Abstract
The main viral protease (3CL pro ) is indispensable for SARS-CoV-2 replication. We delineate the human protein substrate landscape of 3CL pro by TAILS substrate-targeted N-terminomics. We identify more than 100 substrates in human lung and kidney cells supported by analyses of SARS-CoV-2-infected cells. Enzyme kinetics and molecular docking simulations of 3CL pro engaging substrates reveal how noncanonical cleavage sites, which diverge from SARS-CoV, guide substrate specificity. Cleaving the interactors of essential effector proteins, effectively stranding them from their binding partners, amplifies the consequences of proteolysis. We show that 3CL pro targets the Hippo pathway, including inactivation of MAP4K5, and key effectors of transcription, mRNA processing, and translation. We demonstrate that Spike glycoprotein directly binds galectin-8, with galectin-8 cleavage disengaging CALCOCO2/NDP52 to decouple antiviral-autophagy. Indeed, in post-mortem COVID-19 lung samples, NDP52 rarely colocalizes with galectin-8, unlike in healthy lungs. The 3CL pro substrate degradome establishes a foundational substrate atlas to accelerate exploration of SARS-CoV-2 pathology and drug design.
Keywords: COVID-19; SARS-CoV-2 3CL(pro); SARS-CoV-2 main protease; active site structure; degradomics; interactome; proteases; proteomics; subsite specificity; substrates.
【저자키워드】 COVID-19, proteomics, SARS-CoV-2 main protease, Proteases, interactome, SARS-CoV-2 3CL(pro), active site structure, degradomics, subsite specificity, substrates., 【초록키워드】 SARS-CoV-2, pathology, spike, drug design, translation, SARS-CoV, Transcription, molecular docking, lung, 3CL pro, protease, kidney, Protein, specificity, mRNA, inactivation, Lungs, human lung, pathway, cleavage, target, Post-mortem, glycoprotein, SARS-CoV-2 replication, binding, Analysis, SARS-CoV-2-infected cells, substrate, substrates, effector, effector proteins, cleavage sites, Cell, consequence, bind, identify, healthy, supported, amplify, accelerate, MAP4K5, 【제목키워드】 3CL pro, global analysis, the SARS-CoV-2,