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The structure of the human respiratory syncytial virus M2-1 protein bound to the interaction domain of the phosphoprotein P defines the orientation of the complex

DOI: 10.1128/mBio.01554-18 DOI Help

Authors: Muniyandi Selvaraj (University of Leeds) , Kavestri Yegambaram (University of Leeds) , Eleanor J. A. A. Todd (University of Leeds) , Charles-adrien Richard (INRA, Université Paris-Saclay) , Rachel L. Dods (University of Leeds) , Georgia M. Pangratiou (University of Leeds) , Chi H. Trinh (University of Leeds) , Sophie L. Moul (University of Leeds) , James C. Murphy (University of Leeds) , Jamel Mankouri (University of Leeds) , Jean-françois Éléouët (INRA, Université Paris-Saclay) , John N. Barr (University of Leeds) , Thomas A. Edwards (University of Leeds)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Mbio , VOL 9

State: Published (Approved)
Published: November 2018
Diamond Proposal Number(s): 10305

Open Access Open Access

Abstract: Human respiratory syncytial virus (HRSV) is a negative-stranded RNA virus that causes a globally prevalent respiratory infection, which can cause life-threatening illness, particularly in the young, elderly, and immunocompromised. HRSV multiplication depends on replication and transcription of the HRSV genes by the virus-encoded RNA-dependent RNA polymerase (RdRp). For replication, this complex comprises the phosphoprotein (P) and the large protein (L), whereas for transcription, the M2-1 protein is also required. M2-1 is recruited to the RdRp by interaction with P and also interacts with RNA at overlapping binding sites on the M2-1 surface, such that binding of these partners is mutually exclusive. The molecular basis for the transcriptional requirement of M2-1 is unclear, as is the consequence of competition between P and RNA for M2-1 binding, which is likely a critical step in the transcription mechanism. Here, we report the crystal structure at 2.4 Å of M2-1 bound to the P interaction domain, which comprises P residues 90 to 110. The P90–110 peptide is alpha helical, and its position on the surface of M2-1 defines the orientation of the three transcriptase components within the complex. The M2-1/P interface includes ionic, hydrophobic, and hydrogen bond interactions, and the critical contribution of these contacts to complex formation was assessed using a minigenome assay. The affinity of M2-1 for RNA and P ligands was quantified using fluorescence anisotropy, which showed high-affinity RNAs could outcompete P. This has important implications for the mechanism of transcription, particularly the events surrounding transcription termination and synthesis of poly(A) sequences.

Subject Areas: Biology and Bio-materials, Medicine

Instruments: I02-Macromolecular Crystallography