title: Re-analysis of cryoEM data on HCV IRES bound to 40S subunit of human ribosome integrated with recent structural information suggests new contact regions between ribosomal proteins and HCV RNA.
abstract: In this study, we combine available high resolution structural information on eukaryotic ribosomes with low resolution cryo-EM data on the Hepatitis C Viral RNA (IRES) human ribosome complex. Aided further by the prediction of RNA-protein interactions and restrained docking studies, we gain insights on their interaction at the residue level. We identified the components involved at the major and minor contact regions, and propose that there are energetically favorable local interactions between 40S ribosomal proteins and IRES domains. Domain II of the IRES interacts with ribosomal proteins S5 and S25 while the pseudoknot and the downstream domain IV region bind to ribosomal proteins S26, S28 and S5. We also provide support using UV cross-linking studies to validate our proposition of interaction between the S5 and IRES domains II and IV. We found that domain IIIe makes contact with the ribosomal protein S3a (S1e). Our model also suggests that the ribosomal protein S27 interacts with domain IIIc while S7 has a weak contact with a single base RNA bulge between junction IIIabc and IIId. The interacting residues are highly conserved among mammalian homologs while IRES RNA bases involved in contact do not show strict conservation. IRES RNA binding sites for S25 and S3a show the best conservation among related viral IRESs. The new contacts identified between ribosomal proteins and RNA are consistent with previous independent studies on RNA-binding properties of ribosomal proteins reported in literature, though information at the residue level is not available in previous studies.
authors.affiliation: Molecular Biophysics Unit. Indian Institute of Science, Bangalore, India; Present address: Science and Technology Facilities Council, RAL, Harwell, Didcot, UK.
authors.forename: Agnel Praveen
authors.initials: AP
authors.lastname: Joseph
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India.
authors.forename: Prasanna
authors.initials: P
authors.lastname: Bhat
authors.affiliation: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore, India.
authors.forename: Saumitra
authors.initials: S
authors.lastname: Das
authors.affiliation: Molecular Biophysics Unit. Indian Institute of Science, Bangalore, India.
authors.forename: Narayanaswamy
authors.initials: N
authors.lastname: Srinivasan
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D020285:Cryoelectron Microscopy / Q000379:methods*; D016174:Hepacivirus / Q000737:chemistry / Q000235:genetics*; D006801:Humans; D062105:Molecular Docking Simulation; D009690:Nucleic Acid Conformation; D017433:Protein Structure, Secondary; D012367:RNA, Viral / Q000737:chemistry* / Q000378:metabolism; D054658:Ribosome Subunits, Large / Q000737:chemistry* / Q000378:metabolism
chemical_list: D012367:RNA, Viral
keywords: Cryo electron microscopy; Hepatitis C; Protein modeling; RNA-Protein interactions