Macrodomain Mac1 of SARS-CoV-2 nonstructural protein 3 hydrolyzes diverse ADP-ribosylated substrates

Authors

• C Chea
• DY Lee
• J Kato
• H Ishiwata-Endo
• J Moss

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for a global pandemic that resulted in more than 7 million deaths worldwide. The virus encodes non-structural protein 3 that contains macrodomains, i.e., Mac1, Mac2, and Mac3, with potential effects on host cells. Mac1 has been shown to disrupt ADP-ribosylation pathways in mammalian cells and increase SARS-CoV-2 virulence; ADP-ribosylation results from the transfer of the ADP-ribose moiety of NAD+ to various acceptors. Some viral macrodomains cleave the ADP-ribose-acceptor bond, generating free ADP-ribose and interfering with ADP-ribose regulatory pathways in host cells. Here, we examined the potential hydrolytic activities of SARS-CoV-2 Mac1, 2, and 3 on substrates containing ADP-ribose. Mac1 cleaved α-NAD+, but not β-NAD+, to generate ADP-ribose and nicotinamide, consistent with stereospecificity at the C-1” bond. Mac1, but not Mac2 and Mac3, also hydrolyzed O-acetyl-ADP- ribose and ADP-ribose-1”-phosphate. Thus, Mac1 hydrolyzes ADP-ribosylated N- and O- functional groups. However, Mac1 did not cleave α-ADP-ribose-(arginine) and ADP-ribose-(serine)histone H3 peptide as was the case for mammalian ADP-ribose-acceptor hydrolases (ARHs) ARH1 and ARH3 respectively. Unlike ARH1 and ARH3 that catalyze similar reactions to macrodomains, Mac1 did not require Mg2+ for optimal activity. We conclude that SARS-CoV-2 Mac1 may exert anti-viral activity by reversing host-mediated ADP-ribosylation pathways and thus may identify potential SARS-CoV-2 therapeutic targets.

Scientific Focus Area: Virology

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