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Effects of polymorphism of HLA on binding of antigenic peptides to HLA and recognition of peptides complexed with HLA by T cell receptor

HLA多态性对抗原肽与HLA结合及T细胞受体识别HLA复合肽的影响

基本信息

批准号：
03452276
负责人：
NISHIMURA Yasuharu
金额：
$ 4.35万
依托单位：
Kumamoto University School of Medicine (1992-1993)Kyushu University (1991)
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1991
资助国家：
日本
起止时间：
1991 至 1993
项目状态：
已结题

项目摘要

HLA-DR molecules are highly polymorphic molecules and bind antigenic peptides to present it to CD4 positive helper T cells. The particular DR alleles are thought to determine susceptibility to autoimmune diseases because the frequencies of particular DR allelse are increased in the patients group as compared with those in healthy controls. Thus DRB1 ^<**>0405 and DRB1 ^<**>0406 which differs in only four amino acid residues from DRB1 ^<**>0405 control susceptibility to rheumatoid arthritis and insulin autoimmune syndrome respectively. We have investigated the structural characteristics of antigenic peptides bound to either DRB1 ^<**>0405 or DRB1 ^<**>0406 molecules. For this aim, self peptides bound to purified DRB1 ^<**>0405 molecules were isolated and fractionated by a reversed phase HPLC.Amino acid sequences of seven independent peptides were determined and four of them were identified to be fragments of known proteins. These self peptides were synthesized and radio-iodinated, and binding of labeled peptides to DR molecules were investigated. The strongest binder among seven self peptides was identified and this peptide bound to both DRB1 ^<**>0405 and DRB1 ^<**>0406 equally. The binding inhibition assay, in which binding activity of a peptide to be tested was determined by measuring inhibitory effect of the excess amount of peptide on binding of the labeled strongest binder to DR molecule, was established. In order to determine amino acid residues important for DR binding in the strongest binder, analogue peptides having only one amino acid substitution from the original peptides were synthesized and their binding to both DRB1 ^<**>0405 and DRB1 ^<**>0406 was investigated. DR binding peptides consisted of nine to twenty amino acids were bound at three major anchorage amino acid residues of the peptide to DR molecule. These three anchorage residues were separated by two and one intervening amino acids. The first

HLA-DR分子是高度多态的分子，结合抗原肽将其呈递给CD4阳性辅助性T细胞。特定的DR等位基因被认为决定了对自身免疫性疾病的易感性，因为与健康对照组相比，患者组中特定DR等位基因的频率增加。因此，DRB1 ^<**>0405和DRB1 ^<**>0406分别控制类风湿关节炎和胰岛素自身免疫性综合征的易感性，而DRB1 ^<**>0405与DRB1 ^<**>0406仅存在4个氨基酸残基的差异。我们研究了与DRB1 ^<**>0406或DRB1 ^<**>0406分子结合的抗原肽的结构特征。为此，分离与纯化的DRB1 ^<**>0405分子结合的自肽，并通过反相高效液相色谱进行分离。测定了7个独立肽的氨基酸序列，其中4个为已知蛋白的片段。合成了这些自肽并进行了放射性碘化处理，并研究了标记肽与DR分子的结合。结果表明，该肽与DRB1 ^<**>0406和DRB1 ^<**>0406的结合强度最高。建立了结合抑制实验，通过测量过量肽对标记的最强结合剂与DR分子结合的抑制作用来确定待测肽的结合活性。为了确定在最强的结合物中DR结合的重要氨基酸残基，我们合成了与原始肽只有一个氨基酸取代的类似肽，并研究了它们与DRB1 ^<**>0406和DRB1 ^<**>0406的结合。DR结合肽由9 ~ 20个氨基酸组成，在肽的3个主要锚定氨基酸残基上与DR分子结合。这三个锚定残基被两个和一个中间氨基酸分开。第一个