From the Roy J. Carver Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa 50011.
From the Roy J. Carver Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, Iowa 50011
J Biol Chem. 2018 Dec 21;293(51):19899-19908. doi: 10.1074/jbc.RA118.005273. Epub 2018 Oct 25.
Therapeutic mAbs engage Fc γ receptor III (CD16) to elicit a protective cell-mediated response and destroy the target tissue. Newer drugs designed to bind CD16a with increased affinity surprisingly also elicit protective CD16b-mediated responses. However, it is unclear why IgG binds CD16a with more than 10-fold higher affinity than CD16b even though these receptors share more than 97% identity. Here we identified one residue, Gly-129, that contributes to the greater IgG binding affinity of CD16a. The CD16b variant D129G bound IgG1 Fc with 2-fold higher affinity than CD16a and with 90-fold higher affinity than the WT. Conversely, the binding affinity of CD16a-G129D was decreased 128-fold relative to WT CD16a and comparably to that of WT CD16b. The interaction of IgG1 Fc with CD16a, but not with CD16b, is known to be sensitive to the composition of the asparagine-linked carbohydrates (-glycans) attached to the receptor. CD16a and CD16b-D129G displaying minimally processed oligomannose -glycans bound to IgG1 Fc with about 5.2-fold increased affinity compared with variants with highly processed complex-type -glycans. CD16b and the CD16a-G129D variant exhibited a smaller 1.9-fold affinity increase with oligomannose -glycans. A model of glycosylated CD16b bound to IgG1 Fc determined to 2.2 Å resolution combined with a 250-ns all-atom molecular dynamics simulation showed that the larger Asp-129 residue deformed the Fc-binding surface. These results reveal how Asp-129 in CD16b affects its binding affinity for IgG1 Fc and suggest that antibodies engineered to engage CD16b with high affinity must accommodate the Asp-129 side chain.
治疗性单抗与 Fcγ 受体 III（CD16）结合，引发保护性细胞介导的反应并破坏靶组织。新设计的旨在与 CD16a 结合以增加亲和力的药物出乎意料地也引发了保护性 CD16b 介导的反应。然而，目前尚不清楚为什么 IgG 与 CD16a 的结合亲和力比 CD16b 高 10 多倍，尽管这些受体的同一性超过 97%。在这里，我们确定了一个残基 Gly-129，它有助于 CD16a 与 IgG 结合的亲和力更高。CD16b 变体 D129G 与 IgG1 Fc 的结合亲和力比 CD16a 高 2 倍，比 WT 高 90 倍。相反，CD16a-G129D 的结合亲和力相对于 WT CD16a 降低了 128 倍，与 WT CD16b 相当。已知 IgG1 Fc 与 CD16a 的相互作用对附着在受体上的天冬酰胺连接的碳水化合物（N-聚糖）的组成敏感，但与 CD16b 无关。CD16a 和 CD16b-D129G 显示最小加工的寡甘露糖-N-聚糖与 IgG1 Fc 的结合亲和力增加了约 5.2 倍，与具有高度加工的复杂型-N-聚糖的变体相比。CD16b 和 CD16a-G129D 变体与寡甘露糖-N-聚糖的亲和力增加较小，为 1.9 倍。结合 250-ns 全原子分子动力学模拟，确定分辨率为 2.2 Å 的糖基化 CD16b 与 IgG1 Fc 结合的模型表明，较大的 Asp-129 残基使 Fc 结合表面变形。这些结果揭示了 CD16b 中的 Asp-129 如何影响其与 IgG1 Fc 的结合亲和力，并表明设计用于与 CD16b 以高亲和力结合的抗体必须适应 Asp-129 侧链。
