Mutation of interfacial residues disrupts subunit folding and particle assembly of Physalis mottle tymovirus.
Virus-like particles (VLPs) serve as excellent model systems to identify the pathways of virus assembly. To gain insights into the assembly mechanisms of the Physalis mottle tymovirus (PhMV), six interfacial residues, identified based on the crystal structure of the native and recombinant capsids, were targeted for mutagenesis. The Q37E, Y67A, R68Q, D83A, I123A, and S145A mutants of the PhMV recombinant coat protein (rCP) expressed in Escherichia coli were soluble. However, except for the S145A mutant, which assembled into VLPs similar to that of wild type rCP capsids, all the other mutants failed to assemble into VLPs. Furthermore, the purified Q37E, Y67A, R68Q, D83A, and I123A rCP mutants existed essentially as partially folded monomers as revealed by sucrose density gradient analysis, circular dichroism, fluorescence, thermal, and urea denaturation studies. The rCP mutants locked into such conformations probably lack the structural signals/features that would allow them to assemble into capsids. Thus, the mutation of residues involved in inter-subunit interactions in PhMV disrupts both subunit folding and particle assembly.
Umashankar
Mahadevaiah
M
Department of Biochemistry, Indian Institute of Science, Bangalore 560 012, India.
Murthy
Mathur R N
MR
Savithri
Handanahal S
HS
D019943:Amino Acid Substitution; D001483:Base Sequence; D017931:DNA Primers; D006860:Hydrogen Bonding; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D032321:Physalis / Q000235:genetics* / Q000821:virology; D017510:Protein Folding; D017433:Protein Structure, Secondary; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D017806:Tymovirus / Q000235:genetics*; D014764:Viral Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism
D017931:DNA Primers; D021122:Protein Subunits; D011994:Recombinant Proteins; D014764:Viral Proteins