The membrane anchor influences ligand binding two-dimensional kinetic rates and three-dimensional affinity of FcγRIII (CD16)
The membrane anchor influences ligand binding two-dimensional kinetic rates and three-dimensional affinity of FcγRIII (CD16)
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DOI：
10.1074/jbc.275.14.10235
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发表时间：
2000-04-07
影响因子：
4.8
通讯作者：
Zhu, C
中科院分区：
文献类型：
--
作者：
Chesla, SE;Li, P;Zhu, C
动力学速率和亲和力是涉及受体-配体结合的生物过程的基本决定因素。通过使用微量移液器方法，我们测量了人Fc γ受体III（CD 16）与IgG相互作用的动力学，当这两种分子结合到并列的细胞膜。CD 16是已知天然存在于跨膜（TM，CD 16 b）和糖基磷脂酰肌醇（GPI，CD 16 b）同种型中的仅有的四种真核受体之一，这种锚同种型共存的生物学意义尚不清楚。在此，我们发现锚影响动力学速率;与CD 16 a-TM相比，CD 16 a-GPI结合更快，与人和兔IgG的亲和力更高，但与鼠IgG 2a的亲和力更低。用可溶性IgG配体观察到相同的差异性亲和力模式，一种单克隆抗体与CD 16 a-GPI的亲和力高于CD 16 a-TM，而另一种单克隆抗体与CD 16 a-TM的亲和力较强，而与CD 16 a-GPI的亲和力较弱。我们建议作为观察到的锚效应的机制的构象差异，因为它不能被解释为不同的扩散性，灵活性，方向，高度，分布，或细胞膜上的两个分子的集群。这些数据表明，在胞外域羧基末端的IG超家族受体的共价修饰可对配体结合动力学产生影响。
Kinetic rates and affinity are essential determinants for biological processes that involve receptor-ligand binding. By using a micropipette method, we measured the kinetics of human Fc gamma receptor III (CD16) interacting with IgG when the two molecules were bound to apposing cellular membranes. CD16 is one of only four eukaryotic receptors known to exist natively in both the transmembrane (TM, CD16b) and glycosylphosphatidylinositol (GPI, CD16b) isoforms, The biological significance of this anchor isoform coexistence is not clear. Here we showed that the anchor influenced kinetic rates; compared with CD16a-TM, CD16a-GPI bound faster and with higher affinities to human and rabbit IgGs but slower and with lower affinity to murine IgG2a. The same differential affinity patterns were observed using soluble IgG ligands, A monoclonal antibody bound CD16a-GPI with higher affinity than CD16a-TM, whereas another monoclonal antibody reacted strongly with CD16a-TM but weakly with CD16a-GPI, No major differential glycosylation between the two CD16a isoforms was detected by SDS-polyacrylamide gel electrophoresis analysis. We suggest a conformational difference as the mechanism underlying the observed anchor effect, as it cannot be explained by the differing diffusivity, flexibility, orientation, height, distribution, or clustering of the two molecules on the cell membrane. These data demonstrate that a covalent modification of an Ig superfamily receptor at the carboxyl terminus of the ectodomain can have an impact on ligand binding kinetics.