Kelly J M, Sterry S J, Cose S, Turner S J, Fecondo J, Rodda S, Fink P J, Carbone F R
Department of Pathology and Immunology, Monash Medical School, Victoria, Australia.
Eur J Immunol. 1993 Dec;23(12):3318-26. doi: 10.1002/eji.1830231239.
We have analyzed the T cell receptor (TCR) repertoire found in the major histocompatibility complex class I-restricted cytotoxic T lymphocyte (CTL) response to the protein ovalbumin (OVA). Despite skewing towards the expression of V beta 5.2+TCR by OVA-specific CTL from C57BL/6 mice, we found a relatively high degree of diversity in V(D)J usage in both TCR alpha- and beta-chains. Closer examination showed that the majority of these sequences encoded negatively and positively charged residues at their respective TCR alpha- and beta-chain VJ or VDJ junctions. These junctions form the third complementarity-determining regions (CDR3) of the TCR polypeptides involved in the direct interaction with the class I-bound peptide. Crystallographic analyses of Kb-peptide complexes predict that the major determinant from OVA, peptide OVA257-264 (SIINFEKL), contains two exposed charged side chains which can contact the TCR. These are the negatively charged glutamic acid at determinant position 6 (P6) and the positively charged lysine at P7. To examine whether the TCR alpha-chain makes contact with P7 lysine, we established a single chain TCR transgenic C57BL/6 mouse line where all T cells express a TCR beta-chain derived from the V beta 5.2+ clone B3. OVA-specific T cells derived from in vivo primed transgenic mice preferentially expressed TCR alpha-chains that also contained negatively charged junctional residues despite some further variation in V alpha and J alpha sequences. Stimulation of naive TCR beta-chain transgenic T cells with a P7 substitution peptide analogue induced a T cell response that was no longer cross-reactive with the wild-type OVA257-264 determinant, suggesting that the TCR alpha-chain from the T cell clone B3 can determine the specificity for this residue. Consequently, these results reveal the existence of conserved residues in the CDR3 of TCR alpha- and beta-chains specific for OVA257-264 and identify their possible orientation over the peptide-class I complex.
我们分析了在主要组织相容性复合体I类限制性细胞毒性T淋巴细胞（CTL）对卵清蛋白（OVA）的应答中发现的T细胞受体（TCR）库。尽管来自C57BL/6小鼠的OVA特异性CTL偏向于表达Vβ5.2+TCR，但我们发现TCRα链和β链在V（D）J使用上具有相对较高的多样性。进一步检查发现，这些序列中的大多数在其各自的TCRα链和β链VJ或VDJ连接处编码带负电荷和正电荷的残基。这些连接处形成了TCR多肽的第三个互补决定区（CDR3），参与与I类结合肽的直接相互作用。Kb-肽复合物的晶体学分析预测，OVA的主要决定簇，肽OVA257-264（SIINFEKL），包含两个暴露的带电侧链，可与TCR接触。这些是决定簇位置6（P6）处带负电荷的谷氨酸和P7处带正电荷的赖氨酸。为了检查TCRα链是否与P7赖氨酸接触，我们建立了一个单链TCR转基因C57BL/6小鼠品系，其中所有T细胞都表达源自Vβ5.2+克隆B3的TCRβ链。来自体内致敏转基因小鼠的OVA特异性T细胞优先表达TCRα链，这些α链也含有带负电荷的连接残基，尽管Vα和Jα序列存在一些进一步的变异。用P7替代肽类似物刺激幼稚TCRβ链转基因T细胞诱导了一种T细胞应答，该应答不再与野生型OVA257-264决定簇发生交叉反应，这表明来自T细胞克隆B3的TCRα链可以决定对该残基的特异性。因此，这些结果揭示了TCRα链和β链的CDR3中存在对OVA257-264特异的保守残基，并确定了它们在肽-I类复合物上的可能取向。
