Chang H C, Smolyar A, Spoerl R, Witte T, Yao Y, Goyarts E C, Nathenson S G, Reinherz E L
Laboratory of Immunobiology Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA, 02115, USA.
J Mol Biol. 1997 Aug 15;271(2):278-93. doi: 10.1006/jmbi.1997.1169.
The molecular interactions between the CD8 co-receptor dependent N15 and N26 T cell receptors (TCRs) and their common ligand, the vesicular stomatitis virus octapeptide (VSV8) bound to H-2Kb, were studied to define the docking orientation(s) of MHC class I restricted TCRs during immune recognition. Guided by the molecular surfaces of the crystallographically defined peptide/MHC and modeled TCRs, a series of mutations in exposed residues likely contacting the TCR ligand were analyzed for their ability to alter peptide-triggered IL-2 production in T cell transfectants. Critical residues which diminished antigen recognition by 1000 to 10,000-fold in molar terms were identified in both N15 Valpha (alphaE94A or alphaE94R, Y98A and K99) and Vbeta (betaR96A, betaW97A and betaD99A) CDR3 loops. Mutational analysis indicated that the Rp1 residue of VSV8 is critical for antigen recognition of N15 TCR, but R62 of H-2Kb is less critical. More importantly, the alphaE94R mutant could be fully complemented by a reciprocal charge reversal at Kb R62 (R62E). This result suggests a direct interaction between N15 TCR Valpha E94R and Kb R62E residues. As Rp1 of VSV8 is adjacent to R62 in the VSV8/Kb complex and essential for T cell activation, this orientation implies that the N15 Valpha CDR3 loop interacts with the N-terminal residues of VSV8 with the Valpha domain docking to the Kb alpha2 helix while the N15 Vbeta CDR3 loop interacts with the more C-terminal peptide residues and the Vbeta domain overlies the Kb alpha1 helix. An equivalent orientation is suggested for N26, a second VSV8/Kb specific TCR. Given that genetic analysis of two different class II MHC-restricted TCRs and two crystallographic studies of class I restricted TCRs offers a similar overall orientation of V domains relative to alpha-helices, these data raise the possibility of a common docking mode between TCRs and their ligands regardless of MHC restriction.
研究了CD8共受体依赖性N15和N26 T细胞受体（TCR）与其共同配体——与H-2Kb结合的水疱性口炎病毒八肽（VSV8）之间的分子相互作用，以确定免疫识别过程中MHC I类限制性TCR的对接方向。在晶体学定义的肽/MHC和建模的TCR分子表面的指导下，分析了可能与TCR配体接触的暴露残基中的一系列突变，以研究其改变T细胞转染子中肽触发的IL-2产生的能力。在N15 Valpha（alphaE94A或alphaE94R、Y98A和K99）和Vbeta（betaR96A、betaW97A和betaD99A）CDR3环中均鉴定出关键残基，这些残基在摩尔水平上使抗原识别降低了1000至10000倍。突变分析表明，VSV8的Rp1残基对于N15 TCR的抗原识别至关重要，但H-2Kb的R62则不太关键。更重要的是，alphaE94R突变体可以通过Kb R62处的相互电荷反转（R62E）得到完全互补。这一结果表明N15 TCR Valpha E94R和Kb R62E残基之间存在直接相互作用。由于在VSV8/Kb复合物中VSV8的Rp1与R62相邻且对T细胞活化至关重要，这种方向意味着N15 Valpha CDR3环与VSV8的N端残基相互作用，同时Valpha结构域对接至Kb alpha2螺旋，而N15 Vbeta CDR3环与更多C端肽残基相互作用，且Vbeta结构域覆盖Kb alpha1螺旋。对于第二个VSV8/Kb特异性TCR N26，也提出了类似的方向。鉴于对两种不同的II类MHC限制性TCR的遗传分析以及对I类限制性TCR的两项晶体学研究提供了V结构域相对于alpha螺旋的总体相似方向，这些数据增加了TCR与其配体之间存在共同对接模式的可能性，而与MHC限制无关。
