The kinetics of SARS-CoV-2 nsp7-11 polyprotein processing and impact on complexation with nsp16

Abstract

In severe-acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, polyproteins (pp1a/pp1ab) are processed into non-structural proteins (nsps), which largely form the replication/transcription complex (RTC). The polyprotein processing and complex formation is critical and offers potential therapeutic targets. However, the interplay of polyprotein processing and RTC-assembly are poorly understood. Here, we studied two key aspects: The influence of the pp1a terminal nsp11 on the order of polyprotein processing by viral main protease Mpro and the influence of polyprotein processing on core enzyme complex formation. We established a method based on native MS to determine rate constants k considering the structural environment. This enabled us to quantify the multi-reaction kinetics of coronavirus polyprotein processing for the first time. Our results serve as a blueprint for other multi-cleavage reactions. Further, it offers a detailed and quantifiable perspective to the dynamic reactions of SARS-CoV-2 polyprotein processing, which is required for development of novel antivirals.