Related ArticlesSolution structure of mouse hepatitis virus (MHV) nsp3a and determinants of the interaction with MHV nucleocapsid (N) protein.
J Virol. 2013 Jan 9;
Authors: Keane SC, Giedroc DP
Abstract
Coronaviruses (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) × 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 ?2 helix with an affinity K(a) of 8.3 × 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.
PMID: 23302895 [PubMed - as supplied by publisher]
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Solution structure of mouse hepatitis virus (MHV) nsp3a and determinants of the interaction with MHV nucleocapsid (N) protein.
Linear Mode
