Department of Biological Chemistry and Molecular Pharmacology, Blavatnik Institute, Harvard Medical School, Boston, MA 02115, USA.
Department of Cancer Biology, Dana Farber Cancer Institute, Boston, MA 02215, USA.
Cell. 2023 Aug 17;186(17):3632-3641.e10. doi: 10.1016/j.cell.2023.06.026. Epub 2023 Jul 28.
The endopeptidase ADAM10 is a critical catalyst for the regulated proteolysis of key drivers of mammalian development, physiology, and non-amyloidogenic cleavage of APP as the primary α-secretase. ADAM10 function requires the formation of a complex with a C8-tetraspanin protein, but how tetraspanin binding enables positioning of the enzyme active site for membrane-proximal cleavage remains unknown. We present here a cryo-EM structure of a vFab-ADAM10-Tspan15 complex, which shows that Tspan15 binding relieves ADAM10 autoinhibition and acts as a molecular measuring stick to position the enzyme active site about 20 Å from the plasma membrane for membrane-proximal substrate cleavage. Cell-based assays of N-cadherin shedding establish that the positioning of the active site by the interface between the ADAM10 catalytic domain and the bound tetraspanin influences selection of the preferred cleavage site. Together, these studies reveal the molecular mechanism underlying ADAM10 proteolysis at membrane-proximal sites and offer a roadmap for its modulation in disease.
内肽酶 ADAM10 是调节哺乳动物发育、生理的关键驱动蛋白的蛋白水解以及 APP 作为主要的α-分泌酶的非淀粉样蛋白水解的关键催化剂。ADAM10 的功能需要与 C8-四跨膜蛋白形成复合物，但四跨膜蛋白结合如何使酶活性位点定位于靠近质膜的切割位置仍然未知。我们在此展示了一个 vFab-ADAM10-Tspan15 复合物的冷冻电镜结构，该结构表明 Tspan15 结合可解除 ADAM10 的自身抑制作用，并充当分子量规，将酶的活性位点定位在距质膜约 20Å 的位置，以进行靠近质膜的底物切割。基于细胞的 N-钙粘蛋白脱落测定表明，ADAM10 催化结构域与结合的四跨膜蛋白之间的界面将活性位点定位，从而影响优选切割位点的选择。总之，这些研究揭示了 ADAM10 在质膜附近位点进行蛋白水解的分子机制，并为其在疾病中的调节提供了路线图。
