2m0a
From Proteopedia
Solution structure of MHV nsp3a
Structural highlights
Function[R1A_CVMA5] The papain-like proteinase 1 (PL1-PRO) and papain-like proteinase 2 (PL2-PRO) are responsible for the cleavages located at the N-terminus of the replicase polyprotein. In addition, PLP2 possesses a deubiquitinating/deISGylating activity and processes both 'Lys-48'- and 'Lys-63'-linked polyubiquitin chains from cellular substrates. Antagonizes innate immune induction of type I interferon by blocking the phosphorylation, dimerization and subsequent nuclear translocation of host IRF-3 (By similarity). The main proteinase 3CL-PRO is responsible for the majority of cleavages as it cleaves the C-terminus of replicase polyprotein at 11 sites. Recognizes substrates containing the core sequence [ILMVF]-Q-|-[SGACN]. Inhibited by the substrate-analog Cbz-Val-Asn-Ser-Thr-Leu-Gln-CMK. Also contains an ADP-ribose-1-phosphate (ADRP)-binding function (By similarity). Nsp7-nsp8 hexadecamer may possibly confer processivity to the polymerase, maybe by binding to dsRNA or by producing primers utilized by the latter (By similarity). Nsp9 is a ssRNA-binding protein (By similarity). Publication Abstract from PubMedCoronaviruses (CoVs) are positive-sense, single-stranded, enveloped RNA viruses that infect a variety of vertebrate hosts. The CoV nucleocapsid (N) protein contains two structurally independent RNA binding domains denoted the N-terminal domain (NTD) and the dimeric C-terminal (CTD) domain joined by a charged linker region rich in serine and arginine residues (SR-rich linker). An important goal in unraveling N function is to molecularly characterize N-protein interactions. Recent genetic evidence suggests that N interacts with nsp3a, a component of the viral replicase. Here, we present the solution NMR structure of MHV nsp3a and show, using isothermal titration calorimetry, that MHV N219, an N construct that extends into the SR-rich linker (residues 60-219), binds cognate nsp3a with high affinity (K(a) = 1.4 (+/-0.3) x 10(6) M(-1)). In contrast, neither N197, an N construct containing only the folded NTD (residues 60-197), or the CTD dimer (residues 260-380), bind nsp3a with detectable affinity. This indicates that the key nsp3a binding determinants localize to the SR-rich linker, a finding consistent with reverse genetics studies. NMR chemical shift perturbation analysis reveals that the N-terminal region of an MHV N SR-rich linker peptide (residues 198-230) binds to the acidic face of MHV nsp3a containing the acidic alpha2 helix with an affinity K(a) of 8.3 x 10(3) M(-1). These studies reveal that the SR-rich linker of MHV N is necessary but not sufficient to maintain this high-affinity binding to N. Solution structure of mouse hepatitis virus (MHV) nsp3a and determinants of the interaction with MHV nucleocapsid (N) protein.,Keane SC, Giedroc DP J Virol. 2013 Jan 9. PMID:23302895[1] From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine. References
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