Yan X, Evans S V, Kaminki M J, Gillies S D, Reisfeld R A, Houghton A N, Chapman P B
Immunology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
J Immunol. 1996 Aug 15;157(4):1582-8.
mAb against GD3 ganglioside demonstrate homophilic binding in which soluble anti-GD3 mAb bind, through the GD3 binding site, to a VH idiotope (designated IdHOM) on solid phase anti-GD3 mAb. In this way, homophilic binding provides a mechanism for amplifying the binding of mAb to cell surface GD3. We show that serine 52a, within CDR2, is required for IdHOM expression, homophilic binding, and high avidity binding to cell surface GD3. Computer modeling based on the crystal structure of anti-GD3 mAb R24 showed serine 52a situated at the mouth of the GD3 binding pocket, but not directly involved with GD3 binding. Substitutions at position 52a predicted to maintain the GD3 binding pocket (e.g., threonine) resulted in the loss of IdHOM expression and homophilic binding and markedly decreased binding to cell surface GD3, but maintained low avidity GD3 binding as measured by ELISA. All other substitutions at position 52a were predicted to significantly distort the GD3 binding pocket and resulted in the loss of both homophilic binding and any detectable avidity for GD3. We have structurally defined IdHOM and conclude that this idiotope is not required for the GD3 binding pocket, but that the idiotope is necessary for homophilic binding, which is required for high avidity binding to cell surface GD3. We speculate that selection of certain VH genes may result in the expression of idiotopes that allow homophilic binding, and this may represent a general mechanism for increasing the avidity of Abs against T cell-independent Ags.
抗GD3神经节苷脂单克隆抗体表现出嗜同性结合，即可溶性抗GD3单克隆抗体通过GD3结合位点与固相抗GD3单克隆抗体上的VH独特型（命名为IdHOM）结合。通过这种方式，嗜同性结合为放大单克隆抗体与细胞表面GD3的结合提供了一种机制。我们发现，互补决定区2（CDR2）内的丝氨酸52a是IdHOM表达、嗜同性结合以及与细胞表面GD3高亲和力结合所必需的。基于抗GD3单克隆抗体R24晶体结构的计算机建模显示，丝氨酸52a位于GD3结合口袋的开口处，但不直接参与GD3结合。预测在52a位点进行能维持GD3结合口袋的取代（例如苏氨酸）会导致IdHOM表达和嗜同性结合丧失，并显著降低与细胞表面GD3的结合，但通过酶联免疫吸附测定（ELISA）测得仍维持低亲和力的GD3结合。预测52a位点的所有其他取代会显著扭曲GD3结合口袋，并导致嗜同性结合以及对GD3的任何可检测亲和力丧失。我们已从结构上定义了IdHOM，并得出结论，该独特型对于GD3结合口袋并非必需，但对于嗜同性结合是必需的，而嗜同性结合是与细胞表面GD3高亲和力结合所必需的。我们推测，某些VH基因的选择可能导致允许嗜同性结合的独特型表达，这可能代表了一种增加抗体对非T细胞依赖性抗原亲和力的普遍机制。
