IDENTIFICATION OF CONSERVED T-CELL RECEPTOR CDR3 RESIDUES CONTACTING KNOWN EXPOSED PEPTIDE SIDE-CHAINS FROM A MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I-BOUND DETERMINANT
IDENTIFICATION OF CONSERVED T-CELL RECEPTOR CDR3 RESIDUES CONTACTING KNOWN EXPOSED PEPTIDE SIDE-CHAINS FROM A MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I-BOUND DETERMINANT
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DOI：
10.1002/eji.1830231239
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发表时间：
1993-12-01
影响因子：
5.4
通讯作者：
CARBONE, FR
中科院分区：
文献类型：
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作者：
KELLY, JM;STERRY, SJ;CARBONE, FR
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）的反应中发现的T细胞受体（TCR）库。尽管C57BL/6小鼠的ova特异性CTL倾向于表达Vbeta5.2+ TCR，但我们发现TCR α链和β链中V(D)J的使用具有相对高度的多样性。进一步研究表明，这些序列中的大多数在各自的TCR α链和β链VJ或VDJ连接处编码带负电和带正电的残基。这些连接形成了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。K(b)-肽复合物的晶体学分析预测OVA的主要决定因子，肽OVA257-264 (SIINFEKL)，包含两个暴露的带电侧链，可以与TCR接触。它们是带负电的谷氨酸在决定位置6 （P6）和带正电的赖氨酸在P7。为了研究TCR α链是否与P7赖氨酸接触，我们建立了单链TCR转基因C57BL/6小鼠系，其中所有T细胞都表达来自Vbeta5.2+克隆B3的TCR β链。来自体内转基因小鼠的ova特异性T细胞优先表达含有负电荷连接残基的TCR α链，尽管Va和j α序列有进一步的变化。用P7替代肽类似物刺激原始TCR β链转基因T细胞诱导T细胞反应，不再与野生型OVA257-264决定因子交叉反应，这表明来自T细胞克隆B3的TCR α链可以决定该残基的特异性。因此，这些结果揭示了在OVA257-264特异性的TCR α和β链的CDR3中存在保守残基，并确定了它们在肽类I络合物上的可能取向。
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 Vbeta5.2+ 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 K(b)-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 Vbeta5.2+ 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 Va and Jalpha 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, sugesting 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.