6mer)는 에스펩-2(11mer) 대비 48배 떨어짐을 확인하였다.
도 7은 (A)에스펩-1-ND3 및 (B)에스펩-2-ND3의 아미노산 서열을 기반으로 알라닌-스캐닝 방법을 이용하여 아세틸콜린 수용체 결합에 중요한 부위를 확인한 결과이다.
도 8은 Palmitoyl가 결합되어 있는 에스펩-2와 에스펩-2의 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 팔미테이트가 결합되면서 결합력이 향상됨을 확인 할 수 있다.
도 9는 Palmitoyl-에스펩-2가 마이셀을 형성함을 확인하였다. Pal-에스펩-2가 마이셀을 이루면서 avidity 효과에 의해 결합력이 향상되었다.
도 10은 Palmitoyl-에스펩-2를 농도별로 처리하였을 때 AchR 작용 억제 효과를 에스펩-2, Synake, 분가로톡신(Bungarotoxin)과 비교한 결과이다. Pal-에스펩-2의 결합력이 향상되면서 저해능도 에스펩-2 대비 10배 정도 향상되었다.
도 11은 Palmitoyl가 결합된 에스펩-2의 TE671 세포에 대한 독성 결과를 보여주고 있다. 10 ㅅM 고농도에서도 세포독성이 없음을 확인하였다.
도 12는 에스펩-2-ND3를 야생으로 하고, 이의 아미노산 서열 중 K는 R로, R은 K 또는 N으로 치환한 펩타이드의 표면 플라스몬 공명 분석 방법과 동일한 방법으로 AchR과의 결합력을 확인한 것이다. K나 R로 치환했을 때는 활성이 유지되나 N으로 치환시키면 활성이 현저하게 떨어짐을 확인하였다.
도 13은 아세틸콜린 수용체에 대하여 최적화 펩타이드 라이브러리인 8mer L1[(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R), X는 1개의 임의의 아미노산으로 이루어진 서열], 11mer L2[XXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R), X는 1개의 임의의 아미노산으로 이루어진 서열], 14mer L3[XXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R)XXX, X는 1개의 임의의 아미노산으로 이루어진 서열], 18mer L4[XXXXX(K 또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R)XXXXX, X는 1개의 임의의 아미노산으로 이루어진 서열]을 제작한 후 바이오 패닝 단계에서 스크리닝한 파지의 아세틸콜린 수용체에 대한 결합력 및 특이도를 확인한 결과를 보여주고 있다. 박스 친 부분이 첫 번째 라이브러리로써 L1(8mer), L2(11mer), L3(14mer) 라이브러리 모두 첫 번째 라이브러리 자체가 이미 아세틸콜린 수용체에 대한 결합력과 특이성이 높음을 확인할 수 있었으나 펩타이드 크기가 제일 큰 L4(18mer) 라이브러리 자체는 아세틸콜린수용체에 대한 결합력과 특이성이 없음을 확인할 수 있다.
도 14는 최적화 펩타이드 L1(8mer) 라이브러리의 바이오패닝 4,5단계에서의 아세틸콜린 수용체에 특이적인 펩타이드 파지를 스크리닝하여 스트렙트아비딘 흡광도 대비 아세틸콜린 수용체 흡광도 비율을 분석한 결과를 보여주고 있다. 컨트롤(con)로 에스펩-2를 같이 실험하였으며 모든 펩타이드들이 에스펩-2보다 결합력과 특이성이 좋음을 확인할 수 있다.
도 15는 최적화 펩타이드 L2(11mer) 라이브러리의 바이오패닝 4,5단계에서의 아세틸콜린 수용체에 특이적인 펩타이드 파지를 스크리닝하여 스트렙트아비딘 흡광도 대비 아세틸콜린 수용체 흡광도 비율을 분석한 결과를 보여주고 있다. 컨트롤(con)로 에스펩-2를 같이 하였으며 모든 펩타이드들이 에스펩-2보다 결합력과 특이성이 좋음을 확인할 수 있다.
도 16은 최적화 펩타이드 L3(14mer) 라이브러리의 바이오패닝 4,5단계에서의 아세틸콜린 수용체에 특이적인 펩타이드 파지를 스크리닝하여 스트렙트아비딘 흡광도 대비 아세틸콜린 수용체 흡광도 비율을 분석한 결과를 보여주고 있다. 컨트롤(con)로 에스펩-2를 같이 하였으며 모든 펩타이드들이 에스펩-2보다 결합력과 특이성이 좋음을 확인할 수 있다.
도 17은 최적화 펩타이드 L1(8mer)라이브러리에서 특이적으로 우수한 펩타이드를 40개 선별하여 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 결합력이 높음을 확인하였다.
도 18은 최적화 펩타이드 L2(11mer)라이브러리에서 특이적으로 우수한 펩타이드를 40개 선별하여 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 결합력이 높음을 확인하였다.
도 19는 최적화 펩타이드 L3(14mer)라이브러리에서 특이적으로 우수한 펩타이드를 40개 선별하여 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 결합력이 높음을 확인하였다.
도 20은 선별한 최적화 펩타이드 L1(8mer) 40개 중 높은 순으로 20개 펩타이드에 대한 아세틸콜린 수용체 억제효과를 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 저해능이 좋음을 확인하였다.
도 21은 선별한 최적화 펩타이드 L2(11mer) 40개 중 높은 순으로 20개 펩타이드에 대한 아세틸콜린 수용체 억제효과를 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 저해능이 좋음을 확인하였다.
도 22는 선별한 최적화 펩타이드 L3(14mer) 40개 중 높은 순으로 20개 펩타이드에 대한 아세틸콜린 수용체 억제효과를 비교한 결과를 보여주고 있다. 모든 최적화 펩타이드가 에스펩-2 보다 저해능이 좋음을 확인하였다.
도 23은 최적화 펩타이드 L1(8mer), L2(11mer), L3(14mer) 중 가장 친화력이 높은 대표 펩타이드에 대한 아세틸콜린 수용체와의 결합력을 비교한 결과를 보여주고 있다.
도 24는 에스펩-2와 선별된 대표 펩타이드의 아세틸콜린 수용체 억제효과를 농도별로 비교한 결과를 보여주고 있다.
도 25는 에스펩-2와 선별된 대표 펩타이드의 아세틸콜린 수용체 억제효과 배수를 비교한 결과를 보여주고 있다. L3-37 펩타이드가 에스펩-2보다 32배 억제능이 향상되었음을 확인하였다.
도 26은 대표 펩타이드인 L3-37 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 27은 대표 펩타이드인 L3-28 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 28은 대표 펩타이드인 L3-27 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 29는 대표 펩타이드인 L2-110 펩타이드 말단의 다양한 변형을 통한 아세틸콜린 수용체에 대한 결합력을 비교한 결과를 보여주고 있다.
도 30은 Myristic-L3-37과 Stearic-L3-37의 마이셀을 형성한 결과를 보여주고 있다.
도 31은 Palmitoyl-L3-37, Palmitoyl-L3-28, Palmitoyl-L3-27, Palmitoyl-L2-110을 농도별로 처리하였을 때 AchR 작용 억제 효과를 분가로톡신(Bungarotoxin)과 비교한 결과이다.
도 32는 Palmitoy-L3-37의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 33는 Palmitoy-L3-28의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 34는 Palmitoy-L3-27의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 35는 Palmitoy-L2-110의 TE671 세포에 대한 독성 결과를 보여주고 있다.
도 36은 Palmitoyl-L3-37 펩타이드를 CatWalk 장비를 이용하여 동물 효능을 평가한 결과를 보여주고 있다.
도 37는 Palmitoyl-L3-37 펩타이드를 농도별로 동물 효능 평가를 하기 위해 DAS 분석법을 이용한 결과를 보여주고 있다.
도 38는 Palmitoyl-L3-37 펩타이드의 동물 효능을 DAS 분석법으로 평가한 뒤 회복 되는 결과를 보여주고 있다.
도 39은 Palmitoyl-L3-37 펩타이드의 동물 급성 독성 평가 결과를 보여주고 있다.
이하 본 발명의 바람직한 실시예를 상세히 설명하기로 한다. 그러나 본 발명은 여기서 설명되는 실시예에 한정되지 않고 다른 형태로 구체화될 수도 있다. 오히려, 여기서 소개되는 내용이 철저하고 완전해지고, 당업자에게 본 발명의 사상을 충분히 전달하기 위해 제공하는 것이다.
<실시예 1. 바이오패닝을 통한 아세틸콜린 수용체 결합 펩타이드 스크리닝>
본 발명의 발명자들은 선행특허(한국 등록특허 제10-1971092호)에 기재된 방법에 따라 랜덤 펩타이드 라이브러리 DNA 제작 및 형질 전환을 통하여 랜덤 재조합 파지를 생산하고, 바이오 패닝(biopanning)을 통하여 아세틸콜린 수용체(AchR) 결합 펩타이드를 스크리닝 하였다. 이때 AchR와 결합력 및 특이성이 높은 4 및 5 회차 바이오패닝 투입 파지로부터 AchR 특이적인 펩타이드를 선별하여 서열 분석한 결과를 표 1에 나타내었다.
| 펩타이드 명 | 아미노산서열 |
| 에스펩-1 | WTWKGKGTLNR |
| 에스펩-2 | WTWKGRKSLLR |
| 에스펩-3 | WTWKGEDKGKN |
| 에스펩-4 | WTWKGRDKLQM |
| 에스펩-5 | WTWKGQLGQLS |
| 에스펩-6 | WTWKGGRLSAS |
| 에스펩-7 | WTWKGRQLNNQ |
| 에스펩-8 | WTWKGDNLQNN |
| 에스펩-9 | WTWKGLYQRLG |
| 에스펩-10 | WTWKGNKQVKF |
| 에스펩-11 | WTWKGETYDSK |
<실시예 2. 펩타이드의 아세틸콜린 수용체 결합력 확인>
상기 실시예 1에서 선별된 에스펩-1~11를 합성하였고, 표면 플라스몬 공명(surface plasmon resonance, SPR) 분석기기(Biacore3000, Biacore A, 스웨덴)를 이용하여 AchR과의 결합력을 비교하였다.
표면 플라스몬 공명 분석용 바이오센서 칩인 CM5 칩에 EDC/NHS를 이용하여 아세틸콜린 수용체 단백질을 고정하고, 20mM Tris(pH 7.0)의 러닝버퍼(running buffer), 속도 30㎕/분, 펩타이드(에스펩-1~11) 농도 10μM의 조건으로 수행하였으며, 최대 500초까지 결합(association) 및 해리(dissociation)를 관찰하였고, 그 결과를 도 3에 나타내었다. 이때, AchR과 결합하여 주름을 감소시킨다고 알려진 Synake를 양성대조군으로 이용하였다.
도 1에서 보듯이, 에스펩-1, 2, 4 및 10의 경우, AchR과 결합한다고 알려진 Synake와 비교하면 AchR과의 결합력이 높음을 확인하였다.
<실시예 3. 펩타이드의 아세틸콜린 수용체 작용 억제 확인>
상기 실시예 2에서 아세틸콜린 수용체와 결합력이 높은 펩타이드의 AchR 작용 억제 효과를 확인하기 위해 AchR이 과발현 되어 있는 TE671세포를 이용하였다.
TE671 세포를 10% FBS(fetal bovine serum) 및 1% 페니실린/스트렙토마이신이 포함되어 있는 DMEM 배지를 이용하여 4일 동안 배양하였다. 배양한 세포에 트립신(trypsin)을 처리하여 떼어낸 다음, 18㎜의 커버슬라이드를 넣은 12웰 세포 배양 플레이트에 웰 당 2×14개의 세포가 되도록 분주하고 4일 동안 배양하였다. 새로운 12웰 플레이트에 세포가 배양된 커버슬라이드를 옮기고 HBSS(Hanks' balanced salt solution) 버퍼 997㎕와 Fura-2-AM(세포내 칼슘 이온 표지자) 3㎕을 넣고 37℃, 5% CO2 조건의 세포 배양기에서 15분간 배양하였다. 이후에 HBSS 버퍼로 3~4회 세척하여 남아 있는 Fura-2-AM을 제거한 후, HBSS 버퍼 1㎖를 넣었다. 세포가 자라있는 커버슬라이드를 현미경 관찰용 챔버(chamber)에 끼우고 HBSS 버퍼 500㎕를 넣은 후, 상기 실시예 2-3에서 선별된 펩타이드를 0.5, 5 또는 50μM이 되도록 첨가한 후 AchR과 결합하도록 3분 정도 방치하였다. 이후에 니코틴 125μM 또는 아세틸콜린 1μM을 첨가하여 세포 내 Fura-2의 형광 세기를 측정하여 AchR의 작용 여부를 확인하였으며, Synake 및 분가로톡신(bungarotoxin)을 양성대조군으로 처리하였고, 그 결과를 도 2에 나타내었다.
도 2에서 보듯이, 아세틸콜린을 처리한 경우(A~C)에, 에스펩-1은 0.5μM에서는 5%의 AchR 작용 억제 효과를 보이나 5μM에서는 90%, 50μM에서는 100% AchR 작용 억제 효과가 나타나는 것을 확인하였고, 에스펩-2는 0.5μM에서 90%, 5μM 이상에서는 100%의 AchR 작용 억제 효과를 나타냈으며, 에스펩-4 및 10은 50μM에서 각각 17%, 47%의 AchR 작용 억제 효과를 나타내는 것을 확인하였다. 또한, 니코틴을 처리한 경우(D)에, Synake에 비해 에스펩-1 및 2가 니코틴의 아세틸콜린 수용체에 대한 작용 억제 효과가 우수함을 확인하였다.
이를 통해, 에스펩-1 및 에스펩-2가 AchR 작용 억제 효과가 우수함을 알 수 있었고, 특히나, 에스펩-2가 AchR과 결합력이 가장 높고, 이로 인한 AchR 작용 억제 효과가 가장 우수함을 알 수 있었다.
<실시예 4. Synake, 에스펩1, 에스펩-2 세포 독성 측정>
에스펩-1, 에스펩-2의 독성을 평가하기 위해서 TE671 세포에서 WST Assay를 실시하였다. 이때, Synake의 독성 평가도 함께 실시하였고, 그 결과를 도 3에 나타내었다.
도 3에서 보듯이, 에스펩-1, 에스펩-2 및 Synake 모두 100ㅅM 처리 농도에서도 독성이 나타나지 않는 것을 확인하였다.
<실시예 5. 가장 높은 결합력을 가진 에스펩-2 펩타이드의 아세틸콜린 수용체와의 결합능 확인>
상기 실시예 2에서 AchR과의 결합력이 가장 높은 에스펩-2를 이용하여 AchR과의 결합능을 확인하기 위해 상기 실시예 2의 표면 플라스몬 공명 분석기기를 이용하여 동일한 방법을 통해 해리상수(dissociation constant, Kd)를 분석하였고, 그 결과를 도 4에 나타내었다. 이때, 에스펩-2의 농도는 0.12~1μM이 되도록 처리하였고, 양성대조군으로 Synake를 처리하여 해리상수를 비교하였다.
도 4에서 보듯이, 에스펩-2를 흘려주면 결합(association) 곡선이 급격히 증가하였고, 에스펩-2 처리 농도 의존적으로 결합 곡선이 증가하는 것을 확인하였다. 에스펩-2를 모두 흘려주고 난 후, 해리(dissociation)를 위해 러닝 버퍼만 흘려주어 과정에서 증가된 결합 곡선이 감소되지 않는 것을 통해 에스펩-2가 AchR와 지속적으로 결합하고 있음을 알 수 있었다.
또한, 에스펩-2의 AchR에 대한 Kd 값을 분석한 결과, 1.2μM로 Synake의 2,300μM과 대비하여 AchR과의 결합능이 약 2,000배 정도 우수함을 확인하였다.
<실시예 6. 펩타이드의 아세틸콜린 수용체 결합에 중요한 서열 부위 확인>
실시예 6-1. 말단의 아미노산의 순차적 삭제를 통한 아세틸콜린 수용체와의 결합력 확인
AchR과의 결합에 중요하게 작용하는 펩타이드의 부위를 확인하기 위해 상기 실시예 6에서 확인된 에스펩-2의 아미노산 서열을 기반으로 하여 펩타이드를 변형하고, 변형된 펩타이드를 상기 실시예 2의 표면 플라스몬 공명 분석기기에 적용하여 AchR과의 결합력을 확인하였다.
상기 에스펩-2의 아미노산 서열을 기반으로 하여 펩타이드의 N-말단 또는 C-말단의 아미노산 서열을 하나씩 단계적으로 삭제하여 하기 표 2의 펩타이드를 합성하고, 합성한 펩타이드를 이용해 AchR과의 결합력을 비교하여 그 결과를 도 5에 나타내었다.
| 펩타이드 명 | 아미노산서열 |
| 에스펩-2 | WTWKGRKSLLR |
| 에스펩-2-ND1 | TWKGRKSLLR |
| 에스펩-2-ND2 | WKGRKSLLR |
| 에스펩-2-ND3 | KGRKSLLR |
| 에스펩-2-ND4 | GRKSLLR |
| 에스펩-2-ND5 | RKSLLR |
| 에스펩-2-CD1 | WTWKGRKSLL |
| 에스펩-2-CD2 | WTWKGRKSL |
| 에스펩-2-CD3 | WTWKGRKS |
| 에스펩-2-CD4 | WTWKGRK |
| 에스펩-2-CD5 | WTWKGR |
도 5에서 보듯이, 에스펩-2의 N-말단의 아미노산 서열을 단계적으로 삭제시킨 경우(A)에, 에스펩-2-ND1~3은 AchR과의 결합력이 에스펩-2에 비해 50% 정도 감소되나 여전히 AchR과의 결합이 유지되고 있는 반면에, 에스펩-2-ND4~5는 결합력이 현저히 감소되는 것을 확인하였다. 또한, 에스펩-2의 C-말단의 아미노산 서열을 단계적으로 삭제시킨 경우(B)에는 C-말단의 R(Arg)이 삭제된 에스펩-2-CD1~5 모두 AchR과의 결합이 급격히 감소하는 것을 확인하였다.
이를 통해, AchR과 결합하는 에스펩-2의 아미노산 서열 중 4번째의 K(Lys)과 11번째의 R(Arg)이 AchR과의 결합에 있어서 중요한 부위임을 알 수 있었다.
실시예 6-2. 변형된 펩타이드의 아세틸콜린 수용체와의 결합능 확인
상기 실시예 6-1에서 확인된 에스펩-2의 변형된 펩타이드 중, AchR과의 결합에 필요한 가장 최소한의 서열을 가진 것으로 확인된 에스펩-2-ND3와 에스펩-2-ND3보다 더 작은 크기를 가진 에스펩-2-ND5를 이용하여 상기 실시예 2의 표면 플라스몬 공명 분석기기에 적용하여 AchR과의 결합력을 확인하였고, 그 결과를 도 6에 나타내었다.
도 6에서 보듯이, (A)의 에스펩-2-ND3의 경우, (B)의 에스펩-2-ND5와 비교하면 낮은 농도에서도 AchR과의 결합력이 높은 것을 확인하였고, 해리상수를 확인한 결과, 에스펩-2-ND3는 3.1μM인 반면에, 에스펩-2-ND5는 57μM로 에스펩-2-ND3가 에스펩-2-ND5 보다 AchR과의 결합능이 현저히 우수함을 알 수 있었다.
이를 통해 11mer인 에스펩-2와 8mer인 에스펩-2-ND3는 Ach와의 결합력이 큰 차이가 나지 않는 반면에, 6mer인 에스펩-2-ND5는 Ach와의 결합력이 현저히 감소하는 것을 알 수 있었다. 또한 상기 결과를 통해 에스펩-2의 N-말단의 3개의 아미노산은 AchR와의 결합력에 크게 중요하지 않음을 확인할 수 있었다.
<실시예 7. 알라닌-스캐닝을 이용한 펩타이드의 아세틸콜린 수용체 결합에 중요한 부위 확인>
상기 실시예 6에서 확인한 펩타이드 중, 에스펩-2-ND3와 에스펩-2와 아미노산 서열 유사성이 높은 에스펩-1의 N-말단의 아미노산 3개를 삭제한 에스펩-1-ND3(KGKGTLNR)을 이용하여, 각 펩타이드의 서열 중 중요한 부분을 확인하고자 알라닌-스캐닝을 진행하였다.
에스펩-1-ND3 및 에스펩-2-ND3를 야생으로 하고, 각각의 펩타이드의 아미노산 서열을 순차적으로 알라닌(alanine, Ala, A)으로 치환한 하기 표 3의 펩타이드를 합성하여 상기 실시예 2의 표면 플라스몬 공명 분석과 동일한 방법으로 AchR과의 결합력을 비교하였다. 이때, 각 펩타이드의 농도는 20μM가 되도록 처리하였고, 그 결과를 도 7에 나타내었다.
| 펩타이드 명 | 아미노산서열 | 펩타이드 명 | 아미노산서열 |
| 에스펩-1-ND3(WT) | KGKGTLNR | 에스펩-2-ND3(WT) | KGRKSLLR |
| 에스펩-1-ND3(K1A) | AGKGTLNR | 에스펩-2-ND3(K1A) | AGRKSLLR |
| 에스펩-1-ND3(G2A) | KAKGTLNR | 에스펩-2-ND3(G2A) | KARKSLLR |
| 에스펩-1-ND3(K3A) | KGAGTLNR | 에스펩-2-ND3(R3A) | KGAKSLLR |
| 에스펩-1-ND3(G4A) | KGKATLNR | 에스펩-2-ND3(K4A) | KGRASLLR |
| 에스펩-1-ND3(T5A) | KGKGALNR | 에스펩-2-ND3(S5A) | KGRKALLR |
| 에스펩-1-ND3(L6A) | KGKGTANR | 에스펩-2-ND3(L6A) | KGRKSALR |
| 에스펩-1-ND3(N7A) | KGKGTLAR | 에스펩-2-ND3(L7A) | KGRKSLAR |
| 에스펩-1-ND3(R8A) | KGKGTLNA | 에스펩-2-ND3(R8A) | KGRKSLLA |
도 7에서 보듯이, 에스펩-1-ND3의 1번째 K, 3번째 K, 8번째 R을 A로 치환한 경우 AchR에 대한 결합력이 감소하였고(A), 에스펩-2-ND3의 1번째 K, 3번째 R, 4번째 K, 8번째 R을 A로 치환한 경우 AchR에 대한 결합력이 감소하는 것을 확인하였다.
이를 통해, 에스펩-1-ND3 및 에스펩-2-ND3의 K 및 R 부분이 AchR과의 결합에 있어서 중요한 부위임을 알 수 있었다.
<실시예 8. 펩타이드 변형에 따른 지방산 유도체가 결합된 에스펩-2의 아세틸콜린 수용체와의 결합력 확인>
AchR에 결합하는 펩타이드의 변형에 따른 AchR과의 결합력 변화를 확인하고자 하였다. 구체적으로, 상기 실시예 2에서 확인된 펩타이드 중, AchR에 대한 결합력이 가장 높았던 에스펩-2에 지방산 유도체인 팔미토일(palmitoyl)을 결합시킨 펩타이드(Palmitoyl-에스펩-2)를 이용하여, 상기 실시예 2의 표면 플라스몬 공명 분석 방법과 동일한 방법을 통해 AchR과의 결합력을 확인하였고, 그 결과를 도 8에 나타내었다. 이때, 에스펩-2와 AchR과의 결합력을 비교군으로 이용하였다.
도 8에서 보듯이, 에스펩-2에 비해 Palmitoyl-에스펩-2가 AchR과의 결합력이 더 강하다는 것을 확인하였다. 이유는 도 9에서 보듯이 지용성인 Palmitoyl이 안으로 모이고 수용성인 에스펩-2가 밖으로 노출되는 마이셀 구조를 형성하여 AchR의 여러 부위에 결합하는 Avidity 효과를 가지기 때문이다.
<실시예 9. Palmitoyl-에스펩-2의 아세틸콜린 수용체 작용 억제 확인>
상기 실시예 8에서 AchR과의 결합력이 우수하다고 확인된 Palmitoyl-에스펩-2의 AchR 작용 억제 효과를 확인하였다. 이때, 상기 실시예 3과 동일한 방법으로 AchR 작용 억제 효과를 확인하였고, Palmitoyl-에스펩-2를 농도별로 처리하였다. AchR 작용 억제 효과 비교를 위해 에스펩-2, Synake, 분가로톡신(Bungarotoxin)을 농도별로 처리하여 대조군으로 이용하였고, 그 결과를 도 10에 나타내었다.
도 10에서 보듯이, Palmitoyl-에스펩-2가 에스펩-2에 비해 AchR에 대한 억제 효과가 더 우수하다는 것을 알 수 있었다. 더 구체적으로 각 펩타이드의 AchR에 대한 IC50 값을 살펴보면, Synake는 75ㅅM, 에스펩-2는 750nM, Palmitoyl-에스펩-2는 75nM, 분가로톡신은 7.5nM로, Pal-에스펩-2가 에스펩-2보다 AchR에 대한 작용 억제 효과가 10배 더 우수함을 확인하였다.
<실시예 10. Palmitoyl-에스펩-2 세포 독성 측정>
Palmitoyl-에스펩-2의 독성을 평가하기 위해서 TE671 세포에서 WST assay를 실시하였고, 그 결과를 도 11에 나타내었다.
도 11에서 보듯이, Palmitoyl-에스펩-2는 10ㅅM로 처리하더라도 세포 독성이 나타나지 않음을 확인하였다.
<실시예 11. 에스펩-2-ND3의 아미노산 서열 중 K와 R의 아미노산 치환을 통한 AchR과의 결합력 변화 확인>
상기 실시예 7에서 확인된 AchR과의 결합에 있어서 중요한 부위임이 확인된 에스펩-2-ND3의 K 및 R의 아미노산을 K는 R로, R은 K 또는 N으로 치환할 경우 AchR과의 결합력에 변화가 있는지 확인하였다.
에스펩-2-ND3를 야생으로 하고, 이의 아미노산 서열 중 K는 R로, R은 K 또는 N으로 치환한 하기 표 4의 펩타이드를 합성하여 상기 실시예 2의 표면 플라스몬 공명 분석 방법과 동일한 방법으로 AchR과의 결합력을 확인하였다. 이때, 각 펩타이드의 농도는 20μM가 되도록 처리하였고, 그 결과를 도 12에 나타내었다.
| 펩타이드명 | 아미노산 서열 |
| 에스펩-2-ND3(WT) | KGRKSLLR |
| 에스펩-2-K1R | RGRKSLLR |
| 에스펩-2-R3K | KGKKSLLR |
| 에스펩-2-R3N | KGNKSLLR |
| 에스펩-2-K4R | KGRRSLLR |
| 에스펩-2-R8K | KGRKSLLK |
| 에스펩-2-K1R-R3K | RGKKSLLR |
| 에스펩-2-K1R-R3N | RGNKSLLR |
| 에스펩-2-R3N-R8N | KGNKSLLN |
| 에스펩-2-K1R-K4R | RGRRSLLR |
| 에스펩-2-K1R-R3K-K4R-R8K | RGKRSLLK |
도 12에서 보듯이, 에스펩-2-ND3의 K를 R로 치환한 펩타이드의 경우 AchR에 대한 결합력이 증가하였고, R을 K로 치환한 펩타이드의 경우에는 AchR에 대한 결합력이 약간 감소하였으나 큰 차이가 없는 것을 확인하였다. 반면에, R을 N으로 치환한 펩타이드의 경우에는 AchR에 대한 결합력이 현저히 감소하는 것을 확인하였다.이를 통해, 에스펩-2-ND3 아미노산 중 K와 R 부분은 서로 치환하여도 AchR과의 결합에 변화가 없으나, R을 N으로 치환할 경우에는 AchR과의 결합능이 없어지는 것을 알 수 있었다.
<실시예 12. 아세틸콜린 수용체에 대한 결합력이 향상된 최적화 펩타이드 선별을 위한 펩타이드 라이브러리 제작>
실시예 12-1. 랜덤 펩타이드 라이브러리 DNA 제작 및 형질 전환
상기 실시예 6, 7, 11에서 확인한 결과를 통해 에스펩-2-ND3 펩타이드 서열을 변형하여 8mer; (K/R)X(K/R)(K/R)XXX(K/R), 9mer; X(K/R)X(K/R)(K/R)XXX(K/R), (K/R)X(K/R)(K/R)XXX(K/R)X, 10mer; XX(K/R)X(K/R)(K/R)XXX(K/R), (K/R)X(K/R)(K/R)XXX(K/R)XX, X(K/R)X(K/R)(K/R)XXX(K/R)X, 11mer; XXX(K/R)X(K/R)(K/R)XXX(K/R), (K/R)X(K/R)(K/R)XXX(K/R)XXX, X(K/R)X(K/R)(K/R)XXX(K/R)XX, XX(K/R)X(K/R)(K/R)XXX(K/R)X, 12mer; X(K/R)X(K/R)(K/R)XXX(K/R)XXX, XX(K/R)X(K/R)(K/R)XXX(K/R)XX, XXX(K/R)X(K/R)(K/R)XXX(K/R)X, 13mer; XX(K/R)X(K/R)(K/R)XXX(K/R)XXX, XXX(K/R)X(K/R)(K/R)XXX(K/R)XX, 14mer; XXX(K/R)X(K/R)(K/R)XXX(K/R)XXX 및 18mer; XXXXX(K/R)X(K/R)(K/R)XXX(K/R)XXXXX [(K/R)은 K나 R 둘 중 하나, X는 랜덤 아미노산]의 랜덤 펩타이드 라이브러리를 제조하고 아세틸콜린 수용체 저해 효과를 확인하였다. 상기 랜덤 라이브러리는 전체적으로 아세틸콜린 수용체와의 결합력이 에스펩-2와 유사하거나 뛰어난 것을 확인하였다.
이하에서는 특히 아세틸콜린 수용체와의 결합력이 우수한 대표적인 펩타이드 서열인 8mer; (K/R)X(K/R)(K/R)XXX(K/R), 11mer; XXX(K/R)X(K/R)(K/R)XXX(K/R) 및 14mer; XXX(K/R)X(K/R)(K/R)XXX(K/R)XXX 와 18mer; XXXXX(K/R)X(K/R)(K/R)XXX(K/R)XXXXX [(K/R)은 K나 R 둘 중 하나, X는 랜덤 아미노산]의 랜덤 펩타이드 라이브러리를 제조하고 아세틸콜린 수용체 저해 효과를 확인한 실시예를 설명한다.
8mer; (K/R)X(K/R)(K/R)XXX(K/R), 11mer; XXX(K/R)X(K/R)(K/R)XXX(K/R), 14mer; XXX(K/R)X(K/R)(K/R)XXX(K/R)XXX 및 18mer; XXXXX(K/R)X(K/R)(K/R)XXX(K/R)XXXXX [(K/R)은 K나 R 둘 중 하나, X는 랜덤 아미노산]의 펩타이드 라이브러리를 제조하기 위해서 하기 표 5의 DNA 라이브러리를 합성하였다(바이오니아, 한국). 이후, 합성한 표 5의 DNA 라이브러리를 주형으로, 두 개의 단일 가닥의 프라이머(TTCTATGCGGCCCAG, AACAGTTTCTGCGGC)를 이용하여 PCR을 수행하여 이중 가닥의 삽입용 DNA(insert DNA)를 증폭하였다. 증폭한 삽입용 DNA와 파지미드 벡터(phagemid vector)인 pIGT를 제한 효소인 Sfi I 및 Not I로 처리하고, 각각의 DNA 정제하였다. 정제한 삽입용 DNA와 pIGT를 T4 DNA 리가아제를 이용하여 연결한 후, 에탄올로 침전시켜 얻은 침전물을 TE(Tris- EDAT) 버퍼로 용해시켜 랜덤 펩타이드 라이브러리 DNA를 제작하였다.
제작한 랜덤 펩타이드 라이브러리 DNA를 컴피턴트 세포(competent cell)에 넣어 혼합한 후, 전기천공법(electroporation)을 이용하여 형질전환 시켰다. 형질전환 시킨 세포를 20mM 글루코스가 포함된 LB(luria bertani) 액체 배지에 넣고 시험관으로 옮겨 37℃에서 200rpm으로 1시간 동안 배양하였다. 배양한 세포를 20mM 글루코스 및 50㎍/㎖ 암피실린이 포함된 LB 액체 배지에 넣고 30℃에서 하루 동안 배양하였다. 하루 배양 후 4℃에서 4,000×g로 20분 동안 원심 분리하여 상층액을 제거하여 침전된 세포들을 확보하였다. 확보한 세포에 LB 액체배지로 현탁한 후, 글리세롤을 최종 농도가 20% 이상 되도록 넣고, -80℃에 보관하여 랜덤 펩타이드 라이브러리를 확보하였다.
| 라이브러리 | 아미노산 시퀀스 | 염기서열 |
| 8mer (L1) |
(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R) , X=random amino acids | TTCTATGCGGCCCAGCTGGCC ARANNKARAARANNKNNKNNKARA GCGGCCGCAGAAACTGTT |
| 11mer (L2) |
XXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R), X=random amino acids | TTCTATGCGGCCCAGCTGGCC NNKNNKNNKARANNKARAARANNKNNKNNKARA GCGGCCGCAGAAACTGTT |
| 14mer (L3) |
XXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R)XXX, X=random amino acids | TTCTATGCGGCCCAGCTGGCC NNKNNKNNKARANNKARAARANNKNNKNNKARANNKNNKNNK GCGGCCGCAGAAACTGTT |
| 18mer |
XXXXX(K또는 R)-X-(K 또는 R)(K 또는 R)-XXX-(K 또는 R)XXXXX, X=random amino acids | TTCTATGCGGCCCAGCTGGCC NNKNNKNNKNNKNNKARANNKARAARANNKNNKNNKARANNKNNKNNKNNKNNK GCGGCCGCAGAAACTGTT |
| N : A or C or G or T K : G or T R : A or G X : Random amino acids |
||
실시예 12-2. 최적화 펩타이드 재조합 파지 생산
상기 실시예 12-1에서 확보한 랜덤 펩타이드 라이브러리를 SB 액체 배지(3% tryptone, 2% yeast extract, 1% MOPS free acid, 2% glucose)에 추가하여 20분 동안 37℃에서 200rpm으로 배양하고, 1×1010pfu 헬퍼파지(helper phage)와 최종 농도가 50㎍/㎖이 되도록 암피실린을 넣고 다시 1시간 동안 같은 조건으로 배양하였다. 배양액을 50㎍/㎖ 암피실린 및 10㎍/㎖ 카나마이신(kanamycin)이 포함된 SB 액체 배지에 옮겨 같은 조건으로 16시간 배양하여 랜덤 펩타이드 재조합 파지를 생산하였다.
생산된 랜덤 펩타이드 재조합 파지를 4℃에서 5,000rpm으로 10분 동안 원심 분리하여 상층액을 얻고, 상층액에 PEG(polyethylene glycol)/NaCl(20% PEG, 15% NaCl)을 5:1[v:v]로 혼합하여 얼음에 1시간 동안 방치한 후, 4℃, 13,000rpm으로 20분 동안 원심 분리하여 상층액을 제거하고 1㎖의 PBS(phosphate buffered saline)로 침전물을 현탁하여 랜덤 펩타이드 재조합 파지를 확보하였다.
<실시예 13. 아세틸콜린 수용체에 대한 결합력이 향상된 최적화 펩타이드 스크리닝>
실시예 13-1. 바이오패닝(biopanning) 방법
항체를 표면 발현하는 파지 라이브러리를 고정화된 항원에 처리하여 항원에 결합하는 항체 후보를 선별하는 과정을 바이오패닝(biopanning)이라 하며, 결합/세척/유리(bind/wash/elution)의 세 단계로 구성된다. 세척 과정 중 결합력이 약한 항체를 갖는 파지는 제거되고 결과적으로 결합력이 높은 항체를 발현하고 있는 파지만 남게 되는데, 이러한 과정을 3~4회 반복하면 항원 결합력과 특이도가 우수한 항체 후보들을 발굴할 수 있다. 이에, 본 발명의 아세틸콜린 수용체에 대한 결합력과 특이도가 우수한 아세틸콜린 수용체 결합 펩타이드를 스크리닝하기 위해 바이오패닝 방법을 이용하였다.
10㎍/㎖의 AchR α1을 96웰 플레이트의 8개 웰에 웰 당 50㎕씩 넣고, 4℃에서 하룻밤 동안 방치하였다. 다음날 200㎕ Tris(20mM pH 7)로 1회 세척 후, 2% BSA(bovine serum albumin) 200㎕를 넣고 상온에서 2시간 동안 블로킹(blocking) 시킨 뒤 용액을 모두 버리고 200㎕ Tris(20mM pH 7)로 3회 세척하였다. 상기 실시예 13-2의 PBS로 현탁한 랜덤 펩타이드 재조합 파지(투입 파지, input phage) 400㎕ 및 2% BSA 400㎕를 혼합하여 웰 당 100㎕씩 넣고 30℃에서 1시간 동안 방치하였다. 웰의 용액을 모두 제거하고 Tris(20mM pH 7)로 3회 세척한 뒤 0.2M 글리신(glycine)(pH 2.2)을 웰 당 100㎕씩 넣어 20분간 파지를 유리시킨 후, 하나의 e-튜브에 모두 모으고, 1M Tris(pH 9.0) 200㎕를 넣어 중화시켰다(산출 파지, output phage).
바이오패닝을 반복하기 위해 유리된 파지 500㎕를 5㎖의 대장균(E. coli)과 섞어 37℃에서 200rpm으로 30분 동안 배양한 후, 1×1010pfu 헬퍼 파지와 최종 농도가 50㎍/㎖이 되도록 암피실린을 첨가하여 30분 동안 추가 배양하였다. 배양액을 50㎍/㎖ 암피실린 및 10㎍/㎖ 카나마이신이 포함된 SB 액체 배지에 옮겨 같은 조건으로 하룻밤 동안 배양하여 랜덤 펩타이드 재조합 파지를 재생산하였다. 재생산한 랜덤 펩타이드 재조합 파지를 4℃에서 5,000rpm으로 10분 동안 원심 분리하여 상층액을 얻고, 상층액에 PEG/NaCl을 5:1[v:v]로 혼합하여 얼음에 1시간 동안 방치한 후, 4℃, 13,000rpm으로 20분 동안 원심 분리하여 상층액을 제거하고 1㎖의 PBS(phosphate buffered saline)로 침전물을 현탁한 다음, 2차 바이오패닝에 사용하였다.
각 바이오패닝 단계마다 상기와 동일한 방법으로 랜덤 재조합 파지를 재생산하고, 상기의 방법과 동일한 방법으로 바이오패닝을 수행하였다. 이때, 바이오패닝 단계에서 Tris(20mM pH 7)로 세척하는 과정의 횟수를 3회 또는 6회로 하여 AchR에 대하여 5회에 걸쳐 바이오패닝을 실시하였다.
각 바이오패닝마다 투입 파지(input phage) 및 산출 파지(output phage) 수를 측정하기 위해 600㎚에서의 흡광도가 0.7(OD600=0.7)인 대장균(E. coli)에 섞어 주어 암피실린이 포함된 아가 배지(agar plate)에 도말하였고, 그 결과를 하기 표 6~9에 나타내었다.
| 8mer 바이오패닝 | ||||
| 회차 | 세척 횟수 | 투입 파지 | 산출 파지 | 산출/투입 |
| 1st | 3회 | 10.4 x 1011 | 264 x106 | 25.4x10-5 |
| 2nd | 3회 | 2.752 x1011 | 82 x106 | 29.8x10-5 |
| 3rd | 6회 | 1.024 x1011 | 42.8 x106 | 41.8x10-5 |
| 4th | 6회 | 15.8 x1011 | 144 x106 | 9.11x10-5 |
| 5th | 6회 | 12.96 x1011 | 44.8 x106 | 3.46x10-5 |
| 11mer 바이오패닝 | ||||
| 회차 | 세척 횟수 | 투입 파지 | 산출 파지 | 산출/투입 |
| 1st | 3회 | 1.14x1012 | 1.048x108 | 8.77x10-5 |
| 2nd | 6회 | 9.28x1011 | 3.4x106 | 0.366x10-5 |
| 3rd | 6회 | 9.4x1011 | 6.96x107 | 7.4x10-5 |
| 4th | 6회 | 1.02x1011 | 2.32x107 | 22.74x10-5 |
| 14mer 바이오패닝 | ||||
| 회차 | 세척 횟수 | 투입 파지 | 산출 파지 | 산출/투입 |
| 1st | 3회 | 1.52 x 1011 | 112 x106 | 73.68 x10-5 |
| 2nd | 3회 | 2.112 x1011 | 86 x106 | 40.72 x10-5 |
| 3rd | 6회 | 1.0 x1011 | 32 x106 | 32 x10-5 |
| 4th | 6회 | 12.7 x1011 | 192 x106 | 15.12 x10-5 |
| 5th | 6회 | 17.54 x1011 | 29x106 | 1.65x10-5 |
| 18mer 바이오패닝 | ||||
| 회차 | 세척 횟수 | 투입 파지 | 산출 파지 | 산출/투입 |
| 1st | 3회 | 1.28 x 1012 | 5.36 x108 | 41.87x10-5 |
| 2nd | 3회 | 1.6 x 1012 | 6.4 x108 | 40x10-5 |
| 3rd | 6회 | 1.58 x 1012 | 3.92 x107 | 2.48x10-5 |
| 4th | 6회 | 1.49 x 1011 | 9.9 x106 | 6.6x10-5 |
| 5th | 6회 | 1.18 x 1012 | 1.16 x108 | 9.83x10-5 |
실시예 13-2. 랜덤 펩타이드 라이브러리 투입 재조합 파지(input phage)를 이용한 ELISA(enyzme-linked immunosorbent assay) 방법
상기 실시예 13-1의 바이오패닝의 각 회차별 투입 파지를 이용하여 스트렙트아비딘(stereptavidin) 및 AchR에 대해 ELISA를 수행하였다.
96웰 ELISA 플레이트에 10㎍/㎖의 AchR 또는 스트렙트아비딘을 각 웰당 50㎕씩 넣고, 4℃에서 하룻밤 동안 방치하였고, 다음날 Tris(20mM pH7)로 3회 세척하고, PBS로 희석한 2% BSA를 넣고 상온에서 2시간 동안 블로킹 시킨 뒤 용액을 모두 버리고 Tris(20mM pH7)로 3회 세척하였다. 상기 실시예 13-1의 표 6~9에서의 1st, 2nd, 3rd, 4th 및 5th의 투입 파지 800㎕를 10% BSA 200㎕를 혼합하여 웰 당 100㎕씩 넣고 30℃에서 1시간 동안 방치하였다. 웰의 용액을 모두 제거하고 Tris(20mM pH7)로 3회 세척한 뒤, 1:1,000으로 희석한 HRP(horseradish peroxidase)-결합 항-M13 항체(HRP-conjugated anti-M13 Ab, GE Healthcare 사)를 각 웰 당 100㎕씩 넣고 30℃에서 1시간 동안 반응시켰다. 이후, Tris(20mM pH 7)로 3회 세척한 후, 각 웰에 HRP의 기질인 테트라메틸벤지딘(tetramethylbenzidine, TMB) 용액을 100㎕씩 넣고 발색 반응을 유도한 다음 100㎕의 1M HCl을 첨가하여 반응을 중지시키고, 450㎚에서 흡광도(OD450)를 측정하였다. 그 결과를 도 13에 나타내었다.
도 13에서 보듯이 바이오패닝의 투입 파지로부터 얻은 8mer, 11mer, 14mer의 랜덤 펩타이드 라이브러리는 전체적으로 스트렙트아비딘에 비해 AchR에 대한 결합 특이성이 매우 높게 나타났다. 즉, 8mer의 1, 3, 4 및 8번째 아미노산을 리신(K) 또는 아르기닌(R)으로 고정하고, 2 및 5, 6, 7번째 아미노산을 임의의 아미노산으로 형성하여, 일반식: (K 또는 R)-X-(K 또는 R)-(K 또는 R)-XXX-(K 또는 R)로 표현되는 폴리펩타이드의 라이브러리는 2 및 5, 6, 7번째 아미노산(X)의 종류와 관계없이 전체적으로 AchR에 높은 특이성을 갖고 결합하였다.
11mer에서도 일반식: XXX-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-XXX-(K 또는 R)로 표현되는 폴리펩타이드의 라이브러리는 X로 표현되는 임의의 아미노산(1, 2, 3, 5, 8, 9 및 10번째 아미노산)의 종류와 관계없이 4, 6, 7 및 11번째의 아미노산이 리신(K) 또는 아르기닌(R)인 경우, 전체적으로 AchR에 높은 특이성을 갖고 결합하였다.
14mer에서도 마찬가지로 일반식: XXX-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-XXX-(K 또는 R)-XXX로 표현되는 폴리펩타이드의 라이브러리는 X로 표현되는 임의의 아미노산(1, 2, 3, 5, 8, 9, 10, 12, 13 및 14번째 아미노산)의 종류와 관계없이 4, 6, 7 및 11번째의 아미노산이 리신(K) 또는 아르기닌(R)인 경우, 전체적으로 AchR에 높은 특이성을 갖고 결합하였다.
상기의 결과를 통하여 본 발명에 따른 [화학식 1], [화학식 2], [화학식 3], 또는 [화학식 4]로 표현되는 아미노산 서열을 포함하는 아세틸콜린 수용체 결합 펩타이드는 AchR에 높은 특이성을 갖고 결합하는 것을 확인하였다.
[화학식 1]
X
L
-(K 또는 R)-X-(K 또는 R)-X
M
-(K 또는 R)-X
N
(상기 XL, XM, XN은 각각 독립적으로 1~8개, X는 1개의 임의의 아미노산으로 이루어진 서열을 나타냄)
[화학식 2]
(K 또는 R)-X-(K 또는 R)-(K 또는 R)-X
M-1
-(K 또는 R)
(상기 XM-1은 1~3개, X는 1개의 임의의 아미노산으로 이루어진 서열을 나타냄)
[화학식 3]
X
L
-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-X
M-1
-(K 또는 R)
(상기 XL, XM-1은 각각 독립적으로 1~3개, X는 1개의 임의의 아미노산으로 이루어진 서열을 나타냄)
[화학식 4]
X
L
-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-X
M-1
-(K 또는 R)-X
N
(상기 XL, XM-1, XN 은 각각 독립적으로 1~3개, X는 1개의 임의의 아미노산으로 이루어진 서열을 나타냄)
아미노산 서열을 중요한 펩타이드 부위 및 서열을 중심으로 일정한 식으로 표현하고, 상기 펩타이드의 라이브러리를 제작하였으며, 제작된 라이브러리가 전체적으로 아세틸콜린 수용체와 결합하여 그 작용을 억제하는 것을 확인하였다.
그리고, 도 13에서 보듯이 1회차 바이오패닝의 투입 파지부터 얻은 8mer, 11mer, 14mer의 랜덤 펩타이드 라이브러리는 스트렙트아비딘에 비해 AchR에 대한 결합 특이성이 높게 나온 반면에, 1회차 바이오패닝의 투입 파지로부터 얻은 18mer의 랜덤 펩타이드 라이브러리는 AchR에 대한 결합 특이성이 낮은 것을 확인하였다.
이를 통해, 8~14mer까지의 랜덤 펩타이드는 AchR에 대한 결합력이 유지되나, 18mer의 랜덤 펩타이드는 AchR과의 결합력이 감소되는 것을 확인하였다. 이러한 AchR과의 결합력의 차이는 랜덤 아미노산(X)의 갯수에 의해 나타나는 것으로 예상된다.
실시예 13-3. 아세틸콜린 수용체에 대한 결합력이 향상된 최적화 펩타이드 스크리닝
상기 실시예 13-1 및 13-2에서 확인된 AchR에 대한 결합력 및 특이성이 높은 4 및 5회 차 바이오패닝 투입 파지로부터 AchR에 대한 결합력이 향상된 최적화 펩타이드를 선별하기 위한 스크리닝을 진행하였다.
상기 실시예 13-1의 4 및 5회차 바이오패닝 산출 파지를 대장균에 접종시킨 후, 플라크(plaque)가 플레이트 당 100~200개 정도가 생성되도록 아가 배지에 도말하였다. 멸균된 팁을 이용하여 플라크를 50㎍/㎖의 암피실린이 포함된 SB 액체 배지 1㎖에 각각 접종하여 37℃에서 5시간 동안 진탕 배양하고, 헬퍼 파지를 30㎕씩 첨가하여 37℃, 200rpm으로 하루 동안 배양하였다. 배양액을 12,000rpm으로 2분간 원심 분리하여 상층액을 회수한 후, 2% BSA를 넣어 파지 스크리닝에 이용하였다.
96웰 ELISA 플레이트에 5㎍/㎖의 AchR 또는 스트렙트아비딘을 각 웰 당 50㎕씩 넣고, 4℃에서 하루 동안 방치하였고, 다음날 각 웰의 용액을 버리고 2% BSA를 넣고 상온에서 2시간 동안 블로킹 시킨 뒤 용액을 모두 버리고 Tris(20mM pH7)로 3회 세척하였다. 상기에서 준비한 플라크로부터 얻은 각각의 파지 용액 100㎕씩을 웰에 분주하고 30℃에서 1시간 동안 방치하였다. 웰의 용액을 모두 제거하고 Tris(20mM pH7)로 3회 세척한 뒤, 1:2,000으로 희석한 HRP-결합 항-M13 항체를 각 웰 당 100㎕씩 넣고 30℃에서 1시간 동안 반응시켰다. 이후, Tris(20mM pH7)로 3회 세척한 후, 각 웰에 테트라메틸벤지딘 용액을 100㎕씩 넣고 발색 반응을 유도한 다음 100㎕의 1M H2SO4를 첨가하여 반응을 중지시키고, 450㎚에서 흡광도(OD450)를 측정하였다. 측정한 흡광도를 이용하여 스트렙트아비딘 흡광도 대비 AchR 흡광도 비율(AchR signal/streptavidin signal)로 환산하였고, 그 결과를 도 14~16에 나타내었다.
도 14~16에서 보듯이, 파지에 따른 AchR과의 결합력이 차이가 있음을 관찰하였고, 이 중, 스트렙트아비딘 흡광도 대비 AchR 흡광도 비율이 가장 높은 파지를 선별하였다. 그 결과를 표 10~12 에 나타내었다. 8mer (L1), 11mer (L2), 14mer (L3) 최적화 펩타이드 모두 양성대조군인 에스펩-2보다 AchR과 결합력과 특이성이 높게 나옴을 확인하였다.
상기에서 선별한 파지로부터 플라스미드 DNA를 정제하고, 정제한 플라스미드 DNA와 GATTACGCCAAGCTTTGGAGC로 이루어진 염기서열 분석용 프라이머를 이용하여 펩타이드의 서열 분석을 의뢰하였고, 서열이 확인되고 AchR과의 결합 특이성이 우수한 최적화 펩타이드를 선별하였다.
또한 대한민국 공개특허 10-2014-0139010호에 기재된 아미노산 서열의 펩타이드와 비교하기 위하여 표 13과 같이 아미노산 서열의 펩타이드를 합성하여 상기와 동일한 방법으로 AchR과의 결합 특이성을 확인한 결과, 에스펩-2와 유사한 결합특이성을 확인하였다. 그러나 R이 반복된 서열, 또는 8mer의 양단으로 G가 반복된 서열은 11mer, 또는 14mer 라이브러리 전체와 비교하여 낮은 수준의 결합특이성을 나타내었다. 이는 펩타이드의 하전된 전하 또는 입체구조적 특징에 의한 것으로 사료된다.
| 서열번호 | 펩타이드(8mer) | OD | Seq. |
| 1 | L1-1 | 0.693 | KAKKIRQR |
| 2 | L1-2 | 0.68 | KKRKGSAK |
| 3 | L1-3 | 0.649 | RGKRQLGR |
| 4 | L1-4 | 0.696 | KLKKFPVR |
| 5 | L1-5 | 0.657 | RHKKSPWK |
| 6 | L1-6 | 0.652 | RPRRTHIK |
| 7 | L1-7 | 0.68 | KHKKLNQR |
| 8 | L1-8 | 0.653 | RAKRMCCK |
| 9 | L1-9 | 0.639 | RQRRNDMK |
| 10 | L1-10 | 0.621 | KRKKDWWR |
| 11 | L1-11 | 0.703 | RARKDAFK |
| 12 | L1-12 | 0.691 | KWRREGFK |
| 13 | L1-13 | 0.73 | RFRRGVRR |
| 14 | L1-14 | 0.755 | KARRQEDK |
| 15 | L1-15 | 0.67 | RFKRLCMK |
| 16 | L1-16 | 0.62 | KLKRRSRR |
| 17 | L1-17 | 0.653 | RPRRSLDR |
| 18 | L1-18 | 0.627 | KHKKQNVR |
| 19 | L1-19 | 0.575 | KFKRTEGR |
| 20 | L1-20 | 0.619 | RGKKAVRK |
| 21 | L1-21 | 0.65 | KFKKHGVK |
| 22 | L1-22 | 0.732 | KTKKIHSK |
| 23 | L1-23 | 0.712 | RDRKIHDK |
| 24 | L1-24 | 0.742 | RPRRGHQK |
| 25 | L1-25 | 0.709 | KTKKLYEK |
| 26 | L1-26 | 0.647 | RQKKDNQK |
| 27 | L1-27 | 0.662 | RQRKSMQR |
| 28 | L1-28 | 0.656 | RGRRWFVK |
| 29 | L1-29 | 0.646 | RNKRDENK |
| 30 | L1-30 | 0.605 | RLRRWCVK |
| 31 | L1-31 | 0.604 | RQKKPWVK |
| 32 | L1-32 | 0.607 | RWKRSEQR |
| 33 | L1-33 | 0.529 | RLKRAPGR |
| 34 | L1-34 | 0.634 | KMRRAQGR |
| 35 | L1-35 | 0.556 | KIKKAARR |
| 36 | L1-36 | 0.553 | KNRKNLYR |
| 37 | L1-37 | 0.524 | KCKKMTER |
| 38 | L1-38 | 0.514 | KTRRVDLR |
| 39 | L1-39 | 0.509 | KSRKWGWR |
| 40 | L1-40 | 0.424 | KRRRFRWK |
| 41 | L1-41 | 0.613 | KYKKQHTK |
| 42 | L1-42 | 0.695 | RGRKKCGK |
| 43 | L1-43 | 0.601 | KLRKQKLR |
| 44 | L1-44 | 0.59 | RNRKIPLK |
| 45 | L1-45 | 0.597 | RLRRRYRR |
| 46 | L1-46 | 0.637 | KWRKRRIK |
| 47 | L1-47 | 0.54 | KTKRMLAK |
| 48 | L1-48 | 0.519 | KEKRAVHK |
| 49 | L1-49 | 0.504 | RVRRDGHR |
| 50 | L1-50 | 0.583 | RAKKCYHK |
| 51 | L1-51 | 0.66 | KGKRDSNK |
| 52 | L1-52 | 0.56 | KVRKSLWK |
| 53 | L1-53 | 0.78 | RSKKSLYR |
| 54 | L1-54 | 0.569 | RGKRGCER |
| 55 | L1-55 | 0.585 | KQKKTGFR |
| 56 | L1-56 | 0.468 | KYRRNLVR |
| 57 | L1-57 | 0.408 | RIRRGSVR |
| 58 | L1-58 | 0.472 | RSKRMNGK |
| 59 | L1-59 | 0.479 | RHKKFIEK |
| 60 | L1-60 | 0.484 | RIRRVSQR |
| 61 | L1-61 | 0.625 | RPKKWMQK |
| 62 | L1-62 | 0.633 | KERKHEAK |
| 63 | L1-63 | 0.618 | KERKASAK |
| 64 | L1-64 | 0.589 | KMRRIMYK |
| 65 | L1-65 | 0.585 | KWKRNCTK |
| 66 | L1-66 | 0.605 | RCKKNPEK |
| 67 | L1-67 | 0.593 | KQRKPHGR |
| 68 | L1-68 | 0.55 | KAKRGHFK |
| 69 | L1-69 | 0.474 | KSKRHHEK |
| 70 | L1-70 | 0.668 | KSRKHCNR |
| 71 | L1-71 | 0.618 | RSKKINNK |
| 72 | L1-72 | 0.616 | RVRRSWIR |
| 73 | L1-73 | 0.644 | KSRRSAGK |
| 74 | L1-74 | 0.649 | KMRKHDVR |
| 75 | L1-75 | 0.581 | KGKKLSQR |
| 76 | L1-76 | 0.588 | KWRREQFR |
| 77 | L1-77 | 0.636 | KDKRWANR |
| 78 | L1-78 | 0.642 | KHKRTAQR |
| 79 | L1-79 | 0.583 | KPKREMHK |
| 80 | L1-80 | 0.631 | RWKKNINK |
| 81 | L1-81 | 0.63 | KWKKMMER |
| 82 | L1-82 | 0.596 | RIRRWWVR |
| 83 | L1-83 | 0.621 | RIRRHMYR |
| 84 | L1-84 | 0.576 | KPKKPPAR |
| 85 | L1-85 | 0.57 | RLKRGQFK |
| 86 | L1-86 | 0.551 | KNKRTPVK |
| 87 | L1-87 | 0.538 | KWRRAFGR |
| 88 | L1-88 | 0.562 | KNKKCVGR |
| 89 | L1-89 | 0.551 | RTRKCVLK |
| 90 | L1-90 | 0.439 | KIKRDLYK |
| 91 | L1-91 | 0.826 | REKRYMAR |
| 92 | L1-92 | 0.581 | RVKKAETR |
| 93 | L1-93 | 0.616 | RGRRGTMR |
| 94 | L1-94 | 0.496 | RTKRWQLK |
| 95 | L1-95 | 0.535 | RLRKSLGK |
| 96 | L1-96 | 0.494 | RPKRNWAK |
| 97 | L1-97 | 0.546 | KDRKVNSR |
| 98 | L1-98 | 0.54 | KWRRNGQR |
| 99 | L1-99 | 0.543 | RFKRMVWR |
| 100 | L1-100 | 0.737 | KERKGAVR |
| 101 | L1-101 | 0.599 | KTKRGSLK |
| 102 | L1-102 | 0.507 | KTKRVDPK |
| 103 | L1-103 | 0.55 | RARKLHNR |
| 104 | L1-104 | 0.46 | RGKKPWWR |
| 105 | L1-105 | 0.507 | KGKKYYQK |
| 106 | L1-106 | 0.417 | RWRRSVIK |
| 107 | L1-107 | 0.534 | RQKKDFLK |
| 108 | L1-108 | 0.535 | RGRKPIWK |
| 109 | L1-109 | 0.542 | KPRRPDTR |
| 110 | L1-110 | 0.422 | KVKRLWNR |
| 111 | L1-111 | 0.608 | KDRKGVYK |
| 112 | L1-112 | 0.431 | RVKRFVPK |
| 113 | L1-113 | 0.526 | KPKRDQSR |
| 114 | L1-114 | 0.42 | KDKKQWGR |
| 115 | L1-115 | 0.451 | RFRKMIPK |
| 116 | L1-116 | 0.492 | KFRKHVCK |
| 117 | L1-117 | 0.508 | RPKRSPSK |
| 118 | L1-118 | 0.471 | RYKKVPAK |
| 119 | L1-119 | 0.463 | RSRRCPVK |
| 120 | L1-120 | 0.388 | RCKRCDNK |
| 121 | L1-121 | 0.354 | RGKKQLSK |
| 122 | L1-122 | 0.352 | KSKRPEGR |
| 123 | L1-123 | 0.409 | KCRKTMGR |
| 124 | L1-124 | 0.428 | RDRKNPVR |
| 125 | L1-125 | 0.345 | KPKRSAPR |
| 126 | L1-126 | 0.362 | KGKRTGCK |
| 127 | L1-127 | 0.412 | RFRKPTDR |
| 128 | L1-128 | 0.301 | RMKKFHTR |
| 129 | L1-129 | 0.421 | KIKKYYWK |
| 130 | L1-130 | 0.368 | RIKRNNCK |
| 131 | L1-131 | 0.505 | RHKRMLWR |
| 132 | L1-132 | 0.541 | KVRRVFLK |
| 133 | L1-133 | 0.504 | KDKRPECK |
| 134 | L1-134 | 0.365 | KTRRSACR |
| 135 | L1-135 | 0.479 | KFRKGPYR |
| 136 | L1-136 | 0.392 | KFKKSAPR |
| 137 | L1-137 | 0.458 | KPRKIQGR |
| 138 | L1-138 | 0.373 | KMRKGWDK |
| 139 | L1-139 | 0.456 | KFRRQSTR |
| 140 | L1-140 | 0.381 | KNRRADCR |
| 141 | L1-141 | 0.505 | KNKRWVFK |
| 142 | L1-142 | 0.469 | RCRKDTAR |
| 143 | L1-143 | 0.416 | KARRVGTK |
| 144 | L1-144 | 0.503 | KVKKIYYR |
| 145 | L1-145 | 0.54 | KAKRDYLR |
| 146 | L1-146 | 0.362 | KVKRLPYR |
| 147 | L1-147 | 0.415 | RHRRANFR |
| 148 | L1-148 | 0.426 | KPKKGGWK |
| 149 | L1-149 | 0.34 | KEKRLYAR |
| 150 | L1-150 | 0.457 | REKKQEVK |
| 151 | L1-151 | 0.479 | KSRRAVFR |
| 152 | L1-152 | 0.484 | RHRKYVDK |
| 153 | L1-153 | 0.625 | RHKRWSGR |
| 154 | L1-154 | 0.633 | RDRKFVQK |
| 155 | L1-155 | 0.618 | RYRRWFYR |
| 156 | L1-156 | 0.589 | RQRRMTPR |
| 157 | L1-157 | 0.585 | RNKRMDGK |
| 158 | L1-158 | 0.605 | RPKKHQER |
| 159 | L1-159 | 0.593 | KKRKSLLR |
| 160 | L1-160 | 0.55 | KIKRNVFK |
| 161 | L1-161 | 0.474 | KYKREEYR |
| 162 | L1-162 | 0.668 | KIRRVTNK |
| 163 | L1-163 | 0.618 | KVKRVWGR |
| 164 | L1-164 | 0.616 | RIKRDYCK |
| 165 | L1-165 | 0.644 | RIRRAHDR |
| 166 | L1-166 | 0.649 | RARKESHK |
| 167 | L1-167 | 0.581 | RYKKQQYK |
| 168 | L1-168 | 0.588 | KIKKLSQK |
| 169 | L1-169 | 0.636 | KLKRAMLK |
| 170 | L1-170 | 0.642 | KARKNSIR |
| 171 | L1-171 | 0.583 | KERKLWWK |
| 172 | L1-172 | 0.631 | KTRKQDHR |
| 173 | L1-173 | 0.63 | RCKRFVGK |
| 174 | L1-174 | 0.596 | KEKKLVWK |
| 175 | L1-175 | 0.621 | KARRNSLK |
| 176 | L1-176 | 0.479 | KMRRVAPR |
| 177 | L1-177 | 0.484 | RAKKIMFR |
| 178 | L1-178 | 0.625 | KTKKAAER |
| 179 | L1-179 | 0.633 | RGKRHWHR |
| 180 | L1-180 | 0.618 | RTKKENVK |
| 181 | L1-181 | 0.589 | REKKYAYK |
| 182 | L1-182 | 0.585 | KWRRELPR |
| 183 | L1-183 | 0.605 | KSRRMWGR |
| 184 | L1-184 | 0.593 | KCKKDNDK |
| 185 | L1-185 | 0.55 | KDKKMPQR |
| 186 | L1-186 | 0.474 | RHKRQQDK |
| 187 | L1-187 | 0.668 | RLKKHCGK |
| 188 | L1-188 | 0.618 | KARKQEVR |
| 189 | L1-189 | 0.616 | KMRKFYSK |
| 190 | L1-190 | 0.644 | KMRRWWLK |
| 191 | L1-191 | 0.649 | KQKRQWAR |
| 192 | L1-192 | 0.581 | RSRRGIGK |
| 193 | L1-193 | 0.588 | KDKKTPCK |
| 194 | L1-194 | 0.636 | RIRKITWR |
| 195 | L1-195 | 0.642 | RHKRWEVR |
| 196 | L1-196 | 0.583 | RFRRNFHK |
| 197 | L1-197 | 0.631 | KWKRFSQR |
| 198 | L1-198 | 0.63 | KYKKSFTK |
| 199 | L1-199 | 0.596 | KTRKIVMK |
| 200 | L1-200 | 0.621 | RTKKAYVK |
| 에스펩 -2 (con) | 0.303 | WTWKGRKSLLR |
| 서열번호 | 펩타이드 (11mer) | OD | Seq. |
| 201 | L2-1 | 0.705 | EDYKYRRQNYK |
| 202 | L2-2 | 0.749 | GIWKFRRNQCK |
| 203 | L2-3 | 0.796 | SPVRWKRLCLR |
| 204 | L2-4 | 0.757 | ECYRNRKAYCR |
| 205 | L2-5 | 0.752 | TQQRLKRVMEK |
| 206 | L2-6 | 0.78 | PELKCKRMIGR |
| 207 | L2-7 | 0.753 | WQMKTKKEIWK |
| 208 | L2-8 | 0.739 | FGHRGKKEWAR |
| 209 | L2-9 | 0.721 | ATCKPKRPWYK |
| 210 | L2-10 | 0.803 | TSQKVRKMEAK |
| 211 | L2-11 | 0.791 | GIPKYRRGCSR |
| 212 | L2-12 | 0.83 | AYAKARRWGQK |
| 213 | L2-13 | 0.855 | GGLKSKRLTTR |
| 214 | L2-14 | 0.746 | GMWKVRKTVFR |
| 215 | L2-15 | 0.705 | HTTRGKRCDPR |
| 216 | L2-16 | 0.77 | VDVRNKKSNNR |
| 217 | L2-17 | 0.72 | GLNRWRRCHHK |
| 218 | L2-18 | 0.753 | LQSRSKKYSVK |
| 219 | L2-19 | 0.727 | GWNRAKREESK |
| 220 | L2-20 | 0.675 | SWQKCKKAEVR |
| 221 | L2-21 | 0.719 | MVMKWKRWNQR |
| 222 | L2-22 | 0.75 | MMMRHKRCQYR |
| 223 | L2-23 | 0.832 | QTYRLKKPLQK |
| 224 | L2-24 | 0.812 | SFWRERKNGFR |
| 225 | L2-25 | 0.842 | RLERPRRELTK |
| 226 | L2-26 | 0.809 | APWRLKRAPQR |
| 227 | L2-27 | 0.747 | QGEKYRRTESK |
| 228 | L2-28 | 0.762 | CIWKSKRSPAK |
| 229 | L2-29 | 0.756 | AILKGKRIPNK |
| 230 | L2-30 | 0.746 | PTLKWRKPVLR |
| 231 | L2-31 | 0.737 | NNSRSKRFMNR |
| 232 | L2-32 | 0.64 | LYMRCKKQAPR |
| 233 | L2-33 | 0.619 | INYRCRKWVDK |
| 234 | L2-34 | 0.604 | VMDRDRKWWWR |
| 235 | L2-35 | 0.683 | NFTKLRKPGPR |
| 236 | L2-36 | 0.76 | FIPKMRKCSPR |
| 237 | L2-37 | 0.66 | TLWKVRKAYSR |
| 238 | L2-38 | 0.88 | ESEKIKRLSTR |
| 239 | L2-39 | 0.669 | LMTKNRRNSFR |
| 240 | L2-40 | 0.685 | PEVKVRRHSMR |
| 241 | L2-41 | 0.568 | AEVKDKKAYYK |
| 242 | L2-42 | 0.508 | YYAKSKKYMVR |
| 243 | L2-43 | 0.572 | LMDRTRRDMYK |
| 244 | L2-44 | 0.579 | NMGRHKRPFLK |
| 245 | L2-45 | 0.584 | WIYRGRKDVAK |
| 246 | L2-46 | 0.704 | CYWKAKRYPMR |
| 247 | L2-47 | 0.707 | IEDKGRRINPR |
| 248 | L2-48 | 0.629 | VSTKIKKEPQR |
| 249 | L2-49 | 0.734 | YPFKAKRPAEK |
| 250 | L2-50 | 0.656 | SADRNKRHMTR |
| 251 | L2-51 | 0.653 | SLYRQKRHDYK |
| 252 | L2-52 | 0.624 | LPSRPKKPVPK |
| 253 | L2-53 | 0.614 | PAWRCKRCQPK |
| 254 | L2-54 | 0.609 | HHWRFKREMPR |
| 255 | L2-55 | 0.524 | PACRSKRDWQK |
| 256 | L2-56 | 0.713 | FICKSRKFYGK |
| 257 | L2-57 | 0.795 | LEHKERKDDFR |
| 258 | L2-58 | 0.701 | ANPKNRKDNLR |
| 259 | L2-59 | 0.69 | FGVKYRRVICR |
| 260 | L2-60 | 0.697 | HADKFRRFNMR |
| 261 | L2-61 | 0.688 | PTIRVRKSDDR |
| 262 | L2-62 | 0.736 | NLHKPKRDLPK |
| 263 | L2-63 | 0.742 | IDGKWRKICTR |
| 264 | L2-64 | 0.683 | SPFRAKRQDVR |
| 265 | L2-65 | 0.731 | PLWKTRKIEPR |
| 266 | L2-66 | 0.798 | PNYKWRKSRRR |
| 267 | L2-67 | 0.696 | KTGRSKRHRWR |
| 268 | L2-68 | 0.721 | GPDKSRRNLHR |
| 269 | L2-69 | 0.676 | WCCKTKRAVMK |
| 270 | L2-70 | 0.67 | QLLKMRKALSR |
| 271 | L2-71 | 0.651 | PYLRSKRFPPR |
| 272 | L2-72 | 0.638 | APHKWRKQEQR |
| 273 | L2-73 | 0.662 | PPFKFRKPLGR |
| 274 | L2-74 | 0.651 | CIMRVKRMWWK |
| 275 | L2-75 | 0.539 | YILRDKKVPLK |
| 276 | L2-76 | 0.725 | PWVRIRKMASR |
| 277 | L2-77 | 0.733 | VADRQKKTAPK |
| 278 | L2-78 | 0.718 | HHNRMKKQYHR |
| 279 | L2-79 | 0.689 | EGAKYRRDGWR |
| 280 | L2-80 | 0.685 | FWDRVKRNPSK |
| 281 | L2-81 | 0.705 | TMWRQRKMSCK |
| 282 | L2-82 | 0.693 | PDTRWKRVLFK |
| 283 | L2-83 | 0.65 | HQQKCKKTTTK |
| 284 | L2-84 | 0.574 | PWHKGKRDFDR |
| 285 | L2-85 | 0.768 | VFVKWRRQMMR |
| 286 | L2-86 | 0.701 | SQWKIRKRLIR |
| 287 | L2-87 | 0.716 | EWHKVRRVYAK |
| 288 | L2-88 | 0.744 | NAMRTKRMSFK |
| 289 | L2-89 | 0.749 | PPFKFRKPLGR |
| 290 | L2-90 | 0.765 | TSVKKRKQRLR |
| 291 | L2-91 | 0.517 | PIDKFKRGMVR |
| 292 | L2-92 | 0.634 | FVGRWKREYAK |
| 293 | L2-93 | 0.635 | FNPKMRKVCLR |
| 294 | L2-94 | 0.642 | NPGKTKRWLQR |
| 295 | L2-95 | 0.522 | IMDKRRKPGCR |
| 296 | L2-96 | 0.708 | ISDKAKRQHFK |
| 297 | L2-97 | 0.531 | DVNKYRKHSHK |
| 298 | L2-98 | 0.626 | LMLRGKRLTTK |
| 299 | L2-99 | 0.52 | CSLRARKEEWR |
| 300 | L2-100 | 0.551 | MNYRCKRVQER |
| 301 | L2-101 | 0.592 | NGVRERKWQSK |
| 302 | L2-102 | 0.608 | YTSRCKKQPRK |
| 303 | L2-103 | 0.571 | QYNKGRKIHVR |
| 304 | L2-104 | 0.563 | PEDRQRKTWFK |
| 305 | L2-105 | 0.488 | IEMKPRKFGVK |
| 306 | L2-106 | 0.926 | QRWKWRKSLAR |
| 307 | L2-107 | 0.681 | VKHKERKCQRR |
| 308 | L2-108 | 0.716 | QQDRPKRDIPK |
| 309 | L2-109 | 0.596 | YPTKGKRCMIR |
| 310 | L2-110 | 0.831 | RYAKHRKRQTR |
| 311 | L2-111 | 0.594 | GYFRPRKETCK |
| 312 | L2-112 | 0.646 | CFFKMRRCNTK |
| 313 | L2-113 | 0.64 | QNDKDRKLSHR |
| 314 | L2-114 | 0.643 | QVTKSKRVAFK |
| 315 | L2-115 | 0.837 | VRAKHRKSSLR |
| 316 | L2-116 | 0.699 | SHSRQRKTPLR |
| 317 | L2-117 | 0.607 | DNGKIRRCLGK |
| 318 | L2-118 | 0.65 | FLRRLKKVHWK |
| 319 | L2-119 | 0.56 | FCRRSKKIGRR |
| 320 | L2-120 | 0.607 | PAARTKRMYGR |
| 321 | L2-121 | 0.492 | HETKPKKDGLR |
| 322 | L2-122 | 0.558 | MGDRPRKWDSR |
| 323 | L2-123 | 0.473 | HDTKCKKMYAK |
| 324 | L2-124 | 0.556 | NVVRGRRLECR |
| 325 | L2-125 | 0.481 | PADKGRREVMK |
| 326 | L2-126 | 0.605 | AMMKYKKEFPK |
| 327 | L2-127 | 0.569 | YIIRWKRQMTR |
| 328 | L2-128 | 0.516 | SPWRIRRQNIR |
| 329 | L2-129 | 0.603 | TCIKYRRAHTK |
| 330 | L2-130 | 0.64 | GYDRARKGTLR |
| 331 | L2-131 | 0.462 | MTLKHRRVYIK |
| 332 | L2-132 | 0.515 | LFTRIKRLVCK |
| 333 | L2-133 | 0.526 | DFSRDRRCLSK |
| 334 | L2-134 | 0.44 | VWNKVKRWLER |
| 335 | L2-135 | 0.557 | FPGKNRKYCSR |
| 336 | L2-136 | 0.454 | YTSKNKRGCPR |
| 337 | L2-137 | 0.452 | CACKQRRATSR |
| 338 | L2-138 | 0.509 | IMERQRKSQHR |
| 339 | L2-139 | 0.528 | SVLRCRKCSMK |
| 340 | L2-140 | 0.445 | YPQKLRRTALK |
| 341 | L2-141 | 0.462 | SSHKGKRAQSK |
| 342 | L2-142 | 0.512 | DNPRFRKTILK |
| 343 | L2-143 | 0.401 | AIIKFRKVQWK |
| 344 | L2-144 | 0.521 | PYTRCRKEICK |
| 345 | L2-145 | 0.468 | MNEKPKKNDQK |
| 346 | L2-146 | 0.605 | LIGKFKKPFYR |
| 347 | L2-147 | 0.641 | TWCKHKKLDMK |
| 348 | L2-148 | 0.604 | DSNKVRKCSSK |
| 349 | L2-149 | 0.465 | LPMKQRKCEFR |
| 350 | L2-150 | 0.579 | EDVRVKRQTCR |
| 351 | L2-151 | 0.749 | TQCKDRRVSDR |
| 352 | L2-152 | 0.681 | IALKPKRVWLK |
| 353 | L2-153 | 0.688 | GHQRGKREGSR |
| 354 | L2-154 | 0.736 | NPFKYKKICPK |
| 355 | L2-155 | 0.742 | NEARIKKCDVK |
| 356 | L2-156 | 0.683 | YGLRMRKWYMK |
| 357 | L2-157 | 0.731 | NFYKCRKLQCK |
| 358 | L2-158 | 0.73 | MMTKYKKTCCK |
| 359 | L2-159 | 0.696 | CNQKTKKIAEK |
| 360 | L2-160 | 0.721 | ADIRMKKWYPK |
| 361 | L2-161 | 0.579 | AWFRVKRSNCR |
| 362 | L2-162 | 0.584 | NCDRTRKHWAR |
| 363 | L2-163 | 0.725 | PHARTRKNITK |
| 364 | L2-164 | 0.733 | VPTKMKKYETK |
| 365 | L2-165 | 0.718 | AYPKFRKTFNR |
| 366 | L2-166 | 0.579 | QQLRLRKLCGK |
| 367 | L2-167 | 0.584 | MFMRNKKLAWR |
| 368 | L2-168 | 0.725 | GGAKNKKVVSR |
| 369 | L2-169 | 0.733 | HDPRHKKTPTK |
| 370 | L2-170 | 0.718 | QVHRNKRYTDR |
| 371 | L2-171 | 0.689 | YGTRPKKYVSK |
| 372 | L2-172 | 0.685 | CTWRGRRPHDK |
| 373 | L2-173 | 0.705 | SWAKARKLVHR |
| 374 | L2-174 | 0.693 | PLFKSRRAYVR |
| 375 | L2-175 | 0.65 | CVMRCRRSEDK |
| 376 | L2-176 | 0.574 | WWHKHRRAQSK |
| 377 | L2-177 | 0.768 | LNSKPRRVEFK |
| 378 | L2-178 | 0.718 | VPYRHRRMQFK |
| 379 | L2-179 | 0.716 | GIAKSKRNAGR |
| 380 | L2-180 | 0.744 | DPEKWRKFYDR |
| 381 | L2-181 | 0.749 | PGARCRKQDVK |
| 382 | L2-182 | 0.681 | HEQRPKKGQQK |
| 383 | L2-183 | 0.688 | TCDRARKESFR |
| 384 | L2-184 | 0.736 | MHQRERRNFVK |
| 385 | L2-185 | 0.742 | FITRFRKMGEK |
| 386 | L2-186 | 0.683 | PNWKVRRFGDK |
| 387 | L2-187 | 0.731 | TAAKWKKIIMK |
| 388 | L2-188 | 0.73 | CLWRLRKDNGR |
| 389 | L2-189 | 0.696 | AHIKSRKVWSR |
| 390 | L2-190 | 0.721 | NLVKSKKVEEK |
| 391 | L2-191 | 0.689 | PDSRLKKHEAK |
| 392 | L2-192 | 0.685 | EHVKLKRLDFR |
| 393 | L2-193 | 0.705 | PALRMRRWCQK |
| 394 | L2-194 | 0.693 | HLEKHRRCEFK |
| 395 | L2-195 | 0.65 | CQAKTRKAEDR |
| 396 | L2-196 | 0.574 | QNMRMKRFIQR |
| 397 | L2-197 | 0.768 | CPYRIRRGPGK |
| 398 | L2-198 | 0.718 | DIYKGKRTLVK |
| 399 | L2-199 | 0.716 | EICKNRKPANR |
| 400 | L2-200 | 0.744 | QGMKLKRIWSK |
| 에스펩 -2 (con) | 0.303 | WTWKGRKSLLR |
| 서열번호 | 펩타이드 (14mer) |
OD | Seq. |
| 401 | L3-1 | 0.853 | ALGRTKRLHMRVHV |
| 402 | L3-2 | 0.841 | SFYKERKCQFRSGL |
| 403 | L3-3 | 0.88 | SGVRYRKWWIRSVV |
| 404 | L3-4 | 0.905 | MPQKLKKLDIRNHN |
| 405 | L3-5 | 0.82 | NEDRGKRPHIRVLG |
| 406 | L3-6 | 0.77 | EFVKLRKARLRGPQ |
| 407 | L3-7 | 0.803 | EGDKFRRHDIKYNF |
| 408 | L3-8 | 0.777 | PICRWKRAPFKWYF |
| 409 | L3-9 | 0.725 | NCNRFRRTIVRHCH |
| 410 | L3-10 | 0.769 | AHGKDRRYVEKLEV |
| 411 | L3-11 | 0.8 | FMQKCKKWWDRAVF |
| 412 | L3-12 | 0.882 | DPPKKRKSLLRRVS |
| 413 | L3-13 | 0.862 | YCTRIRREGMKGSE |
| 414 | L3-14 | 0.892 | FPGKSKKQWHRLWP |
| 415 | L3-15 | 0.859 | YEPRFKRPYGKWCH |
| 416 | L3-16 | 0.797 | DYLRSRKMEERFQE |
| 417 | L3-17 | 0.812 | VGPKFRKNHRRQNR |
| 418 | L3-18 | 0.806 | EYQKCKKPSFRLTM |
| 419 | L3-19 | 0.796 | YMKKKRKSLLRTSL |
| 420 | L3-20 | 0.755 | CCFKLKKAYNKGPF |
| 421 | L3-21 | 0.843 | NPMKLRKAETKHNV |
| 422 | L3-22 | 0.83 | HSLKTRKSAFKSNT |
| 423 | L3-23 | 0.799 | QYHKVRKLWFRVEP |
| 424 | L3-24 | 0.846 | FVRKKRKSLLRDTR |
| 425 | L3-25 | 0.807 | CFQRKRKSLLRVLR |
| 426 | L3-26 | 0.802 | VPMKYRRCGNRQSN |
| 427 | L3-27 | 0.83 | RARKKRKSLLRRQV |
| 428 | L3-28 | 0.803 | PNGKVRKRIRRRYF |
| 429 | L3-29 | 0.789 | CASKPRRTYLRAAN |
| 430 | L3-30 | 0.771 | WAHKCKKPGQRIPP |
| 431 | L3-31 | 0.763 | LSDKKRKSLLRYDY |
| 432 | L3-32 | 0.845 | CATRGKKVFSKRTM |
| 433 | L3-33 | 0.751 | LRQRSKRVLEKLRP |
| 434 | L3-34 | 0.74 | LGWKKRKSLLRRHV |
| 435 | L3-35 | 0.747 | LQCKYRRGSDKQPQ |
| 436 | L3-36 | 0.787 | AFSRIKRGVLKLLS |
| 437 | L3-37 | 0.69 | SNVKRKRGRCKPYR |
| 438 | L3-38 | 0.669 | TVVKSRKCSVRYNW |
| 439 | L3-39 | 0.654 | DDVKQRKKHPRVQT |
| 440 | L3-40 | 0.733 | ASPKGRRPTFRPQH |
| 441 | L3-41 | 0.81 | GWLKAKRFPSRPPT |
| 442 | L3-42 | 0.71 | ETNRTRKQCYKTTF |
| 443 | L3-43 | 0.93 | FTHKNRRDSLRVWM |
| 444 | L3-44 | 0.719 | PPFRLRKPLWRPQR |
| 445 | L3-45 | 0.735 | PGNRMKKYQNRVHG |
| 446 | L3-46 | 0.618 | SAHKKRKQTLRCSE |
| 447 | L3-47 | 0.558 | LYEKYKKHNNREDD |
| 448 | L3-48 | 0.622 | PHCRQKKFWIRCGT |
| 449 | L3-49 | 0.629 | VAFKTRRRVQRQSG |
| 450 | L3-50 | 0.634 | VFDKFRKTENRGVI |
| 451 | L3-51 | 0.754 | NHHKTKRCSVRFNI |
| 452 | L3-52 | 0.757 | GTFKWRKSGARQYL |
| 453 | L3-53 | 0.679 | HCLRTKKLINKICS |
| 454 | L3-54 | 0.784 | FFLRCRRLLGKVQV |
| 455 | L3-55 | 0.706 | HFPRARRFEHRCML |
| 456 | L3-56 | 0.703 | QISKLKRPSYRGDD |
| 457 | L3-57 | 0.674 | EEIKQKKLHLRVWF |
| 458 | L3-58 | 0.664 | SLPKWRKGGDRVFT |
| 459 | L3-59 | 0.659 | HVYKNRRVWGKGWP |
| 460 | L3-60 | 0.574 | EDLRCKKLELRSVI |
| 461 | L3-61 | 0.768 | THDKCKKHNDKQAH |
| 462 | L3-62 | 0.766 | YPERPRKLQDKSYS |
| 463 | L3-63 | 0.794 | CPWRNRKAMIKGII |
| 464 | L3-64 | 0.799 | HEIKQKKYFHRGHD |
| 465 | L3-65 | 0.731 | CLEKLRKAVHRQRR |
| 466 | L3-66 | 0.738 | YISKSKKTAGRWFW |
| 467 | L3-67 | 0.786 | VTWKFRKAEKRWGY |
| 468 | L3-68 | 0.792 | YSSKSRKLSPRTPR |
| 469 | L3-69 | 0.733 | CVNRVKKSDSKGTW |
| 470 | L3-70 | 0.781 | DHERERRLWSRFPF |
| 471 | L3-71 | 0.78 | PLIKVKRGVGKLWN |
| 472 | L3-72 | 0.746 | TGCRCKRSMYKNLH |
| 473 | L3-73 | 0.771 | WTCRMRKYQLRTSE |
| 474 | L3-74 | 0.726 | IAIRPRKMTLKIHP |
| 475 | L3-75 | 0.72 | QIPKQKKQEQRAIS |
| 476 | L3-76 | 0.701 | QMGKFKKISLKNTF |
| 477 | L3-77 | 0.688 | VLIKQRKWQDRSCS |
| 478 | L3-78 | 0.712 | FITKSRRQQFRNQG |
| 479 | L3-79 | 0.701 | QQFKYRRECVKYGS |
| 480 | L3-80 | 0.589 | MEGREKKSYNKGEN |
| 481 | L3-81 | 0.775 | PTWRHRRYCAKDIG |
| 482 | L3-82 | 0.783 | HMCRSKRVAWKNLI |
| 483 | L3-83 | 0.768 | SYLKCKRSDYKEVP |
| 484 | L3-84 | 0.739 | HTFKLRKLCQKLFE |
| 485 | L3-85 | 0.735 | QLVKLRKLARRVSY |
| 486 | L3-86 | 0.755 | SEVRPKKDHSRLFI |
| 487 | L3-87 | 0.743 | FLEKCRKFIIRVST |
| 488 | L3-88 | 0.7 | PETRHRKMFIRDFW |
| 489 | L3-89 | 0.624 | EWCKNKRWHSREYP |
| 490 | L3-90 | 0.818 | LWNRYKKLLMRIFW |
| 491 | L3-91 | 0.749 | CWCRQRKCFHKPWI |
| 492 | L3-92 | 0.657 | NLERTKKHGLKGYM |
| 493 | L3-93 | 0.7 | EGGRIKRPNYRGDG |
| 494 | L3-94 | 0.61 | PVLKLRKGRVRAQP |
| 495 | L3-95 | 0.657 | SISRLRKAHQKFIP |
| 496 | L3-96 | 0.567 | EWHKHRREMVRWGP |
| 497 | L3-97 | 0.684 | WSAKGRRMGCKSTM |
| 498 | L3-98 | 0.685 | PNGKVRKRIRRRYF |
| 499 | L3-99 | 0.692 | PCWKFRRAQMKWGI |
| 500 | L3-100 | 0.572 | LEERPKKHCAKNHC |
| 501 | L3-101 | 0.758 | QPWRQKKFWCRCWG |
| 502 | L3-102 | 0.581 | IMVKIRRCPARPLC |
| 503 | L3-103 | 0.676 | NDWKLRKDFWRNHF |
| 504 | L3-104 | 0.57 | ASVRDRKMGYKSDG |
| 505 | L3-105 | 0.601 | YYEKIRRGEIKIAE |
| 506 | L3-106 | 0.642 | DLVRKRKTMLRQLV |
| 507 | L3-107 | 0.658 | DLSKVRRGHGKNDI |
| 508 | L3-108 | 0.621 | ESYRLRRGTDKEQW |
| 509 | L3-109 | 0.613 | YITKFRKFLMKDQF |
| 510 | L3-110 | 0.538 | AWAKYKKVQPKHHT |
| 511 | L3-111 | 0.976 | TWLRGRRWPDKQPQ |
| 512 | L3-112 | 0.731 | ALPRVRKDSHREEI |
| 513 | L3-113 | 0.766 | CLFKYRKSCVRLCG |
| 514 | L3-114 | 0.646 | LQNRMRKIYWKTFD |
| 515 | L3-115 | 0.685 | NDPRLRKHNHRCCT |
| 516 | L3-116 | 0.644 | YGSKQRRFEEKICG |
| 517 | L3-117 | 0.696 | CGHKNKKWHNRMDP |
| 518 | L3-118 | 0.69 | YGIRDRRPMTKHTQ |
| 519 | L3-119 | 0.693 | TPLKSRKYWNKHAY |
| 520 | L3-120 | 0.887 | FLYKAKKALMRDSL |
| 521 | L3-121 | 0.655 | CDDKPKKTVVKLTW |
| 522 | L3-122 | 0.691 | MHPRPRKMSHKMSC |
| 523 | L3-123 | 0.654 | PGERLKKEDHRASG |
| 524 | L3-124 | 0.515 | GAQRARKPVLRAVG |
| 525 | L3-125 | 0.629 | MQEKCRRCVFKGNI |
| 526 | L3-126 | 0.542 | YVDRWRRMQYKLSI |
| 527 | L3-127 | 0.608 | WVDRSRKFEDRNCL |
| 528 | L3-128 | 0.523 | IVNRAKRSQIRFDV |
| 529 | L3-129 | 0.606 | IHPRYKRHQARSCC |
| 530 | L3-130 | 0.531 | EHSKHRKDLFRMVG |
| 531 | L3-131 | 0.655 | YTPKFRRIFWRIID |
| 532 | L3-132 | 0.619 | TITRVKRASHKTPS |
| 533 | L3-133 | 0.566 | VDSREKKWRQKCQC |
| 534 | L3-134 | 0.653 | YWFRIKRAHAKGCP |
| 535 | L3-135 | 0.69 | NDHRMRRSVDRGET |
| 536 | L3-136 | 0.512 | PFDRARRDLHRIMM |
| 537 | L3-137 | 0.565 | QVVRLRRGKNRGTV |
| 538 | L3-138 | 0.576 | TNQRPRRETAKTIE |
| 539 | L3-139 | 0.49 | FHHRHKRIATKHPA |
| 540 | L3-140 | 0.607 | EHCKWKKMFDKEGE |
| 541 | L3-141 | 0.504 | FFYRLRRCLTRWWV |
| 542 | L3-142 | 0.502 | LQCKYRRGSDKTVV |
| 543 | L3-143 | 0.559 | QTMKFKRCCNKEMV |
| 544 | L3-144 | 0.578 | CSYKLRRCDMRLWG |
| 545 | L3-145 | 0.495 | MHIKMKREQVKDEE |
| 546 | L3-146 | 0.512 | AVWRNRKDNVKASE |
| 547 | L3-147 | 0.562 | ADHRAKKTLSKFWT |
| 548 | L3-148 | 0.451 | MMARCRRIMCRSFN |
| 549 | L3-149 | 0.571 | PFGKFKRLENKDEC |
| 550 | L3-150 | 0.518 | YLSRTKRWLARYVE |
| 551 | L3-151 | 0.624 | HTLKPKRLYPRPSE |
| 552 | L3-152 | 0.818 | VVIKCRKWTHKMDH |
| 553 | L3-153 | 0.768 | FHTKLRKPIPKIDM |
| 554 | L3-154 | 0.766 | ENIKSRKYFVKLTW |
| 555 | L3-155 | 0.794 | AWTRFRRFQCKGMS |
| 556 | L3-156 | 0.799 | IVPRDRKQALRWTN |
| 557 | L3-157 | 0.731 | CMLRWKRWHMRFNP |
| 558 | L3-158 | 0.738 | FDMRDRRPNARVMP |
| 559 | L3-159 | 0.786 | QIAKLKRDQMKEAW |
| 560 | L3-160 | 0.792 | LTSRCKKTFQKNSE |
| 561 | L3-161 | 0.733 | MAAKWKKDGMKSHY |
| 562 | L3-162 | 0.781 | ANWRTRRDLAKEHV |
| 563 | L3-163 | 0.78 | PFTKMRKQFQRMSA |
| 564 | L3-164 | 0.746 | QINRVKKSDAKFCT |
| 565 | L3-165 | 0.771 | YLVKLRRFWAKIDM |
| 566 | L3-166 | 0.629 | EYVKAKRTPWKYVV |
| 567 | L3-167 | 0.634 | AMCREKRFPYRAIY |
| 568 | L3-168 | 0.775 | MGLRARKYEDKTLH |
| 569 | L3-169 | 0.783 | FHMKSKKDGDKFVM |
| 570 | L3-170 | 0.768 | CIFKFKKNMFKGYS |
| 571 | L3-171 | 0.629 | LDTKNRRGASKWDY |
| 572 | L3-172 | 0.634 | VTGKGRRVTFRCMN |
| 573 | L3-173 | 0.775 | QLGKQKREATREYV |
| 574 | L3-174 | 0.783 | SSGKWKRPHAKYIV |
| 575 | L3-175 | 0.768 | TCHKTKRSIMRATS |
| 576 | L3-176 | 0.739 | MIGKCRKCGTKCYA |
| 577 | L3-177 | 0.735 | FQPKSRRAETKHSY |
| 578 | L3-178 | 0.755 | PDLKFRKHQDRTAI |
| 579 | L3-179 | 0.743 | YNARVKREGIKNIT |
| 580 | L3-180 | 0.7 | QGIRPRKHLQRMPN |
| 581 | L3-181 | 0.624 | GPCRPRRHSIRLLW |
| 582 | L3-182 | 0.818 | FYLRERRHQLKTEF |
| 583 | L3-183 | 0.768 | NMARSRKNECKYIA |
| 584 | L3-184 | 0.766 | DMDKVKKWVWRSFP |
| 585 | L3-185 | 0.794 | VLPKERRWLTRFLI |
| 586 | L3-186 | 0.799 | YEHRHKRFFHKDMP |
| 587 | L3-187 | 0.731 | HGHKPRRWIYRMPM |
| 588 | L3-188 | 0.738 | DGIRDKKCCWRDLI |
| 589 | L3-189 | 0.786 | ELVKHRRLDFRDPM |
| 590 | L3-190 | 0.792 | VHHKLRRVHLRLDV |
| 591 | L3-191 | 0.739 | GINRGRKVSPKDEQ |
| 592 | L3-192 | 0.735 | NGYRARKENLKFPE |
| 593 | L3-193 | 0.755 | AAYRARKIVEKSGW |
| 594 | L3-194 | 0.743 | WAPRDRRNMGRDPA |
| 595 | L3-195 | 0.7 | IEDKSKRFNCKECG |
| 596 | L3-196 | 0.746 | IEMRYRRDCSRFVN |
| 597 | L3-197 | 0.771 | QQIKYKRPTYRMVC |
| 598 | L3-198 | 0.78 | LPWRERRNGGRSSE |
| 599 | L3-199 | 0.781 | PQTRYRKYFYRPQM |
| 600 | L3-200 | 0.733 | HMCRWRRQINKGVS |
| 에스펩 -2 (con) | 0.303 | WTWKGRKSLLR |
| 서열번호 | 펩타이드 | O.D | Seq. |
| 601 | L2-201 | 0.307 | RRRRRRRRRRR |
| 602 | L2-202 | 0.314 | GGGRKKRRQRR |
| 603 | L2-203 | 0.302 | GGRKKRRQRRR |
| 604 | L3-201 | 0.315 | GGGRKKRRQRRRGG |
| 605 | L3-202 | 0.301 | GGRKKRRQRRRGGG |
| 605 | L2-204 | 0.311 | GGGRRRRRRRR |
| 에스펩-2 (con) | 0.303 | WTWKGRKSLLR |
<실시예 14. 최적화 펩타이드의 아세틸콜린 수용체와의 결합력 확인>
상기 실시예 13-3에서 AchR과의 결합 특이성이 우수한 최적화 펩타이드를 라이브러리별 40개 펩타이드를 선별하여 합성하였고, 실시예 2의 표면 플라스몬 공명(surface plasmon resonance, SPR) 분석 방법을 이용하여 AchR과의 결합력(Ru; Resonance Units)을 비교하였다. 이때, 최적화 펩타이드는 3μM, 10μM의 농도 조건으로 수행하였고, 그 결과를 도 17~19에 나타내었다. 이때, 에스펩-2를 양성대조군으로 이용하였다. 8mer (L1), 11mer (L2), 14mer (L3) 최적화 펩타이드 40개 모두 양성대조군인 에스펩-2보다 AchR과 결합력이 높게 나왔으며, 여기에서 각 라이브러리 마다 AchR과의 결합력이 높은 펩타이드 20개를 도출하였다.
<실시예 15. 각 최적화 펩타이드(L1, L2, L3)의 아세틸콜린 수용체 작용 억제 확인>
상기 실시예 14에서 확인된 AchR에 대한 결합력이 우수한 8mer, 11mer 및 14mer 펩타이드의 AchR 작용 억제 효과를 확인하기 위해, 상기 실시예 3와 동일한 방법으로 AchR 작용 억제 효과를 확인하였다. 이때, 각 펩타이드는 20μM이 되도록 처리하였고, 대조군으로 에스펩-2을 함께 처리하였다. 또한, 최적화 펩타이드의 AchR의 저해능 우수성을 확인하기 위하여 고농도 니코닌을 400μM 또는 600μM을 첨가하였고, 그 결과를 도 20~22에 나타내었다.
도 20~22에서 보듯이, 400μM의 고농도의 니코틴을 첨가하고, 각 펩타이드를 20μM이 되도록 처리한 경우, 대조군인 에스펩-2의 AchR 작용 억제율이 약 10%로 효과가 거의 없는 반면에, 8mer, 11mer, 14mer 펩타이드의 AchR에 대한 작용 억제율은 모두 50% 이상으로 에스펩-2에 비해 억제 효과가 우수함을 확인하였다. 이중 8mer의 L1-13, 11mer의 L2-110, 14mer의 L3-27, 28, 37 펩타이드는 100%에 가까운 억제 효과를 보였다. 나아가, 펩타이드의 세포에서의 AchR의 작용 억제율은 상기 실시예 14의 각 펩타이드의 AchR과의 결합력과 거의 일치한다는 것을 확인하였다.
<실시예 16. 대표 펩타이드의 아세틸콜린 수용체 결합력 확인>
상기 실시예 15에서 확인한 대표 펩타이드 L1-13, L2-110, L3-27, L3-28, L3-37 를, 실시예 2의 표면 플라스몬 공명(surface plasmon resonance, SPR) 분석 방법을 이용하여 AchR과의 결합력(Ru; Resonance Units)을 비교하였다. 이때, 펩타이드는 3μM의 동일한 농도 조건으로 수행하였다. 그 결과를 도 23에 나타내었다.
도 23에서 보듯이 11mer의 L2-110과 14mer의 L3-27, 28, 37이 8mer의 L1-13 보다 AchR에 대한 결합력이 더욱 높았으며, 14mer인 L3-27, 28, 37 펩타이드가 가장 높은 결합력을 보였다. 이를 통해, 본 발명에 따른 화학식에 의한 펩타이드가 8mer, 11mer로 형성될 때에도 AchR과 높은 결합을 나타내며, 14mer일 때 최적의 결합력을 갖는 것을 확인하였다.
<실시예 17. 대표 펩타이드 L2-110, L3-27,28,37 아세틸콜린 수용체 작용 억제 확인>
상기 실시예 16에서 확인된 AchR에 대한 결합력이 우수한 L2-110, L3-27, L3-28, L3-37 펩타이드의 AchR 작용 억제효과를 확인하기 위해 상기 실시예 3과 동일한 방법으로 AchR 작용 억제 효과를 확인하였다. 이때 각 펩타이드는 농도별로 처리하였고 대조군으로 에스펩-2를 함께 처리하였다. 그 결과를 도 24에 나타내었다.
도 24를 보듯이 400 μM 고농도 니코틴에서 에스펩-2는 80μM에서 AchR의 작용을 100% 억제하였으나 70μM의 농도에서는 억제하지 못했다. 11mer인 L2-110과 14mer인 L3-27은 10μM에서 AchR의 작용을 100% 억제하였으며, 5μM은 억제하지 못하였다. 14mer인 L3-28은 5μM에서 AchR의 작용을 100% 억제하였으며, 2.5μM에서는 억제하지 못하였다. 그리고 역시 14mer인 L3-37은 2.5μM에서 100% 억제하였고 1μM에서도 50% 까지 억제하였다.
이 결과를 토대로 도 25에 니코틴성 아세틸콜린 수용체에 대한 저해능을 배수로 나타내었다. 에스펩-2 보다 L2-110과 L3-27이 8배, L3-28이 16배, L3-37이 32배 더 저해효과가 좋았다. 최종적으로 실시예 15의 아세틸콜린 수용체 결합력과 비례하여 억제 효과도 좋게 나온 것을 확인할 수 있었다.
<실시예 18. 대표 펩타이드의 각 말단의 다양한 변형에 따른 아세틸콜린 수용체와의 결합력 확인>
상기 실시예 17에서 확인된 L2-110, L3-27, L3-28, L3-37 펩타이드의 각 말단을 다양하게 변형하여 AchR과의 결합력 변화를 확인하고자 하였다. 구체적으로 상기 실시예 8과 같은 방법으로 지방산 유도체인 팔미토일을 포함하여 myristic acid, stearic acid을 붙이거나 아세틸화 (Acetylation)를 시키거나, 페길화 (PEGylation)시켜서 변형하지 않은 기본 펩타이드와 비교하였다. 그 결과를 도 26~29에 나타내었다.
도 26~29를 보듯이, 본 발명에 따른 펩타이드의 말단에 지방산 유도체를 결합시켰을 때는 모든 펩타이드가 결합력이 약 10배 가까이 증가 하였다(팔미토일-, 미리스틸-, 스테아릭-). 이때 팔미토일을 붙여 변형한 펩타이드가 다른 지방산 유도체를 붙인 것보다 높은 결합력을 보여주는 것을 확인하였다.
본 발명에 따른 펩타이드의 말단을 아세틸화(acetylation)한 경우, AchR과의 결합력은 각 펩타이드의 말단을 변형하지 않았을 때의 AchR과의 결합력과 비슷하였다. 그리고 각 펩타이드의 말단을 페길화(PEGylation)한 경우, 각 펩타이드의 말단을 변형하지 않았을 때에 비해 AchR과의 결합력은 약 50% 감소하였다.
또한 도 30을 보듯이, 본 발명에 따른 펩타이드의 말단에 지방산 유도체인 myristic acid, stearic acid을 붙였을 때 상기 실시예 8과 마찬가지로 팔미토일을 결합한 것처럼 마이셀 구조를 형성함을 확인하였다.
<실시예 19. Palitoyl-L2-110, Palmitoyl-L3-27, Palmitoyl-L3-28, Palmitoyl-L3-37의 아세틸콜린 수용체 작용 억제 확인>
상기 실시예 18에서 AchR에 대한 결합력이 우수한 Palmitoyl-L2-110, Palmitoyl-L3-27, Palmitoyl-L3-28, Palmitoyl-L3-37 펩타이드에 대해 상기 실시예 3과 동일한 방법으로 AchR 작용 억제 효과를 확인하였다. 이때 대조군으로 분가로톡신을 함께 처리하다. 그 결과를 도 31에 나타내었다.
도 31을 보듯이, AchR과의 결합력은 palmitoyl-L3-37이 가장 높았으며, 다음으로 palmitoyl-L3-28이 좋았고, palmitoyl-L3-27과 palmitoyl-L2-110은 비슷한 수준을 나타내었다.
<실시예 20. Palitoyl-L2-110, Palmitoyl-L3-27, Palmitoyl-L3-28, Palmitoyl-L3-37 펩타이드의 세포 독성 확인>
Palitoyl-L2-110, Palmitoyl-L3-27, Palmitoyl-L3-28, Palmitoyl-L3-37 펩타이드의 독성을 평가하기 위해서 TE671 세포에서 WST assay를 실시하였고, 그 결과를 도 32~35에 나타내었다.
도 32~35에서 보듯이, 펩타이드 모두 10ㅅM로 처리하더라도 세포 독성이 나타나지 않음을 확인하였다.
<실시예 21. Palmitoyl-L3-37의 in vivo 효능 분석>
상시 실시예 18에서 아세틸콜린 수용체에 대한 결합력이 가장 좋았던 Palmitoyl-L3-37를 이용하여 동물 효능 평가를 하였다. 6주령 암컷 BALB/c 마우스의 오른쪽 뒷다리 허벅지 근육에(IM) Palmitoyl-L3-37를 10mg/kg 농도로 주사하였다. 그리고 주사하기 전 마우스의 CatWalk 데이터와 주사 후 30분 경과시의 마우스의 CatWalk 데이터를 비교 분석하였고, 그 결과를 도 36에 나타내었다.
도 36에서 보듯이, Palmitoyl-L3-37의 주사 전에 비해 주사 후의 마우스 오른쪽 뒷다리의 발바닥이 바닥에 닿는 면적이 감소한 것을 확인하였고(A), 마우스의 발바닥이 바닥에 닿는 면적(B)과 면적의 최대 값의 평균 값(C)도 Palmitoyl-L3-37 주사 후에 감소하는 것을 확인하였다.
이를 통해, Palmitoyl-L3-37가 아세틸콜린수용체에 결합하여 아세틸콜린의 저해를 통해 동물의 근육 이완에 영향을 준다는 것을 알 수 있었다.
<실시예 22. Palmitoyl-L3-37의 동물 효능(ED50) 및 동물 독성(LD50) 평가>
동물 효능(ED50) 평가는 DAS(Digit Abduction Score) 분석법으로 하였다. DAS 분석법은 마우스 뒷다리 골격근 근육 내(IM) 주사 된 약물의 국소 근육 약화 효능을 측정하기 위해 개발된 방법으로 보톡스도 같은 방법으로 평가한다(Aoki, 1999).
구체적으로, 상기 실시예 18의 Palmitoyl-L3-37에 대한 DAS 값을 평가 하였다. 6주령 암컷 BALB/c 마우스 45마리를 5마리씩 무작위적으로 나누어Palmitoyl-L3-37를 0.1mg/kg, 1mg/kg, 3mg/kg, 10mg/kg, 25mg/kg과 비교군으로 염류 용액(saline)(멸균된 0.9% NaCl) 또는 분가로톡신 70μg/kg을 처리할 그룹으로 구분하였다. 그 다음 Palmitoyl-L3-37와 염류용액, 분가로톡신을 상기와 같은 농도로 마우스의 뒷다리 허벅지 근육에 50μl씩 주사하였다. 이후, 1일 마다 마우스 무게 및 주사한 부분의 발바닥을 관찰하여 기록하였고, 그 결과를 도 37~39에 나타내었다.
도 37의 주사 후 7일간 관찰한 결과, 각 시료 투여 후 10분(0일차)만에 Palmitoyl-L3-37 1mg/kg 투여 그룹은 DAS 2, Palmitoyl-L3-37 3mg/kg, 10mg/kg 및 25mg/kg 투여 그룹은 효능이 DAS 4로 평가되었으며, Palmitoyl-L3-37 1mg/kg 투여 그룹과 비교군인 염류용액 또는 분가로톡신 투여 그룹은 효능이 DAS 0으로 평가되었다. 이를 통해 Palmitoyl-L3-37의 이를 통해 동물실험에서 Palmitoyl-L3-37의 효능(ED50)은 1mg/kg인 것을 알 수 있었다. 또한, 7일 간 관찰하는 동안 Palmitoyl-L3-37 25mg/kg 투여 그룹을 제외한 나머지 그룹은 DAS 값이 내려가면서 회복이 되는 것을 확인하였으나, Palmitoyl-L3-37 25mg/kg투여 그룹은 DAS 4를 유지하는 것으로 나타났다.
도 38의 Palmitoyl-L3-37 25mg/kg 투여 그룹을 투여 후 21일까지 일주일 간격으로 관찰한 결과, Palmitoyl-L3-37 25mg/kg 투여 그룹도 7일 이후에는 DAS값이 점차 감소하여 Palmitoyl-L3-37 투여 19일 이후 DAS값이 1 이하로 내려가면서 회복되는 것을 확인하였다. 이를 통해 Palmitoyl-L3-37의 효능이 가역적이라는 것을 알 수 있었다.
마지막으로 투여 후 7일 동안 1일 마다 무게를 측정하여 Palmitoyl-L3-37의 급성 독성 평가한 결과(도 39 참조), 모든 그룹에서 7일 동안 점차적으로 마우스의 무게가 증가하였고, Palmitoyl-L3-37를 처리한 그룹에서 처리 농도가 가장 높은 25mg/kg을 처리한 그룹에서도 독성이 없다는 것을 확인하였다.
<110> amicogen
<120> Peptide for inhibition of acetylcholine receptor and use thereof
<130> DPC-19-0077
<150> KR 10-2018-0169425
<151> 2018-12-26
<160> 606
<170> KoPatentIn 3.0
<210> 1
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 1
Lys Ala Lys Lys Ile Arg Gln Arg
1 5
<210> 2
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 2
Lys Lys Arg Lys Gly Ser Ala Lys
1 5
<210> 3
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 3
Arg Gly Lys Arg Gln Leu Gly Arg
1 5
<210> 4
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 4
Lys Leu Lys Lys Phe Pro Val Arg
1 5
<210> 5
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 5
Arg His Lys Lys Ser Pro Trp Lys
1 5
<210> 6
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 6
Arg Pro Arg Arg Thr His Ile Lys
1 5
<210> 7
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 7
Lys His Lys Lys Leu Asn Gln Arg
1 5
<210> 8
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 8
Arg Ala Lys Arg Met Cys Cys Lys
1 5
<210> 9
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 9
Arg Gln Arg Arg Asn Asp Met Lys
1 5
<210> 10
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 10
Lys Arg Lys Lys Asp Trp Trp Arg
1 5
<210> 11
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 11
Arg Ala Arg Lys Asp Ala Phe Lys
1 5
<210> 12
<211> 8
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<220>
<223> AchR binding peptide
<400> 12
Lys Trp Arg Arg Glu Gly Phe Lys
1 5
<210> 13
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 13
Arg Phe Arg Arg Gly Val Arg Arg
1 5
<210> 14
<211> 8
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<220>
<223> AchR binding peptide
<400> 14
Lys Ala Arg Arg Gln Glu Asp Lys
1 5
<210> 15
<211> 8
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<220>
<223> AchR binding peptide
<400> 15
Arg Phe Lys Arg Leu Cys Met Lys
1 5
<210> 16
<211> 8
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<223> AchR binding peptide
<400> 16
Lys Leu Lys Arg Arg Ser Arg Arg
1 5
<210> 17
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 17
Arg Pro Arg Arg Ser Leu Asp Arg
1 5
<210> 18
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 18
Lys His Lys Lys Gln Asn Val Arg
1 5
<210> 19
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 19
Lys Phe Lys Arg Thr Glu Gly Arg
1 5
<210> 20
<211> 8
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<220>
<223> AchR binding peptide
<400> 20
Arg Gly Lys Lys Ala Val Arg Lys
1 5
<210> 21
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 21
Lys Phe Lys Lys His Gly Val Lys
1 5
<210> 22
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 22
Lys Thr Lys Lys Ile His Ser Lys
1 5
<210> 23
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 23
Arg Asp Arg Lys Ile His Asp Lys
1 5
<210> 24
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 24
Arg Pro Arg Arg Gly His Gln Lys
1 5
<210> 25
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 25
Lys Thr Lys Lys Leu Tyr Glu Lys
1 5
<210> 26
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 26
Arg Gln Lys Lys Asp Asn Gln Lys
1 5
<210> 27
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 27
Arg Gln Arg Lys Ser Met Gln Arg
1 5
<210> 28
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 28
Arg Gly Arg Arg Trp Phe Val Lys
1 5
<210> 29
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 29
Arg Asn Lys Arg Asp Glu Asn Lys
1 5
<210> 30
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 30
Arg Leu Arg Arg Trp Cys Val Lys
1 5
<210> 31
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 31
Arg Gln Lys Lys Pro Trp Val Lys
1 5
<210> 32
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 32
Arg Trp Lys Arg Ser Glu Gln Arg
1 5
<210> 33
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 33
Arg Leu Lys Arg Ala Pro Gly Arg
1 5
<210> 34
<211> 8
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<220>
<223> AchR binding peptide
<400> 34
Lys Met Arg Arg Ala Gln Gly Arg
1 5
<210> 35
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 35
Lys Ile Lys Lys Ala Ala Arg Arg
1 5
<210> 36
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 36
Lys Asn Arg Lys Asn Leu Tyr Arg
1 5
<210> 37
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 37
Lys Cys Lys Lys Met Thr Glu Arg
1 5
<210> 38
<211> 8
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<220>
<223> AchR binding peptide
<400> 38
Lys Thr Arg Arg Val Asp Leu Arg
1 5
<210> 39
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 39
Lys Ser Arg Lys Trp Gly Trp Arg
1 5
<210> 40
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 40
Lys Arg Arg Arg Phe Arg Trp Lys
1 5
<210> 41
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 41
Lys Tyr Lys Lys Gln His Thr Lys
1 5
<210> 42
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 42
Arg Gly Arg Lys Lys Cys Gly Lys
1 5
<210> 43
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 43
Lys Leu Arg Lys Gln Lys Leu Arg
1 5
<210> 44
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 44
Arg Asn Arg Lys Ile Pro Leu Lys
1 5
<210> 45
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 45
Arg Leu Arg Arg Arg Tyr Arg Arg
1 5
<210> 46
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 46
Lys Trp Arg Lys Arg Arg Ile Lys
1 5
<210> 47
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 47
Lys Thr Lys Arg Met Leu Ala Lys
1 5
<210> 48
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 48
Lys Glu Lys Arg Ala Val His Lys
1 5
<210> 49
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 49
Arg Val Arg Arg Asp Gly His Arg
1 5
<210> 50
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 50
Arg Ala Lys Lys Cys Tyr His Lys
1 5
<210> 51
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 51
Lys Gly Lys Arg Asp Ser Asn Lys
1 5
<210> 52
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 52
Lys Val Arg Lys Ser Leu Trp Lys
1 5
<210> 53
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 53
Arg Ser Lys Lys Ser Leu Tyr Arg
1 5
<210> 54
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 54
Arg Gly Lys Arg Gly Cys Glu Arg
1 5
<210> 55
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 55
Lys Gln Lys Lys Thr Gly Phe Arg
1 5
<210> 56
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 56
Lys Tyr Arg Arg Asn Leu Val Arg
1 5
<210> 57
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 57
Arg Ile Arg Arg Gly Ser Val Arg
1 5
<210> 58
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 58
Arg Ser Lys Arg Met Asn Gly Lys
1 5
<210> 59
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 59
Arg His Lys Lys Phe Ile Glu Lys
1 5
<210> 60
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 60
Arg Ile Arg Arg Val Ser Gln Arg
1 5
<210> 61
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 61
Arg Pro Lys Lys Trp Met Gln Lys
1 5
<210> 62
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 62
Lys Glu Arg Lys His Glu Ala Lys
1 5
<210> 63
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 63
Lys Glu Arg Lys Ala Ser Ala Lys
1 5
<210> 64
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 64
Lys Met Arg Arg Ile Met Tyr Lys
1 5
<210> 65
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 65
Lys Trp Lys Arg Asn Cys Thr Lys
1 5
<210> 66
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 66
Arg Cys Lys Lys Asn Pro Glu Lys
1 5
<210> 67
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 67
Lys Gln Arg Lys Pro His Gly Arg
1 5
<210> 68
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 68
Lys Ala Lys Arg Gly His Phe Lys
1 5
<210> 69
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 69
Lys Ser Lys Arg His His Glu Lys
1 5
<210> 70
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 70
Lys Ser Arg Lys His Cys Asn Arg
1 5
<210> 71
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 71
Arg Ser Lys Lys Ile Asn Asn Lys
1 5
<210> 72
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 72
Arg Val Arg Arg Ser Trp Ile Arg
1 5
<210> 73
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 73
Lys Ser Arg Arg Ser Ala Gly Lys
1 5
<210> 74
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 74
Lys Met Arg Lys His Asp Val Arg
1 5
<210> 75
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 75
Lys Gly Lys Lys Leu Ser Gln Arg
1 5
<210> 76
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 76
Lys Trp Arg Arg Glu Gln Phe Arg
1 5
<210> 77
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 77
Lys Asp Lys Arg Trp Ala Asn Arg
1 5
<210> 78
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 78
Lys His Lys Arg Thr Ala Gln Arg
1 5
<210> 79
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 79
Lys Pro Lys Arg Glu Met His Lys
1 5
<210> 80
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 80
Arg Trp Lys Lys Asn Ile Asn Lys
1 5
<210> 81
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 81
Lys Trp Lys Lys Met Met Glu Arg
1 5
<210> 82
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 82
Arg Ile Arg Arg Trp Trp Val Arg
1 5
<210> 83
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 83
Arg Ile Arg Arg His Met Tyr Arg
1 5
<210> 84
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 84
Lys Pro Lys Lys Pro Pro Ala Arg
1 5
<210> 85
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 85
Arg Leu Lys Arg Gly Gln Phe Lys
1 5
<210> 86
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 86
Lys Asn Lys Arg Thr Pro Val Lys
1 5
<210> 87
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 87
Lys Trp Arg Arg Ala Phe Gly Arg
1 5
<210> 88
<211> 8
<212> PRT
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<220>
<223> AchR binding peptide
<400> 88
Lys Asn Lys Lys Cys Val Gly Arg
1 5
<210> 89
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 89
Arg Thr Arg Lys Cys Val Leu Lys
1 5
<210> 90
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 90
Lys Ile Lys Arg Asp Leu Tyr Lys
1 5
<210> 91
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 91
Arg Glu Lys Arg Tyr Met Ala Arg
1 5
<210> 92
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 92
Arg Val Lys Lys Ala Glu Thr Arg
1 5
<210> 93
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 93
Arg Gly Arg Arg Gly Thr Met Arg
1 5
<210> 94
<211> 8
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<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 94
Arg Thr Lys Arg Trp Gln Leu Lys
1 5
<210> 95
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 95
Arg Leu Arg Lys Ser Leu Gly Lys
1 5
<210> 96
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 96
Arg Pro Lys Arg Asn Trp Ala Lys
1 5
<210> 97
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 97
Lys Asp Arg Lys Val Asn Ser Arg
1 5
<210> 98
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 98
Lys Trp Arg Arg Asn Gly Gln Arg
1 5
<210> 99
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 99
Arg Phe Lys Arg Met Val Trp Arg
1 5
<210> 100
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 100
Lys Glu Arg Lys Gly Ala Val Arg
1 5
<210> 101
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 101
Lys Thr Lys Arg Gly Ser Leu Lys
1 5
<210> 102
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 102
Lys Thr Lys Arg Val Asp Pro Lys
1 5
<210> 103
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 103
Arg Ala Arg Lys Leu His Asn Arg
1 5
<210> 104
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 104
Arg Gly Lys Lys Pro Trp Trp Arg
1 5
<210> 105
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 105
Lys Gly Lys Lys Tyr Tyr Gln Lys
1 5
<210> 106
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 106
Arg Trp Arg Arg Ser Val Ile Lys
1 5
<210> 107
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 107
Arg Gln Lys Lys Asp Phe Leu Lys
1 5
<210> 108
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 108
Arg Gly Arg Lys Pro Ile Trp Lys
1 5
<210> 109
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 109
Lys Pro Arg Arg Pro Asp Thr Arg
1 5
<210> 110
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 110
Lys Val Lys Arg Leu Trp Asn Arg
1 5
<210> 111
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 111
Lys Asp Arg Lys Gly Val Tyr Lys
1 5
<210> 112
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 112
Arg Val Lys Arg Phe Val Pro Lys
1 5
<210> 113
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 113
Lys Pro Lys Arg Asp Gln Ser Arg
1 5
<210> 114
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 114
Lys Asp Lys Lys Gln Trp Gly Arg
1 5
<210> 115
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 115
Arg Phe Arg Lys Met Ile Pro Lys
1 5
<210> 116
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 116
Lys Phe Arg Lys His Val Cys Lys
1 5
<210> 117
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 117
Arg Pro Lys Arg Ser Pro Ser Lys
1 5
<210> 118
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 118
Arg Tyr Lys Lys Val Pro Ala Lys
1 5
<210> 119
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 119
Arg Ser Arg Arg Cys Pro Val Lys
1 5
<210> 120
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 120
Arg Cys Lys Arg Cys Asp Asn Lys
1 5
<210> 121
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 121
Arg Gly Lys Lys Gln Leu Ser Lys
1 5
<210> 122
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 122
Lys Ser Lys Arg Pro Glu Gly Arg
1 5
<210> 123
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 123
Lys Cys Arg Lys Thr Met Gly Arg
1 5
<210> 124
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 124
Arg Asp Arg Lys Asn Pro Val Arg
1 5
<210> 125
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 125
Lys Pro Lys Arg Ser Ala Pro Arg
1 5
<210> 126
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 126
Lys Gly Lys Arg Thr Gly Cys Lys
1 5
<210> 127
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 127
Arg Phe Arg Lys Pro Thr Asp Arg
1 5
<210> 128
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 128
Arg Met Lys Lys Phe His Thr Arg
1 5
<210> 129
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 129
Lys Ile Lys Lys Tyr Tyr Trp Lys
1 5
<210> 130
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 130
Arg Ile Lys Arg Asn Asn Cys Lys
1 5
<210> 131
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 131
Arg His Lys Arg Met Leu Trp Arg
1 5
<210> 132
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 132
Lys Val Arg Arg Val Phe Leu Lys
1 5
<210> 133
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 133
Lys Asp Lys Arg Pro Glu Cys Lys
1 5
<210> 134
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 134
Lys Thr Arg Arg Ser Ala Cys Arg
1 5
<210> 135
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 135
Lys Phe Arg Lys Gly Pro Tyr Arg
1 5
<210> 136
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 136
Lys Phe Lys Lys Ser Ala Pro Arg
1 5
<210> 137
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 137
Lys Pro Arg Lys Ile Gln Gly Arg
1 5
<210> 138
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 138
Lys Met Arg Lys Gly Trp Asp Lys
1 5
<210> 139
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 139
Lys Phe Arg Arg Gln Ser Thr Arg
1 5
<210> 140
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 140
Lys Asn Arg Arg Ala Asp Cys Arg
1 5
<210> 141
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 141
Lys Asn Lys Arg Trp Val Phe Lys
1 5
<210> 142
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 142
Arg Cys Arg Lys Asp Thr Ala Arg
1 5
<210> 143
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 143
Lys Ala Arg Arg Val Gly Thr Lys
1 5
<210> 144
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 144
Lys Val Lys Lys Ile Tyr Tyr Arg
1 5
<210> 145
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 145
Lys Ala Lys Arg Asp Tyr Leu Arg
1 5
<210> 146
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 146
Lys Val Lys Arg Leu Pro Tyr Arg
1 5
<210> 147
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 147
Arg His Arg Arg Ala Asn Phe Arg
1 5
<210> 148
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 148
Lys Pro Lys Lys Gly Gly Trp Lys
1 5
<210> 149
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 149
Lys Glu Lys Arg Leu Tyr Ala Arg
1 5
<210> 150
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 150
Arg Glu Lys Lys Gln Glu Val Lys
1 5
<210> 151
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 151
Lys Ser Arg Arg Ala Val Phe Arg
1 5
<210> 152
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 152
Arg His Arg Lys Tyr Val Asp Lys
1 5
<210> 153
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 153
Arg His Lys Arg Trp Ser Gly Arg
1 5
<210> 154
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 154
Arg Asp Arg Lys Phe Val Gln Lys
1 5
<210> 155
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 155
Arg Tyr Arg Arg Trp Phe Tyr Arg
1 5
<210> 156
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 156
Arg Gln Arg Arg Met Thr Pro Arg
1 5
<210> 157
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 157
Arg Asn Lys Arg Met Asp Gly Lys
1 5
<210> 158
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 158
Arg Pro Lys Lys His Gln Glu Arg
1 5
<210> 159
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 159
Lys Lys Arg Lys Ser Leu Leu Arg
1 5
<210> 160
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 160
Lys Ile Lys Arg Asn Val Phe Lys
1 5
<210> 161
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 161
Lys Tyr Lys Arg Glu Glu Tyr Arg
1 5
<210> 162
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 162
Lys Ile Arg Arg Val Thr Asn Lys
1 5
<210> 163
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 163
Lys Val Lys Arg Val Trp Gly Arg
1 5
<210> 164
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 164
Arg Ile Lys Arg Asp Tyr Cys Lys
1 5
<210> 165
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 165
Arg Ile Arg Arg Ala His Asp Arg
1 5
<210> 166
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 166
Arg Ala Arg Lys Glu Ser His Lys
1 5
<210> 167
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 167
Arg Tyr Lys Lys Gln Gln Tyr Lys
1 5
<210> 168
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 168
Lys Ile Lys Lys Leu Ser Gln Lys
1 5
<210> 169
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 169
Lys Leu Lys Arg Ala Met Leu Lys
1 5
<210> 170
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 170
Lys Ala Arg Lys Asn Ser Ile Arg
1 5
<210> 171
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 171
Lys Glu Arg Lys Leu Trp Trp Lys
1 5
<210> 172
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 172
Lys Thr Arg Lys Gln Asp His Arg
1 5
<210> 173
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 173
Arg Cys Lys Arg Phe Val Gly Lys
1 5
<210> 174
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 174
Lys Glu Lys Lys Leu Val Trp Lys
1 5
<210> 175
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 175
Lys Ala Arg Arg Asn Ser Leu Lys
1 5
<210> 176
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 176
Lys Met Arg Arg Val Ala Pro Arg
1 5
<210> 177
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 177
Arg Ala Lys Lys Ile Met Phe Arg
1 5
<210> 178
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 178
Lys Thr Lys Lys Ala Ala Glu Arg
1 5
<210> 179
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 179
Arg Gly Lys Arg His Trp His Arg
1 5
<210> 180
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 180
Arg Thr Lys Lys Glu Asn Val Lys
1 5
<210> 181
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 181
Arg Glu Lys Lys Tyr Ala Tyr Lys
1 5
<210> 182
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 182
Lys Trp Arg Arg Glu Leu Pro Arg
1 5
<210> 183
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 183
Lys Ser Arg Arg Met Trp Gly Arg
1 5
<210> 184
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 184
Lys Cys Lys Lys Asp Asn Asp Lys
1 5
<210> 185
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 185
Lys Asp Lys Lys Met Pro Gln Arg
1 5
<210> 186
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 186
Arg His Lys Arg Gln Gln Asp Lys
1 5
<210> 187
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 187
Arg Leu Lys Lys His Cys Gly Lys
1 5
<210> 188
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 188
Lys Ala Arg Lys Gln Glu Val Arg
1 5
<210> 189
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 189
Lys Met Arg Lys Phe Tyr Ser Lys
1 5
<210> 190
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 190
Lys Met Arg Arg Trp Trp Leu Lys
1 5
<210> 191
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 191
Lys Gln Lys Arg Gln Trp Ala Arg
1 5
<210> 192
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 192
Arg Ser Arg Arg Gly Ile Gly Lys
1 5
<210> 193
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 193
Lys Asp Lys Lys Thr Pro Cys Lys
1 5
<210> 194
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 194
Arg Ile Arg Lys Ile Thr Trp Arg
1 5
<210> 195
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 195
Arg His Lys Arg Trp Glu Val Arg
1 5
<210> 196
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 196
Arg Phe Arg Arg Asn Phe His Lys
1 5
<210> 197
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 197
Lys Trp Lys Arg Phe Ser Gln Arg
1 5
<210> 198
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 198
Lys Tyr Lys Lys Ser Phe Thr Lys
1 5
<210> 199
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 199
Lys Thr Arg Lys Ile Val Met Lys
1 5
<210> 200
<211> 8
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 200
Arg Thr Lys Lys Ala Tyr Val Lys
1 5
<210> 201
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 201
Glu Asp Tyr Lys Tyr Arg Arg Gln Asn Tyr Lys
1 5 10
<210> 202
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 202
Gly Ile Trp Lys Phe Arg Arg Asn Gln Cys Lys
1 5 10
<210> 203
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 203
Ser Pro Val Arg Trp Lys Arg Leu Cys Leu Arg
1 5 10
<210> 204
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 204
Glu Cys Tyr Arg Asn Arg Lys Ala Tyr Cys Arg
1 5 10
<210> 205
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 205
Thr Gln Gln Arg Leu Lys Arg Val Met Glu Lys
1 5 10
<210> 206
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 206
Pro Glu Leu Lys Cys Lys Arg Met Ile Gly Arg
1 5 10
<210> 207
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 207
Trp Gln Met Lys Thr Lys Lys Glu Ile Trp Lys
1 5 10
<210> 208
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 208
Phe Gly His Arg Gly Lys Lys Glu Trp Ala Arg
1 5 10
<210> 209
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 209
Ala Thr Cys Lys Pro Lys Arg Pro Trp Tyr Lys
1 5 10
<210> 210
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 210
Thr Ser Gln Lys Val Arg Lys Met Glu Ala Lys
1 5 10
<210> 211
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 211
Gly Ile Pro Lys Tyr Arg Arg Gly Cys Ser Arg
1 5 10
<210> 212
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 212
Ala Tyr Ala Lys Ala Arg Arg Trp Gly Gln Lys
1 5 10
<210> 213
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 213
Gly Gly Leu Lys Ser Lys Arg Leu Thr Thr Arg
1 5 10
<210> 214
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 214
Gly Met Trp Lys Val Arg Lys Thr Val Phe Arg
1 5 10
<210> 215
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 215
His Thr Thr Arg Gly Lys Arg Cys Asp Pro Arg
1 5 10
<210> 216
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 216
Val Asp Val Arg Asn Lys Lys Ser Asn Asn Arg
1 5 10
<210> 217
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 217
Gly Leu Asn Arg Trp Arg Arg Cys His His Lys
1 5 10
<210> 218
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 218
Leu Gln Ser Arg Ser Lys Lys Tyr Ser Val Lys
1 5 10
<210> 219
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 219
Gly Trp Asn Arg Ala Lys Arg Glu Glu Ser Lys
1 5 10
<210> 220
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 220
Ser Trp Gln Lys Cys Lys Lys Ala Glu Val Arg
1 5 10
<210> 221
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 221
Met Val Met Lys Trp Lys Arg Trp Asn Gln Arg
1 5 10
<210> 222
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 222
Met Met Met Arg His Lys Arg Cys Gln Tyr Arg
1 5 10
<210> 223
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 223
Gln Thr Tyr Arg Leu Lys Lys Pro Leu Gln Lys
1 5 10
<210> 224
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 224
Ser Phe Trp Arg Glu Arg Lys Asn Gly Phe Arg
1 5 10
<210> 225
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 225
Arg Leu Glu Arg Pro Arg Arg Glu Leu Thr Lys
1 5 10
<210> 226
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 226
Ala Pro Trp Arg Leu Lys Arg Ala Pro Gln Arg
1 5 10
<210> 227
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 227
Gln Gly Glu Lys Tyr Arg Arg Thr Glu Ser Lys
1 5 10
<210> 228
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 228
Cys Ile Trp Lys Ser Lys Arg Ser Pro Ala Lys
1 5 10
<210> 229
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 229
Ala Ile Leu Lys Gly Lys Arg Ile Pro Asn Lys
1 5 10
<210> 230
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 230
Pro Thr Leu Lys Trp Arg Lys Pro Val Leu Arg
1 5 10
<210> 231
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 231
Asn Asn Ser Arg Ser Lys Arg Phe Met Asn Arg
1 5 10
<210> 232
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 232
Leu Tyr Met Arg Cys Lys Lys Gln Ala Pro Arg
1 5 10
<210> 233
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 233
Ile Asn Tyr Arg Cys Arg Lys Trp Val Asp Lys
1 5 10
<210> 234
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 234
Val Met Asp Arg Asp Arg Lys Trp Trp Trp Arg
1 5 10
<210> 235
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 235
Asn Phe Thr Lys Leu Arg Lys Pro Gly Pro Arg
1 5 10
<210> 236
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 236
Phe Ile Pro Lys Met Arg Lys Cys Ser Pro Arg
1 5 10
<210> 237
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 237
Thr Leu Trp Lys Val Arg Lys Ala Tyr Ser Arg
1 5 10
<210> 238
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 238
Glu Ser Glu Lys Ile Lys Arg Leu Ser Thr Arg
1 5 10
<210> 239
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 239
Leu Met Thr Lys Asn Arg Arg Asn Ser Phe Arg
1 5 10
<210> 240
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 240
Pro Glu Val Lys Val Arg Arg His Ser Met Arg
1 5 10
<210> 241
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 241
Ala Glu Val Lys Asp Lys Lys Ala Tyr Tyr Lys
1 5 10
<210> 242
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 242
Tyr Tyr Ala Lys Ser Lys Lys Tyr Met Val Arg
1 5 10
<210> 243
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 243
Leu Met Asp Arg Thr Arg Arg Asp Met Tyr Lys
1 5 10
<210> 244
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 244
Asn Met Gly Arg His Lys Arg Pro Phe Leu Lys
1 5 10
<210> 245
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 245
Trp Ile Tyr Arg Gly Arg Lys Asp Val Ala Lys
1 5 10
<210> 246
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 246
Cys Tyr Trp Lys Ala Lys Arg Tyr Pro Met Arg
1 5 10
<210> 247
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 247
Ile Glu Asp Lys Gly Arg Arg Ile Asn Pro Arg
1 5 10
<210> 248
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 248
Val Ser Thr Lys Ile Lys Lys Glu Pro Gln Arg
1 5 10
<210> 249
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 249
Tyr Pro Phe Lys Ala Lys Arg Pro Ala Glu Lys
1 5 10
<210> 250
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 250
Ser Ala Asp Arg Asn Lys Arg His Met Thr Arg
1 5 10
<210> 251
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 251
Ser Leu Tyr Arg Gln Lys Arg His Asp Tyr Lys
1 5 10
<210> 252
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 252
Leu Pro Ser Arg Pro Lys Lys Pro Val Pro Lys
1 5 10
<210> 253
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 253
Pro Ala Trp Arg Cys Lys Arg Cys Gln Pro Lys
1 5 10
<210> 254
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 254
His His Trp Arg Phe Lys Arg Glu Met Pro Arg
1 5 10
<210> 255
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 255
Pro Ala Cys Arg Ser Lys Arg Asp Trp Gln Lys
1 5 10
<210> 256
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 256
Phe Ile Cys Lys Ser Arg Lys Phe Tyr Gly Lys
1 5 10
<210> 257
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 257
Leu Glu His Lys Glu Arg Lys Asp Asp Phe Arg
1 5 10
<210> 258
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 258
Ala Asn Pro Lys Asn Arg Lys Asp Asn Leu Arg
1 5 10
<210> 259
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 259
Phe Gly Val Lys Tyr Arg Arg Val Ile Cys Arg
1 5 10
<210> 260
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 260
His Ala Asp Lys Phe Arg Arg Phe Asn Met Arg
1 5 10
<210> 261
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 261
Pro Thr Ile Arg Val Arg Lys Ser Asp Asp Arg
1 5 10
<210> 262
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 262
Asn Leu His Lys Pro Lys Arg Asp Leu Pro Lys
1 5 10
<210> 263
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 263
Ile Asp Gly Lys Trp Arg Lys Ile Cys Thr Arg
1 5 10
<210> 264
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 264
Ser Pro Phe Arg Ala Lys Arg Gln Asp Val Arg
1 5 10
<210> 265
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 265
Pro Leu Trp Lys Thr Arg Lys Ile Glu Pro Arg
1 5 10
<210> 266
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 266
Pro Asn Tyr Lys Trp Arg Lys Ser Arg Arg Arg
1 5 10
<210> 267
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 267
Lys Thr Gly Arg Ser Lys Arg His Arg Trp Arg
1 5 10
<210> 268
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 268
Gly Pro Asp Lys Ser Arg Arg Asn Leu His Arg
1 5 10
<210> 269
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 269
Trp Cys Cys Lys Thr Lys Arg Ala Val Met Lys
1 5 10
<210> 270
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 270
Gln Leu Leu Lys Met Arg Lys Ala Leu Ser Arg
1 5 10
<210> 271
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 271
Pro Tyr Leu Arg Ser Lys Arg Phe Pro Pro Arg
1 5 10
<210> 272
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 272
Ala Pro His Lys Trp Arg Lys Gln Glu Gln Arg
1 5 10
<210> 273
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 273
Pro Pro Phe Lys Phe Arg Lys Pro Leu Gly Arg
1 5 10
<210> 274
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 274
Cys Ile Met Arg Val Lys Arg Met Trp Trp Lys
1 5 10
<210> 275
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 275
Tyr Ile Leu Arg Asp Lys Lys Val Pro Leu Lys
1 5 10
<210> 276
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 276
Pro Trp Val Arg Ile Arg Lys Met Ala Ser Arg
1 5 10
<210> 277
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 277
Val Ala Asp Arg Gln Lys Lys Thr Ala Pro Lys
1 5 10
<210> 278
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 278
His His Asn Arg Met Lys Lys Gln Tyr His Arg
1 5 10
<210> 279
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 279
Glu Gly Ala Lys Tyr Arg Arg Asp Gly Trp Arg
1 5 10
<210> 280
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 280
Phe Trp Asp Arg Val Lys Arg Asn Pro Ser Lys
1 5 10
<210> 281
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 281
Thr Met Trp Arg Gln Arg Lys Met Ser Cys Lys
1 5 10
<210> 282
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 282
Pro Asp Thr Arg Trp Lys Arg Val Leu Phe Lys
1 5 10
<210> 283
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 283
His Gln Gln Lys Cys Lys Lys Thr Thr Thr Lys
1 5 10
<210> 284
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 284
Pro Trp His Lys Gly Lys Arg Asp Phe Asp Arg
1 5 10
<210> 285
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 285
Val Phe Val Lys Trp Arg Arg Gln Met Met Arg
1 5 10
<210> 286
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 286
Ser Gln Trp Lys Ile Arg Lys Arg Leu Ile Arg
1 5 10
<210> 287
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 287
Glu Trp His Lys Val Arg Arg Val Tyr Ala Lys
1 5 10
<210> 288
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 288
Asn Ala Met Arg Thr Lys Arg Met Ser Phe Lys
1 5 10
<210> 289
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 289
Pro Pro Phe Lys Phe Arg Lys Pro Leu Gly Arg
1 5 10
<210> 290
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 290
Thr Ser Val Lys Lys Arg Lys Gln Arg Leu Arg
1 5 10
<210> 291
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 291
Pro Ile Asp Lys Phe Lys Arg Gly Met Val Arg
1 5 10
<210> 292
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 292
Phe Val Gly Arg Trp Lys Arg Glu Tyr Ala Lys
1 5 10
<210> 293
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 293
Phe Asn Pro Lys Met Arg Lys Val Cys Leu Arg
1 5 10
<210> 294
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 294
Asn Pro Gly Lys Thr Lys Arg Trp Leu Gln Arg
1 5 10
<210> 295
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 295
Ile Met Asp Lys Arg Arg Lys Pro Gly Cys Arg
1 5 10
<210> 296
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 296
Ile Ser Asp Lys Ala Lys Arg Gln His Phe Lys
1 5 10
<210> 297
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 297
Asp Val Asn Lys Tyr Arg Lys His Ser His Lys
1 5 10
<210> 298
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 298
Leu Met Leu Arg Gly Lys Arg Leu Thr Thr Lys
1 5 10
<210> 299
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 299
Cys Ser Leu Arg Ala Arg Lys Glu Glu Trp Arg
1 5 10
<210> 300
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 300
Met Asn Tyr Arg Cys Lys Arg Val Gln Glu Arg
1 5 10
<210> 301
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 301
Asn Gly Val Arg Glu Arg Lys Trp Gln Ser Lys
1 5 10
<210> 302
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 302
Tyr Thr Ser Arg Cys Lys Lys Gln Pro Arg Lys
1 5 10
<210> 303
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 303
Gln Tyr Asn Lys Gly Arg Lys Ile His Val Arg
1 5 10
<210> 304
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 304
Pro Glu Asp Arg Gln Arg Lys Thr Trp Phe Lys
1 5 10
<210> 305
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 305
Ile Glu Met Lys Pro Arg Lys Phe Gly Val Lys
1 5 10
<210> 306
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 306
Gln Arg Trp Lys Trp Arg Lys Ser Leu Ala Arg
1 5 10
<210> 307
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 307
Val Lys His Lys Glu Arg Lys Cys Gln Arg Arg
1 5 10
<210> 308
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 308
Gln Gln Asp Arg Pro Lys Arg Asp Ile Pro Lys
1 5 10
<210> 309
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 309
Tyr Pro Thr Lys Gly Lys Arg Cys Met Ile Arg
1 5 10
<210> 310
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 310
Arg Tyr Ala Lys His Arg Lys Arg Gln Thr Arg
1 5 10
<210> 311
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 311
Gly Tyr Phe Arg Pro Arg Lys Glu Thr Cys Lys
1 5 10
<210> 312
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 312
Cys Phe Phe Lys Met Arg Arg Cys Asn Thr Lys
1 5 10
<210> 313
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 313
Gln Asn Asp Lys Asp Arg Lys Leu Ser His Arg
1 5 10
<210> 314
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 314
Gln Val Thr Lys Ser Lys Arg Val Ala Phe Lys
1 5 10
<210> 315
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 315
Val Arg Ala Lys His Arg Lys Ser Ser Leu Arg
1 5 10
<210> 316
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 316
Ser His Ser Arg Gln Arg Lys Thr Pro Leu Arg
1 5 10
<210> 317
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 317
Asp Asn Gly Lys Ile Arg Arg Cys Leu Gly Lys
1 5 10
<210> 318
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 318
Phe Leu Arg Arg Leu Lys Lys Val His Trp Lys
1 5 10
<210> 319
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 319
Phe Cys Arg Arg Ser Lys Lys Ile Gly Arg Arg
1 5 10
<210> 320
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 320
Pro Ala Ala Arg Thr Lys Arg Met Tyr Gly Arg
1 5 10
<210> 321
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 321
His Glu Thr Lys Pro Lys Lys Asp Gly Leu Arg
1 5 10
<210> 322
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 322
Met Gly Asp Arg Pro Arg Lys Trp Asp Ser Arg
1 5 10
<210> 323
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 323
His Asp Thr Lys Cys Lys Lys Met Tyr Ala Lys
1 5 10
<210> 324
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 324
Asn Val Val Arg Gly Arg Arg Leu Glu Cys Arg
1 5 10
<210> 325
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 325
Pro Ala Asp Lys Gly Arg Arg Glu Val Met Lys
1 5 10
<210> 326
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 326
Ala Met Met Lys Tyr Lys Lys Glu Phe Pro Lys
1 5 10
<210> 327
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 327
Tyr Ile Ile Arg Trp Lys Arg Gln Met Thr Arg
1 5 10
<210> 328
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 328
Ser Pro Trp Arg Ile Arg Arg Gln Asn Ile Arg
1 5 10
<210> 329
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 329
Thr Cys Ile Lys Tyr Arg Arg Ala His Thr Lys
1 5 10
<210> 330
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 330
Gly Tyr Asp Arg Ala Arg Lys Gly Thr Leu Arg
1 5 10
<210> 331
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 331
Met Thr Leu Lys His Arg Arg Val Tyr Ile Lys
1 5 10
<210> 332
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 332
Leu Phe Thr Arg Ile Lys Arg Leu Val Cys Lys
1 5 10
<210> 333
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 333
Asp Phe Ser Arg Asp Arg Arg Cys Leu Ser Lys
1 5 10
<210> 334
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 334
Val Trp Asn Lys Val Lys Arg Trp Leu Glu Arg
1 5 10
<210> 335
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 335
Phe Pro Gly Lys Asn Arg Lys Tyr Cys Ser Arg
1 5 10
<210> 336
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 336
Tyr Thr Ser Lys Asn Lys Arg Gly Cys Pro Arg
1 5 10
<210> 337
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 337
Cys Ala Cys Lys Gln Arg Arg Ala Thr Ser Arg
1 5 10
<210> 338
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 338
Ile Met Glu Arg Gln Arg Lys Ser Gln His Arg
1 5 10
<210> 339
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 339
Ser Val Leu Arg Cys Arg Lys Cys Ser Met Lys
1 5 10
<210> 340
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 340
Tyr Pro Gln Lys Leu Arg Arg Thr Ala Leu Lys
1 5 10
<210> 341
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 341
Ser Ser His Lys Gly Lys Arg Ala Gln Ser Lys
1 5 10
<210> 342
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 342
Asp Asn Pro Arg Phe Arg Lys Thr Ile Leu Lys
1 5 10
<210> 343
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 343
Ala Ile Ile Lys Phe Arg Lys Val Gln Trp Lys
1 5 10
<210> 344
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 344
Pro Tyr Thr Arg Cys Arg Lys Glu Ile Cys Lys
1 5 10
<210> 345
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 345
Met Asn Glu Lys Pro Lys Lys Asn Asp Gln Lys
1 5 10
<210> 346
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 346
Leu Ile Gly Lys Phe Lys Lys Pro Phe Tyr Arg
1 5 10
<210> 347
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 347
Thr Trp Cys Lys His Lys Lys Leu Asp Met Lys
1 5 10
<210> 348
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 348
Asp Ser Asn Lys Val Arg Lys Cys Ser Ser Lys
1 5 10
<210> 349
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 349
Leu Pro Met Lys Gln Arg Lys Cys Glu Phe Arg
1 5 10
<210> 350
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 350
Glu Asp Val Arg Val Lys Arg Gln Thr Cys Arg
1 5 10
<210> 351
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 351
Thr Gln Cys Lys Asp Arg Arg Val Ser Asp Arg
1 5 10
<210> 352
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 352
Ile Ala Leu Lys Pro Lys Arg Val Trp Leu Lys
1 5 10
<210> 353
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 353
Gly His Gln Arg Gly Lys Arg Glu Gly Ser Arg
1 5 10
<210> 354
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 354
Asn Pro Phe Lys Tyr Lys Lys Ile Cys Pro Lys
1 5 10
<210> 355
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 355
Asn Glu Ala Arg Ile Lys Lys Cys Asp Val Lys
1 5 10
<210> 356
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 356
Tyr Gly Leu Arg Met Arg Lys Trp Tyr Met Lys
1 5 10
<210> 357
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 357
Asn Phe Tyr Lys Cys Arg Lys Leu Gln Cys Lys
1 5 10
<210> 358
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 358
Met Met Thr Lys Tyr Lys Lys Thr Cys Cys Lys
1 5 10
<210> 359
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 359
Cys Asn Gln Lys Thr Lys Lys Ile Ala Glu Lys
1 5 10
<210> 360
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 360
Ala Asp Ile Arg Met Lys Lys Trp Tyr Pro Lys
1 5 10
<210> 361
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 361
Ala Trp Phe Arg Val Lys Arg Ser Asn Cys Arg
1 5 10
<210> 362
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 362
Asn Cys Asp Arg Thr Arg Lys His Trp Ala Arg
1 5 10
<210> 363
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 363
Pro His Ala Arg Thr Arg Lys Asn Ile Thr Lys
1 5 10
<210> 364
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 364
Val Pro Thr Lys Met Lys Lys Tyr Glu Thr Lys
1 5 10
<210> 365
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 365
Ala Tyr Pro Lys Phe Arg Lys Thr Phe Asn Arg
1 5 10
<210> 366
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 366
Gln Gln Leu Arg Leu Arg Lys Leu Cys Gly Lys
1 5 10
<210> 367
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 367
Met Phe Met Arg Asn Lys Lys Leu Ala Trp Arg
1 5 10
<210> 368
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 368
Gly Gly Ala Lys Asn Lys Lys Val Val Ser Arg
1 5 10
<210> 369
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 369
His Asp Pro Arg His Lys Lys Thr Pro Thr Lys
1 5 10
<210> 370
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 370
Gln Val His Arg Asn Lys Arg Tyr Thr Asp Arg
1 5 10
<210> 371
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 371
Tyr Gly Thr Arg Pro Lys Lys Tyr Val Ser Lys
1 5 10
<210> 372
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 372
Cys Thr Trp Arg Gly Arg Arg Pro His Asp Lys
1 5 10
<210> 373
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 373
Ser Trp Ala Lys Ala Arg Lys Leu Val His Arg
1 5 10
<210> 374
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 374
Pro Leu Phe Lys Ser Arg Arg Ala Tyr Val Arg
1 5 10
<210> 375
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 375
Cys Val Met Arg Cys Arg Arg Ser Glu Asp Lys
1 5 10
<210> 376
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 376
Trp Trp His Lys His Arg Arg Ala Gln Ser Lys
1 5 10
<210> 377
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 377
Leu Asn Ser Lys Pro Arg Arg Val Glu Phe Lys
1 5 10
<210> 378
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 378
Val Pro Tyr Arg His Arg Arg Met Gln Phe Lys
1 5 10
<210> 379
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 379
Gly Ile Ala Lys Ser Lys Arg Asn Ala Gly Arg
1 5 10
<210> 380
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 380
Asp Pro Glu Lys Trp Arg Lys Phe Tyr Asp Arg
1 5 10
<210> 381
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 381
Pro Gly Ala Arg Cys Arg Lys Gln Asp Val Lys
1 5 10
<210> 382
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 382
His Glu Gln Arg Pro Lys Lys Gly Gln Gln Lys
1 5 10
<210> 383
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 383
Thr Cys Asp Arg Ala Arg Lys Glu Ser Phe Arg
1 5 10
<210> 384
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 384
Met His Gln Arg Glu Arg Arg Asn Phe Val Lys
1 5 10
<210> 385
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 385
Phe Ile Thr Arg Phe Arg Lys Met Gly Glu Lys
1 5 10
<210> 386
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 386
Pro Asn Trp Lys Val Arg Arg Phe Gly Asp Lys
1 5 10
<210> 387
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 387
Thr Ala Ala Lys Trp Lys Lys Ile Ile Met Lys
1 5 10
<210> 388
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 388
Cys Leu Trp Arg Leu Arg Lys Asp Asn Gly Arg
1 5 10
<210> 389
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 389
Ala His Ile Lys Ser Arg Lys Val Trp Ser Arg
1 5 10
<210> 390
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 390
Asn Leu Val Lys Ser Lys Lys Val Glu Glu Lys
1 5 10
<210> 391
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 391
Pro Asp Ser Arg Leu Lys Lys His Glu Ala Lys
1 5 10
<210> 392
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 392
Glu His Val Lys Leu Lys Arg Leu Asp Phe Arg
1 5 10
<210> 393
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 393
Pro Ala Leu Arg Met Arg Arg Trp Cys Gln Lys
1 5 10
<210> 394
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 394
His Leu Glu Lys His Arg Arg Cys Glu Phe Lys
1 5 10
<210> 395
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 395
Cys Gln Ala Lys Thr Arg Lys Ala Glu Asp Arg
1 5 10
<210> 396
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 396
Gln Asn Met Arg Met Lys Arg Phe Ile Gln Arg
1 5 10
<210> 397
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 397
Cys Pro Tyr Arg Ile Arg Arg Gly Pro Gly Lys
1 5 10
<210> 398
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 398
Asp Ile Tyr Lys Gly Lys Arg Thr Leu Val Lys
1 5 10
<210> 399
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 399
Glu Ile Cys Lys Asn Arg Lys Pro Ala Asn Arg
1 5 10
<210> 400
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 400
Gln Gly Met Lys Leu Lys Arg Ile Trp Ser Lys
1 5 10
<210> 401
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 401
Ala Leu Gly Arg Thr Lys Arg Leu His Met Arg Val His Val
1 5 10
<210> 402
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 402
Ser Phe Tyr Lys Glu Arg Lys Cys Gln Phe Arg Ser Gly Leu
1 5 10
<210> 403
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 403
Ser Gly Val Arg Tyr Arg Lys Trp Trp Ile Arg Ser Val Val
1 5 10
<210> 404
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 404
Met Pro Gln Lys Leu Lys Lys Leu Asp Ile Arg Asn His Asn
1 5 10
<210> 405
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 405
Asn Glu Asp Arg Gly Lys Arg Pro His Ile Arg Val Leu Gly
1 5 10
<210> 406
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 406
Glu Phe Val Lys Leu Arg Lys Ala Arg Leu Arg Gly Pro Gln
1 5 10
<210> 407
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 407
Glu Gly Asp Lys Phe Arg Arg His Asp Ile Lys Tyr Asn Phe
1 5 10
<210> 408
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 408
Pro Ile Cys Arg Trp Lys Arg Ala Pro Phe Lys Trp Tyr Phe
1 5 10
<210> 409
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 409
Asn Cys Asn Arg Phe Arg Arg Thr Ile Val Arg His Cys His
1 5 10
<210> 410
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 410
Ala His Gly Lys Asp Arg Arg Tyr Val Glu Lys Leu Glu Val
1 5 10
<210> 411
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 411
Phe Met Gln Lys Cys Lys Lys Trp Trp Asp Arg Ala Val Phe
1 5 10
<210> 412
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 412
Asp Pro Pro Lys Lys Arg Lys Ser Leu Leu Arg Arg Val Ser
1 5 10
<210> 413
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 413
Tyr Cys Thr Arg Ile Arg Arg Glu Gly Met Lys Gly Ser Glu
1 5 10
<210> 414
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 414
Phe Pro Gly Lys Ser Lys Lys Gln Trp His Arg Leu Trp Pro
1 5 10
<210> 415
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 415
Tyr Glu Pro Arg Phe Lys Arg Pro Tyr Gly Lys Trp Cys His
1 5 10
<210> 416
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 416
Asp Tyr Leu Arg Ser Arg Lys Met Glu Glu Arg Phe Gln Glu
1 5 10
<210> 417
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 417
Val Gly Pro Lys Phe Arg Lys Asn His Arg Arg Gln Asn Arg
1 5 10
<210> 418
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 418
Glu Tyr Gln Lys Cys Lys Lys Pro Ser Phe Arg Leu Thr Met
1 5 10
<210> 419
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 419
Tyr Met Lys Lys Lys Arg Lys Ser Leu Leu Arg Thr Ser Leu
1 5 10
<210> 420
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 420
Cys Cys Phe Lys Leu Lys Lys Ala Tyr Asn Lys Gly Pro Phe
1 5 10
<210> 421
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 421
Asn Pro Met Lys Leu Arg Lys Ala Glu Thr Lys His Asn Val
1 5 10
<210> 422
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 422
His Ser Leu Lys Thr Arg Lys Ser Ala Phe Lys Ser Asn Thr
1 5 10
<210> 423
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 423
Gln Tyr His Lys Val Arg Lys Leu Trp Phe Arg Val Glu Pro
1 5 10
<210> 424
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 424
Phe Val Arg Lys Lys Arg Lys Ser Leu Leu Arg Asp Thr Arg
1 5 10
<210> 425
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 425
Cys Phe Gln Arg Lys Arg Lys Ser Leu Leu Arg Val Leu Arg
1 5 10
<210> 426
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 426
Val Pro Met Lys Tyr Arg Arg Cys Gly Asn Arg Gln Ser Asn
1 5 10
<210> 427
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 427
Arg Ala Arg Lys Lys Arg Lys Ser Leu Leu Arg Arg Gln Val
1 5 10
<210> 428
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 428
Pro Asn Gly Lys Val Arg Lys Arg Ile Arg Arg Arg Tyr Phe
1 5 10
<210> 429
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 429
Cys Ala Ser Lys Pro Arg Arg Thr Tyr Leu Arg Ala Ala Asn
1 5 10
<210> 430
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 430
Trp Ala His Lys Cys Lys Lys Pro Gly Gln Arg Ile Pro Pro
1 5 10
<210> 431
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 431
Leu Ser Asp Lys Lys Arg Lys Ser Leu Leu Arg Tyr Asp Tyr
1 5 10
<210> 432
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 432
Cys Ala Thr Arg Gly Lys Lys Val Phe Ser Lys Arg Thr Met
1 5 10
<210> 433
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 433
Leu Arg Gln Arg Ser Lys Arg Val Leu Glu Lys Leu Arg Pro
1 5 10
<210> 434
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 434
Leu Gly Trp Lys Lys Arg Lys Ser Leu Leu Arg Arg His Val
1 5 10
<210> 435
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 435
Leu Gln Cys Lys Tyr Arg Arg Gly Ser Asp Lys Gln Pro Gln
1 5 10
<210> 436
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 436
Ala Phe Ser Arg Ile Lys Arg Gly Val Leu Lys Leu Leu Ser
1 5 10
<210> 437
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 437
Ser Asn Val Lys Arg Lys Arg Gly Arg Cys Lys Pro Tyr Arg
1 5 10
<210> 438
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 438
Thr Val Val Lys Ser Arg Lys Cys Ser Val Arg Tyr Asn Trp
1 5 10
<210> 439
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 439
Asp Asp Val Lys Gln Arg Lys Lys His Pro Arg Val Gln Thr
1 5 10
<210> 440
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 440
Ala Ser Pro Lys Gly Arg Arg Pro Thr Phe Arg Pro Gln His
1 5 10
<210> 441
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 441
Gly Trp Leu Lys Ala Lys Arg Phe Pro Ser Arg Pro Pro Thr
1 5 10
<210> 442
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 442
Glu Thr Asn Arg Thr Arg Lys Gln Cys Tyr Lys Thr Thr Phe
1 5 10
<210> 443
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 443
Phe Thr His Lys Asn Arg Arg Asp Ser Leu Arg Val Trp Met
1 5 10
<210> 444
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 444
Pro Pro Phe Arg Leu Arg Lys Pro Leu Trp Arg Pro Gln Arg
1 5 10
<210> 445
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 445
Pro Gly Asn Arg Met Lys Lys Tyr Gln Asn Arg Val His Gly
1 5 10
<210> 446
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 446
Ser Ala His Lys Lys Arg Lys Gln Thr Leu Arg Cys Ser Glu
1 5 10
<210> 447
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 447
Leu Tyr Glu Lys Tyr Lys Lys His Asn Asn Arg Glu Asp Asp
1 5 10
<210> 448
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 448
Pro His Cys Arg Gln Lys Lys Phe Trp Ile Arg Cys Gly Thr
1 5 10
<210> 449
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 449
Val Ala Phe Lys Thr Arg Arg Arg Val Gln Arg Gln Ser Gly
1 5 10
<210> 450
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 450
Val Phe Asp Lys Phe Arg Lys Thr Glu Asn Arg Gly Val Ile
1 5 10
<210> 451
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 451
Asn His His Lys Thr Lys Arg Cys Ser Val Arg Phe Asn Ile
1 5 10
<210> 452
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 452
Gly Thr Phe Lys Trp Arg Lys Ser Gly Ala Arg Gln Tyr Leu
1 5 10
<210> 453
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 453
His Cys Leu Arg Thr Lys Lys Leu Ile Asn Lys Ile Cys Ser
1 5 10
<210> 454
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 454
Phe Phe Leu Arg Cys Arg Arg Leu Leu Gly Lys Val Gln Val
1 5 10
<210> 455
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 455
His Phe Pro Arg Ala Arg Arg Phe Glu His Arg Cys Met Leu
1 5 10
<210> 456
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 456
Gln Ile Ser Lys Leu Lys Arg Pro Ser Tyr Arg Gly Asp Asp
1 5 10
<210> 457
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 457
Glu Glu Ile Lys Gln Lys Lys Leu His Leu Arg Val Trp Phe
1 5 10
<210> 458
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 458
Ser Leu Pro Lys Trp Arg Lys Gly Gly Asp Arg Val Phe Thr
1 5 10
<210> 459
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 459
His Val Tyr Lys Asn Arg Arg Val Trp Gly Lys Gly Trp Pro
1 5 10
<210> 460
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 460
Glu Asp Leu Arg Cys Lys Lys Leu Glu Leu Arg Ser Val Ile
1 5 10
<210> 461
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 461
Thr His Asp Lys Cys Lys Lys His Asn Asp Lys Gln Ala His
1 5 10
<210> 462
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 462
Tyr Pro Glu Arg Pro Arg Lys Leu Gln Asp Lys Ser Tyr Ser
1 5 10
<210> 463
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 463
Cys Pro Trp Arg Asn Arg Lys Ala Met Ile Lys Gly Ile Ile
1 5 10
<210> 464
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 464
His Glu Ile Lys Gln Lys Lys Tyr Phe His Arg Gly His Asp
1 5 10
<210> 465
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 465
Cys Leu Glu Lys Leu Arg Lys Ala Val His Arg Gln Arg Arg
1 5 10
<210> 466
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 466
Tyr Ile Ser Lys Ser Lys Lys Thr Ala Gly Arg Trp Phe Trp
1 5 10
<210> 467
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 467
Val Thr Trp Lys Phe Arg Lys Ala Glu Lys Arg Trp Gly Tyr
1 5 10
<210> 468
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 468
Tyr Ser Ser Lys Ser Arg Lys Leu Ser Pro Arg Thr Pro Arg
1 5 10
<210> 469
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 469
Cys Val Asn Arg Val Lys Lys Ser Asp Ser Lys Gly Thr Trp
1 5 10
<210> 470
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 470
Asp His Glu Arg Glu Arg Arg Leu Trp Ser Arg Phe Pro Phe
1 5 10
<210> 471
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 471
Pro Leu Ile Lys Val Lys Arg Gly Val Gly Lys Leu Trp Asn
1 5 10
<210> 472
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 472
Thr Gly Cys Arg Cys Lys Arg Ser Met Tyr Lys Asn Leu His
1 5 10
<210> 473
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 473
Trp Thr Cys Arg Met Arg Lys Tyr Gln Leu Arg Thr Ser Glu
1 5 10
<210> 474
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 474
Ile Ala Ile Arg Pro Arg Lys Met Thr Leu Lys Ile His Pro
1 5 10
<210> 475
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 475
Gln Ile Pro Lys Gln Lys Lys Gln Glu Gln Arg Ala Ile Ser
1 5 10
<210> 476
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 476
Gln Met Gly Lys Phe Lys Lys Ile Ser Leu Lys Asn Thr Phe
1 5 10
<210> 477
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 477
Val Leu Ile Lys Gln Arg Lys Trp Gln Asp Arg Ser Cys Ser
1 5 10
<210> 478
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 478
Phe Ile Thr Lys Ser Arg Arg Gln Gln Phe Arg Asn Gln Gly
1 5 10
<210> 479
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 479
Gln Gln Phe Lys Tyr Arg Arg Glu Cys Val Lys Tyr Gly Ser
1 5 10
<210> 480
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 480
Met Glu Gly Arg Glu Lys Lys Ser Tyr Asn Lys Gly Glu Asn
1 5 10
<210> 481
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 481
Pro Thr Trp Arg His Arg Arg Tyr Cys Ala Lys Asp Ile Gly
1 5 10
<210> 482
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 482
His Met Cys Arg Ser Lys Arg Val Ala Trp Lys Asn Leu Ile
1 5 10
<210> 483
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 483
Ser Tyr Leu Lys Cys Lys Arg Ser Asp Tyr Lys Glu Val Pro
1 5 10
<210> 484
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 484
His Thr Phe Lys Leu Arg Lys Leu Cys Gln Lys Leu Phe Glu
1 5 10
<210> 485
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 485
Gln Leu Val Lys Leu Arg Lys Leu Ala Arg Arg Val Ser Tyr
1 5 10
<210> 486
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 486
Ser Glu Val Arg Pro Lys Lys Asp His Ser Arg Leu Phe Ile
1 5 10
<210> 487
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 487
Phe Leu Glu Lys Cys Arg Lys Phe Ile Ile Arg Val Ser Thr
1 5 10
<210> 488
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 488
Pro Glu Thr Arg His Arg Lys Met Phe Ile Arg Asp Phe Trp
1 5 10
<210> 489
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 489
Glu Trp Cys Lys Asn Lys Arg Trp His Ser Arg Glu Tyr Pro
1 5 10
<210> 490
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 490
Leu Trp Asn Arg Tyr Lys Lys Leu Leu Met Arg Ile Phe Trp
1 5 10
<210> 491
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 491
Cys Trp Cys Arg Gln Arg Lys Cys Phe His Lys Pro Trp Ile
1 5 10
<210> 492
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 492
Asn Leu Glu Arg Thr Lys Lys His Gly Leu Lys Gly Tyr Met
1 5 10
<210> 493
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 493
Glu Gly Gly Arg Ile Lys Arg Pro Asn Tyr Arg Gly Asp Gly
1 5 10
<210> 494
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 494
Pro Val Leu Lys Leu Arg Lys Gly Arg Val Arg Ala Gln Pro
1 5 10
<210> 495
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 495
Ser Ile Ser Arg Leu Arg Lys Ala His Gln Lys Phe Ile Pro
1 5 10
<210> 496
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 496
Glu Trp His Lys His Arg Arg Glu Met Val Arg Trp Gly Pro
1 5 10
<210> 497
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 497
Trp Ser Ala Lys Gly Arg Arg Met Gly Cys Lys Ser Thr Met
1 5 10
<210> 498
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 498
Pro Asn Gly Lys Val Arg Lys Arg Ile Arg Arg Arg Tyr Phe
1 5 10
<210> 499
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 499
Pro Cys Trp Lys Phe Arg Arg Ala Gln Met Lys Trp Gly Ile
1 5 10
<210> 500
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 500
Leu Glu Glu Arg Pro Lys Lys His Cys Ala Lys Asn His Cys
1 5 10
<210> 501
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 501
Gln Pro Trp Arg Gln Lys Lys Phe Trp Cys Arg Cys Trp Gly
1 5 10
<210> 502
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 502
Ile Met Val Lys Ile Arg Arg Cys Pro Ala Arg Pro Leu Cys
1 5 10
<210> 503
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 503
Asn Asp Trp Lys Leu Arg Lys Asp Phe Trp Arg Asn His Phe
1 5 10
<210> 504
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 504
Ala Ser Val Arg Asp Arg Lys Met Gly Tyr Lys Ser Asp Gly
1 5 10
<210> 505
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 505
Tyr Tyr Glu Lys Ile Arg Arg Gly Glu Ile Lys Ile Ala Glu
1 5 10
<210> 506
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 506
Asp Leu Val Arg Lys Arg Lys Thr Met Leu Arg Gln Leu Val
1 5 10
<210> 507
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 507
Asp Leu Ser Lys Val Arg Arg Gly His Gly Lys Asn Asp Ile
1 5 10
<210> 508
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 508
Glu Ser Tyr Arg Leu Arg Arg Gly Thr Asp Lys Glu Gln Trp
1 5 10
<210> 509
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 509
Tyr Ile Thr Lys Phe Arg Lys Phe Leu Met Lys Asp Gln Phe
1 5 10
<210> 510
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 510
Ala Trp Ala Lys Tyr Lys Lys Val Gln Pro Lys His His Thr
1 5 10
<210> 511
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 511
Thr Trp Leu Arg Gly Arg Arg Trp Pro Asp Lys Gln Pro Gln
1 5 10
<210> 512
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 512
Ala Leu Pro Arg Val Arg Lys Asp Ser His Arg Glu Glu Ile
1 5 10
<210> 513
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 513
Cys Leu Phe Lys Tyr Arg Lys Ser Cys Val Arg Leu Cys Gly
1 5 10
<210> 514
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 514
Leu Gln Asn Arg Met Arg Lys Ile Tyr Trp Lys Thr Phe Asp
1 5 10
<210> 515
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 515
Asn Asp Pro Arg Leu Arg Lys His Asn His Arg Cys Cys Thr
1 5 10
<210> 516
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 516
Tyr Gly Ser Lys Gln Arg Arg Phe Glu Glu Lys Ile Cys Gly
1 5 10
<210> 517
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 517
Cys Gly His Lys Asn Lys Lys Trp His Asn Arg Met Asp Pro
1 5 10
<210> 518
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 518
Tyr Gly Ile Arg Asp Arg Arg Pro Met Thr Lys His Thr Gln
1 5 10
<210> 519
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 519
Thr Pro Leu Lys Ser Arg Lys Tyr Trp Asn Lys His Ala Tyr
1 5 10
<210> 520
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 520
Phe Leu Tyr Lys Ala Lys Lys Ala Leu Met Arg Asp Ser Leu
1 5 10
<210> 521
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 521
Cys Asp Asp Lys Pro Lys Lys Thr Val Val Lys Leu Thr Trp
1 5 10
<210> 522
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 522
Met His Pro Arg Pro Arg Lys Met Ser His Lys Met Ser Cys
1 5 10
<210> 523
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 523
Pro Gly Glu Arg Leu Lys Lys Glu Asp His Arg Ala Ser Gly
1 5 10
<210> 524
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 524
Gly Ala Gln Arg Ala Arg Lys Pro Val Leu Arg Ala Val Gly
1 5 10
<210> 525
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 525
Met Gln Glu Lys Cys Arg Arg Cys Val Phe Lys Gly Asn Ile
1 5 10
<210> 526
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 526
Tyr Val Asp Arg Trp Arg Arg Met Gln Tyr Lys Leu Ser Ile
1 5 10
<210> 527
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 527
Trp Val Asp Arg Ser Arg Lys Phe Glu Asp Arg Asn Cys Leu
1 5 10
<210> 528
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 528
Ile Val Asn Arg Ala Lys Arg Ser Gln Ile Arg Phe Asp Val
1 5 10
<210> 529
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 529
Ile His Pro Arg Tyr Lys Arg His Gln Ala Arg Ser Cys Cys
1 5 10
<210> 530
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 530
Glu His Ser Lys His Arg Lys Asp Leu Phe Arg Met Val Gly
1 5 10
<210> 531
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 531
Tyr Thr Pro Lys Phe Arg Arg Ile Phe Trp Arg Ile Ile Asp
1 5 10
<210> 532
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 532
Thr Ile Thr Arg Val Lys Arg Ala Ser His Lys Thr Pro Ser
1 5 10
<210> 533
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 533
Val Asp Ser Arg Glu Lys Lys Trp Arg Gln Lys Cys Gln Cys
1 5 10
<210> 534
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 534
Tyr Trp Phe Arg Ile Lys Arg Ala His Ala Lys Gly Cys Pro
1 5 10
<210> 535
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 535
Asn Asp His Arg Met Arg Arg Ser Val Asp Arg Gly Glu Thr
1 5 10
<210> 536
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 536
Pro Phe Asp Arg Ala Arg Arg Asp Leu His Arg Ile Met Met
1 5 10
<210> 537
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 537
Gln Val Val Arg Leu Arg Arg Gly Lys Asn Arg Gly Thr Val
1 5 10
<210> 538
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 538
Thr Asn Gln Arg Pro Arg Arg Glu Thr Ala Lys Thr Ile Glu
1 5 10
<210> 539
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 539
Phe His His Arg His Lys Arg Ile Ala Thr Lys His Pro Ala
1 5 10
<210> 540
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 540
Glu His Cys Lys Trp Lys Lys Met Phe Asp Lys Glu Gly Glu
1 5 10
<210> 541
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 541
Phe Phe Tyr Arg Leu Arg Arg Cys Leu Thr Arg Trp Trp Val
1 5 10
<210> 542
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 542
Leu Gln Cys Lys Tyr Arg Arg Gly Ser Asp Lys Thr Val Val
1 5 10
<210> 543
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 543
Gln Thr Met Lys Phe Lys Arg Cys Cys Asn Lys Glu Met Val
1 5 10
<210> 544
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 544
Cys Ser Tyr Lys Leu Arg Arg Cys Asp Met Arg Leu Trp Gly
1 5 10
<210> 545
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 545
Met His Ile Lys Met Lys Arg Glu Gln Val Lys Asp Glu Glu
1 5 10
<210> 546
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 546
Ala Val Trp Arg Asn Arg Lys Asp Asn Val Lys Ala Ser Glu
1 5 10
<210> 547
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 547
Ala Asp His Arg Ala Lys Lys Thr Leu Ser Lys Phe Trp Thr
1 5 10
<210> 548
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 548
Met Met Ala Arg Cys Arg Arg Ile Met Cys Arg Ser Phe Asn
1 5 10
<210> 549
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 549
Pro Phe Gly Lys Phe Lys Arg Leu Glu Asn Lys Asp Glu Cys
1 5 10
<210> 550
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 550
Tyr Leu Ser Arg Thr Lys Arg Trp Leu Ala Arg Tyr Val Glu
1 5 10
<210> 551
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 551
His Thr Leu Lys Pro Lys Arg Leu Tyr Pro Arg Pro Ser Glu
1 5 10
<210> 552
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 552
Val Val Ile Lys Cys Arg Lys Trp Thr His Lys Met Asp His
1 5 10
<210> 553
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 553
Phe His Thr Lys Leu Arg Lys Pro Ile Pro Lys Ile Asp Met
1 5 10
<210> 554
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 554
Glu Asn Ile Lys Ser Arg Lys Tyr Phe Val Lys Leu Thr Trp
1 5 10
<210> 555
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 555
Ala Trp Thr Arg Phe Arg Arg Phe Gln Cys Lys Gly Met Ser
1 5 10
<210> 556
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 556
Ile Val Pro Arg Asp Arg Lys Gln Ala Leu Arg Trp Thr Asn
1 5 10
<210> 557
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 557
Cys Met Leu Arg Trp Lys Arg Trp His Met Arg Phe Asn Pro
1 5 10
<210> 558
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 558
Phe Asp Met Arg Asp Arg Arg Pro Asn Ala Arg Val Met Pro
1 5 10
<210> 559
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 559
Gln Ile Ala Lys Leu Lys Arg Asp Gln Met Lys Glu Ala Trp
1 5 10
<210> 560
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 560
Leu Thr Ser Arg Cys Lys Lys Thr Phe Gln Lys Asn Ser Glu
1 5 10
<210> 561
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 561
Met Ala Ala Lys Trp Lys Lys Asp Gly Met Lys Ser His Tyr
1 5 10
<210> 562
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 562
Ala Asn Trp Arg Thr Arg Arg Asp Leu Ala Lys Glu His Val
1 5 10
<210> 563
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 563
Pro Phe Thr Lys Met Arg Lys Gln Phe Gln Arg Met Ser Ala
1 5 10
<210> 564
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 564
Gln Ile Asn Arg Val Lys Lys Ser Asp Ala Lys Phe Cys Thr
1 5 10
<210> 565
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 565
Tyr Leu Val Lys Leu Arg Arg Phe Trp Ala Lys Ile Asp Met
1 5 10
<210> 566
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 566
Glu Tyr Val Lys Ala Lys Arg Thr Pro Trp Lys Tyr Val Val
1 5 10
<210> 567
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 567
Ala Met Cys Arg Glu Lys Arg Phe Pro Tyr Arg Ala Ile Tyr
1 5 10
<210> 568
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 568
Met Gly Leu Arg Ala Arg Lys Tyr Glu Asp Lys Thr Leu His
1 5 10
<210> 569
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 569
Phe His Met Lys Ser Lys Lys Asp Gly Asp Lys Phe Val Met
1 5 10
<210> 570
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 570
Cys Ile Phe Lys Phe Lys Lys Asn Met Phe Lys Gly Tyr Ser
1 5 10
<210> 571
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 571
Leu Asp Thr Lys Asn Arg Arg Gly Ala Ser Lys Trp Asp Tyr
1 5 10
<210> 572
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 572
Val Thr Gly Lys Gly Arg Arg Val Thr Phe Arg Cys Met Asn
1 5 10
<210> 573
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 573
Gln Leu Gly Lys Gln Lys Arg Glu Ala Thr Arg Glu Tyr Val
1 5 10
<210> 574
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 574
Ser Ser Gly Lys Trp Lys Arg Pro His Ala Lys Tyr Ile Val
1 5 10
<210> 575
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 575
Thr Cys His Lys Thr Lys Arg Ser Ile Met Arg Ala Thr Ser
1 5 10
<210> 576
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 576
Met Ile Gly Lys Cys Arg Lys Cys Gly Thr Lys Cys Tyr Ala
1 5 10
<210> 577
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 577
Phe Gln Pro Lys Ser Arg Arg Ala Glu Thr Lys His Ser Tyr
1 5 10
<210> 578
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 578
Pro Asp Leu Lys Phe Arg Lys His Gln Asp Arg Thr Ala Ile
1 5 10
<210> 579
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 579
Tyr Asn Ala Arg Val Lys Arg Glu Gly Ile Lys Asn Ile Thr
1 5 10
<210> 580
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 580
Gln Gly Ile Arg Pro Arg Lys His Leu Gln Arg Met Pro Asn
1 5 10
<210> 581
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 581
Gly Pro Cys Arg Pro Arg Arg His Ser Ile Arg Leu Leu Trp
1 5 10
<210> 582
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 582
Phe Tyr Leu Arg Glu Arg Arg His Gln Leu Lys Thr Glu Phe
1 5 10
<210> 583
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 583
Asn Met Ala Arg Ser Arg Lys Asn Glu Cys Lys Tyr Ile Ala
1 5 10
<210> 584
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 584
Asp Met Asp Lys Val Lys Lys Trp Val Trp Arg Ser Phe Pro
1 5 10
<210> 585
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 585
Val Leu Pro Lys Glu Arg Arg Trp Leu Thr Arg Phe Leu Ile
1 5 10
<210> 586
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 586
Tyr Glu His Arg His Lys Arg Phe Phe His Lys Asp Met Pro
1 5 10
<210> 587
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 587
His Gly His Lys Pro Arg Arg Trp Ile Tyr Arg Met Pro Met
1 5 10
<210> 588
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 588
Asp Gly Ile Arg Asp Lys Lys Cys Cys Trp Arg Asp Leu Ile
1 5 10
<210> 589
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 589
Glu Leu Val Lys His Arg Arg Leu Asp Phe Arg Asp Pro Met
1 5 10
<210> 590
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 590
Val His His Lys Leu Arg Arg Val His Leu Arg Leu Asp Val
1 5 10
<210> 591
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 591
Gly Ile Asn Arg Gly Arg Lys Val Ser Pro Lys Asp Glu Gln
1 5 10
<210> 592
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 592
Asn Gly Tyr Arg Ala Arg Lys Glu Asn Leu Lys Phe Pro Glu
1 5 10
<210> 593
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 593
Ala Ala Tyr Arg Ala Arg Lys Ile Val Glu Lys Ser Gly Trp
1 5 10
<210> 594
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 594
Trp Ala Pro Arg Asp Arg Arg Asn Met Gly Arg Asp Pro Ala
1 5 10
<210> 595
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 595
Ile Glu Asp Lys Ser Lys Arg Phe Asn Cys Lys Glu Cys Gly
1 5 10
<210> 596
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 596
Ile Glu Met Arg Tyr Arg Arg Asp Cys Ser Arg Phe Val Asn
1 5 10
<210> 597
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 597
Gln Gln Ile Lys Tyr Lys Arg Pro Thr Tyr Arg Met Val Cys
1 5 10
<210> 598
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 598
Leu Pro Trp Arg Glu Arg Arg Asn Gly Gly Arg Ser Ser Glu
1 5 10
<210> 599
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 599
Pro Gln Thr Arg Tyr Arg Lys Tyr Phe Tyr Arg Pro Gln Met
1 5 10
<210> 600
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 600
His Met Cys Arg Trp Arg Arg Gln Ile Asn Lys Gly Val Ser
1 5 10
<210> 601
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 601
Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg Arg
1 5 10
<210> 602
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 602
Gly Gly Gly Arg Lys Lys Arg Arg Gln Arg Arg
1 5 10
<210> 603
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 603
Gly Gly Arg Lys Lys Arg Arg Gln Arg Arg Arg
1 5 10
<210> 604
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 604
Gly Gly Gly Arg Lys Lys Arg Arg Gln Arg Arg Arg Gly Gly
1 5 10
<210> 605
<211> 14
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 605
Gly Gly Arg Lys Lys Arg Arg Gln Arg Arg Arg Gly Gly Gly
1 5 10
<210> 606
<211> 11
<212> PRT
<213> Artificial Sequence
<220>
<223> AchR binding peptide
<400> 606
Gly Gly Gly Arg Arg Arg Arg Arg Arg Arg Arg
1 5 10
Claims (6)
- 하기 화학식 3으로 표현되는 아미노산 서열로 이루어지며, 하기 XL, XM -1은 각각 3개, X는 1개의 임의의 아미노산으로 이루어진 아세틸콜린 수용체 결합 펩타이드를 포함하는 아세틸콜린 수용체 과잉활성 관련 질환 예방 또는 치료용 약학 조성물에 있어서,
상기 아세틸콜린 수용체 과잉활성 관련 질환은 경부근긴장이상증(cervical dystonia), 사지 긴장 이상증(limb dystonia), 체부 근육긴장이상증(truncal dystonia), 안검경련(blepharospasm, 얼굴떨림), 경직(spasticity), 반측안면경련(hemifacial spasm), 사시(strabismus), 안구떨림(nystagmus), 틱(tics), 만성 통증(chronic pain), 만성 편두통(chronic migraine), 신경병성 통증, 피부 주름, 사각턱, 신경인성 방광(neurogenic bladder), 배뇨근-괄약근 협동장애(detrusorsphincter dyssynergia), 식도이완 불능증(achalasia cardia), 다한증(hyperhidrosis) 및 타액 분비 과다(sialorrhea)로 이루어진 군에서 선택되는 어느 하나 이상이며,
상기 아세틸콜린 수용체 결합 펩타이드는 서열번호 201 내지 400 으로 이루어진 군에서 선택되는 어느 하나의 서열의 펩타이드인 것을 특징으로 하는 아세틸콜린 수용체 과잉활성 관련 질환 예방 또는 치료용 약학 조성물.
[화학식 3]
XL-(K 또는 R)-X-(K 또는 R)-(K 또는 R)-XM -1-(K 또는 R) - 제1항에 있어서,
상기 펩타이드는 N-말단 또는 C-말단이 변형(modification)된 것을 특징으로 하는 아세틸콜린 수용체 과잉활성 관련 질환 예방 또는 치료용 약학 조성물. - 제2항에 있어서,
상기 N-말단 또는 C-말단이 변형(modification)된 것은 팔미토일화(palmitoylation), 아세틸화(acetylation), 아미드화(amidation), 포르밀화(formylation) 또는 페길화(PEGylation)되거나, 또는 2-메르캅토아세트산(2-mercaptoacetic acid), 3-메르캅토프로피오닉산(3-mercaptopropionic acid), 6-메르캅토헥사노익산(6-mercaptohexanoic acid), 피로글루탐산(pyroglutamic acid), 숙신이미드산(succinimide acid), 시스트라민(cystramine), 시스테아민(cysteamine), 메틸에스테르(methyl ester), 에틸에스테르(ethyl ester), 벤질에스테르(benzyl ester), 미리스트산(myristic acid), 스테아르산(stearic acid), 팔미트산(palmitic acid), 콜레스테롤(cholesterol), 6-아미노 헥사노익산(6-aminohexanoic acid) 및 8-아미노 옥타노익산(8-amino octanoic acid)로 이루어진 군으로부터 선택된 1 이상의 결합에 의한 것을 특징으로 하는 아세틸콜린 수용체 과잉활성 관련 질환 예방 또는 치료용 약학 조성물. - 제1항의 조성물에 포함된 펩타이드를 포함하는 것을 특징으로 하는 주름 개선용 화장료 조성물.
- 제1항의 조성물에 포함된 펩타이드를 포함하는 것을 특징으로 하는 아세틸콜린 수용체 과잉활성 관련 질환 개선용 건강기능식품 조성물에 있어서,
상기 아세틸콜린 수용체 과잉활성 관련 질환은 경부근긴장이상증(cervical dystonia), 사지 긴장 이상증(limb dystonia), 체부 근육긴장이상증(truncaldystonia), 안검경련(blepharospasm, 얼굴떨림), 경직(spasticity), 반측안면경련(hemifacialspasm), 사시(strabismus), 안구떨림(nystagmus), 틱(tics), 만성 통증(chronic pain), 만성 편두통(chronic migraine), 신경병성 통증, 피부주름, 사각턱, 신경인성 방광(neurogenicbladder), 배뇨근-괄약근협동장애(detrusor-sphincter dyssynergia), 식도이완 불능증(achalasiacardia), 다한증(hyperhidrosis) 및 타액 분비 과다(sialorrhea)로 이루어진 군에서 선택되는 어느 하나 이상인 것을 특징으로 하는 아세틸콜린 수용체 과잉활성 관련 질환 개선용 건강기능식품 조성물. - 제1항의 조성물에 포함된 펩타이드를 포함하는 것을 특징으로 하는 아세틸콜린 수용체 결합 펩타이드를 포함하는 필러.
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| WO2004057329A1 (en) * | 2002-12-23 | 2004-07-08 | Bayer Healthcare Ag | Diagnostics and therapeutics for diseases associated with muscarinic acetylcholine receptor m1 (acm1) |
| WO2015134920A1 (en) * | 2014-03-07 | 2015-09-11 | The Arizona Board Of Regents On Behalf Of The University Of Arizona | Non-narcotic crmp2 peptides targeting sodium channels for chronic pain |
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| US8946166B2 (en) * | 2009-07-24 | 2015-02-03 | Lipotec, S.A. | Peptide-based compounds and compositions which inhibit muscle contraction |
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| KR102607079B1 (ko) | 2016-09-09 | 2023-11-28 | 주식회사 엘지생활건강 | 신경세포 투과성이 향상된 신경전달물질 조절 펩타이드 |
| KR101971092B1 (ko) * | 2017-06-28 | 2019-04-23 | 한국세라믹기술원 | 아세틸콜린수용체 결합 펩타이드 |
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| WO2004057329A1 (en) * | 2002-12-23 | 2004-07-08 | Bayer Healthcare Ag | Diagnostics and therapeutics for diseases associated with muscarinic acetylcholine receptor m1 (acm1) |
| WO2015134920A1 (en) * | 2014-03-07 | 2015-09-11 | The Arizona Board Of Regents On Behalf Of The University Of Arizona | Non-narcotic crmp2 peptides targeting sodium channels for chronic pain |
| WO2016040517A1 (en) * | 2014-09-09 | 2016-03-17 | Merial Inc. | Mammalian cell lines expressing functional nematode acetylcholine receptors and use thereof for high-throughput screening assays |
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