Architecture of physalis mottle tymovirus as probed by monoclonal antibodies and cross-linking studies.
Physalis mottle tymovirus (previously named belladonna mottle virus, Iowa strain) RNA was cross-linked to its coat protein by exposure of the intact virus to ultraviolet light. The site of cross-linking of the coat protein with the RNA was identified as Lys-10 by sequencing the oligonucleotide-linked tryptic peptide obtained upon HPLC separation subsequent to enzymatic digestion of the cross-linked and dissociated virus. Three monoclonal antibodies PA3B2, PB5G9, and PF12C9, obtained using denatured coat protein as antigen, cross-reacted effectively with the intact virus indicating that the epitopes recognized by these monoclonals are on the surface of the virus. Using the peptides generated by digestion with CNBr, clostripain, V-8 protease, or trypsin and a recombinant protein lacking the N-terminal 21 residues expressed from a cDNA clone, it was shown that PA3B2 recognizes the sequence 22-36 on the coat protein while PB5G9 and PF12C9 recognize region 75-110. These results suggest that Lys-10 is one of the specific sites through which the RNA interacts in the intact virus. The polypeptide segment (region 22-36) following this buried portion as well as the epitope within the region 75-110 are exposed in the intact virus. These observations are consistent with the canonical beta-barrel structure observed in certain other plant viruses.
Kekuda
R
R
Department of Biochemistry, Indian Institute of Science, Bangalore.
Karande
A A
AA
Jacob
A N
AN
Savithri
H S
HS
D000595:Amino Acid Sequence; D000911:Antibodies, Monoclonal; D015153:Blotting, Western; D002213:Capsid / Q000737:chemistry* / Q000302:isolation & purification; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000737:chemistry*; D010446:Peptide Fragments / Q000302:isolation & purification; D010449:Peptide Mapping; D017433:Protein Structure, Secondary; D012367:RNA, Viral / Q000737:chemistry / Q000302:isolation & purification; D014357:Trypsin
D000911:Antibodies, Monoclonal; D010446:Peptide Fragments; D012367:RNA, Viral; D014357:Trypsin