Atassi M Z, Yoshioka M, Bean M, Bixler G S
Verna and Marrs McLean Department of Biochemistry, Baylor College of Medicine, Houston, TX 77030.
Biochem J. 1987 Sep 1;246(2):307-12. doi: 10.1042/bj2460307.
Six regions (T sites) of myoglobin (Mb) were found by a comprehensive synthetic strategy to stimulate Mb-primed lymph-node cells. To define precisely the N-terminal boundary of the immunodominant T site (residues 107-120) with site-specific T-cell clones and to determine the effects of peptide size on their stimulation, two sets of peptides were employed. In one set, the peptides were elongated to the left from His-113 by one-residue increments of the Mb sequence. The other set represented an identical stepwise elongation by one-residue increments of the Mb sequence, but which were extended by additional unrelated ('nonsense') residues to a uniform size of 14 residues. Examination of the proliferative responses of eight T-cell clones, derived from Mb-primed DBA/2 (H-2d) or SJL (H-2s) mice, revealed a dramatic non-specific size requirement. In every clone, the longer nonsense-extended peptides achieved maximum stimulating activity at a lower optimum peptide dose than its natural-sequence, but shorter, analogue. In addition, slight (one-residue) differences in the N-terminal boundaries among the clones was observed. Thus, the fine specificity of each clone was mapped to the region from residue 111 or 112 to about residue 120 of Mb, which coincides with the site of B-cell recognition and resides in a small discrete surface region of the protein chain.
通过一种全面的合成策略发现，肌红蛋白（Mb）的六个区域（T位点）能够刺激经Mb致敏的淋巴结细胞。为了利用位点特异性T细胞克隆精确界定免疫显性T位点（第107 - 120位氨基酸残基）的N端边界，并确定肽大小对其刺激作用的影响，使用了两组肽。在一组中，肽从His - 113开始向左以Mb序列的一个氨基酸残基增量进行延伸。另一组代表通过Mb序列的一个氨基酸残基增量进行相同的逐步延伸，但通过额外的无关（“无义”）残基延伸至统一的14个残基大小。对源自经Mb致敏的DBA/2（H - 2d）或SJL（H - 2s）小鼠的八个T细胞克隆的增殖反应进行检测，结果显示出显著的非特异性大小需求。在每个克隆中，较长的无义延伸肽在比其天然序列但较短的类似物更低的最佳肽剂量下达到最大刺激活性。此外，观察到各克隆之间N端边界存在轻微（一个氨基酸残基）差异。因此，每个克隆的精细特异性被定位到Mb第111或112位氨基酸残基至约第120位氨基酸残基的区域，该区域与B细胞识别位点重合，且位于蛋白质链的一个小的离散表面区域。
