Carbone F R, Fox B S, Schwartz R H, Paterson Y
J Immunol. 1987 Mar 15;138(6):1838-44.
The B10.A T cell proliferative response to pigeon cytochrome c is largely directed to a single site in the molecule located at the carboxyl terminus within the amino acid sequence of residues 81 to 104. This study uses the pigeon cytochrome c-specific T cell clone A.E7 and synthetic peptide analogs to clarify the role of certain residues within this sequence in T cell recognition. By using the helically constrained amino acid, alpha-aminoisobutyric acid, alternated with alanine in an amino-terminal leader sequence, we generated a series of molecules of similar length and alpha-helical conformation but which contain increasing lengths of the native sequence. By comparing the stimulatory ability of this series of peptides, we have clearly identified that the isoleucyl residue at position 95 in pigeon cytochrome c is essential for T cell recognition. This series, when compared with a series containing the same native sequences but without the leader sequence, also showed that the presence of the leader sequence has a general effect on enhancement of T cell recognition. An analysis of the conformational preferences of the peptides using circular dichroism indicated that all of the peptides with leader sequences have a strong preference for the alpha-helical conformation in nonpolar solvents. However, the introduction of helix-breaking residues into these peptides, with a concomitant measured reduction in alpha-helix, did not affect their recognition by clone A.E7. This implies that factors other than conformational stabilization are responsible for the full potency of these peptides. Binding studies to phospholipid vesicles indicated that residues in the leader sequence and in the amino terminus of segment 81-104 beyond residue 95 were important in increasing the ability of the antigens to bind to membranes. These results suggest that the capacity to bind to membranes may be a significant factor in the dose response of T cells to exogenously presented peptides.
B10.A T细胞对鸽细胞色素c的增殖反应主要针对该分子中位于氨基酸序列81至104位羧基末端的单个位点。本研究使用鸽细胞色素c特异性T细胞克隆A.E7和合成肽类似物来阐明该序列中某些残基在T细胞识别中的作用。通过在氨基末端前导序列中使用与丙氨酸交替的螺旋约束氨基酸α-氨基异丁酸，我们生成了一系列长度相似且具有α-螺旋构象但包含越来越长天然序列的分子。通过比较这一系列肽的刺激能力，我们清楚地确定鸽细胞色素c中第95位的异亮氨酸残基对于T细胞识别至关重要。与一系列包含相同天然序列但无前导序列的肽相比，该系列肽还表明前导序列的存在对增强T细胞识别具有普遍作用。使用圆二色性对肽的构象偏好进行分析表明，所有带有前导序列的肽在非极性溶剂中都强烈偏好α-螺旋构象。然而，在这些肽中引入破坏螺旋的残基，同时伴随着α-螺旋的测量减少，并不影响克隆A.E7对它们的识别。这意味着除了构象稳定之外的因素对这些肽的全部效力负责。与磷脂囊泡的结合研究表明，前导序列和81 - 104片段氨基末端中95位残基以外的残基在增加抗原与膜结合的能力方面很重要。这些结果表明，与膜结合的能力可能是T细胞对外源呈递肽剂量反应的一个重要因素。
