TY - JOUR
T1 - SARS-CoV-2 polyprotein substrate regulates the stepwise Mpro cleavage reaction
AU - Narwal, Manju
AU - Armache, Jean Paul
AU - Edwards, Thomas J.
AU - Murakami, Katsuhiko S.
N1 - Funding Information:
This research was supported in part by the National Cancer Institute ’s National Cryo-EM Facility at the Frederick National Laboratory for Cancer Research under contract HSSN261200800001E. This work was supported by an NIH grant ( R35 GM131860 ) to K. S. M. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Funding Information:
We thank Joyce Jose for providing a plasmid with the Mpro-encoding gene. We thank Cryo-EM facility at Penn State for the initial screening and optimization of the cryo-EM specimen preparations. We thank Boris Krichel at the University of Wisconsin-Madison, Ruchi Yadav, Xavi Ruiz, and Eddy Arnold at Rutgers University, Valentine Courouble and Patrick Griffin at Scripps Research Institute, Florida, for critical discussion of the manuscript. M. N. and K. S. M. conceptualization; M. N. investigation; M. N. J.-P. A. and K. S. M. formal analysis; M. N. J.-P. A. and K. S. M. writing – review and editing; T. J. E. resources. This research was supported in part by the National Cancer Institute's National Cryo-EM Facility at the Frederick National Laboratory for Cancer Research under contract HSSN261200800001E. This work was supported by an NIH grant (R35 GM131860) to K. S. M. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Publisher Copyright:
© 2023 The Authors
PY - 2023/5
Y1 - 2023/5
N2 - The processing of the Coronavirus polyproteins pp1a and pp1ab by the main protease Mpro to produce mature proteins is a crucial event in virus replication and a promising target for antiviral drug development. Mpro cleaves polyproteins in a defined order, but how Mpro and/or the polyproteins determine the order of cleavage remains enigmatic due to a lack of structural information about polyprotein-bound Mpro. Here, we present the cryo-EM structures of SARS-CoV-2 Mpro in an apo form and in complex with the nsp7-10 region of the pp1a polyprotein. The complex structure shows that Mpro interacts with only the recognition site residues between nsp9 and nsp10, without any association with the rest of the polyprotein. Comparison between the apo form and polyprotein-bound structures of Mpro highlights the flexible nature of the active site region of Mpro, which allows it to accommodate ten recognition sites found in the polyprotein. These observations suggest that the role of Mpro in selecting a preferred cleavage site is limited and underscores the roles of the structure, conformation, and/or dynamics of the polyproteins in determining the sequence of polyprotein cleavage by Mpro.
AB - The processing of the Coronavirus polyproteins pp1a and pp1ab by the main protease Mpro to produce mature proteins is a crucial event in virus replication and a promising target for antiviral drug development. Mpro cleaves polyproteins in a defined order, but how Mpro and/or the polyproteins determine the order of cleavage remains enigmatic due to a lack of structural information about polyprotein-bound Mpro. Here, we present the cryo-EM structures of SARS-CoV-2 Mpro in an apo form and in complex with the nsp7-10 region of the pp1a polyprotein. The complex structure shows that Mpro interacts with only the recognition site residues between nsp9 and nsp10, without any association with the rest of the polyprotein. Comparison between the apo form and polyprotein-bound structures of Mpro highlights the flexible nature of the active site region of Mpro, which allows it to accommodate ten recognition sites found in the polyprotein. These observations suggest that the role of Mpro in selecting a preferred cleavage site is limited and underscores the roles of the structure, conformation, and/or dynamics of the polyproteins in determining the sequence of polyprotein cleavage by Mpro.
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U2 - 10.1016/j.jbc.2023.104697
DO - 10.1016/j.jbc.2023.104697
M3 - Article
C2 - 37044215
AN - SCOPUS:85158834874
SN - 0021-9258
VL - 299
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 5
M1 - 104697
ER -