title: Formation of covalently modified folding intermediates of simian virus 40 Vp1 in large T antigen-expressing cells.
abstract: The folding and pentamer assembly of the simian virus 40 (SV40) major capsid protein Vp1, which take place in the infected cytoplasm, have been shown to progress through disulfide-bonded Vp1 folding intermediates. In this report, we further demonstrate the existence of another category of Vp1 folding or assembly intermediates: the nonreducible, covalently modified mdVp1s. These species were present in COS-7 cells that expressed a recombinant SV40 Vp1, Vp1ΔC, through plasmid transfection. The mdVp1s persisted under cell and lysate treatment and SDS-PAGE conditions that are expected to have suppressed the formation of artifactual disulfide cross-links. As shown through a pulse-chase analysis, the mdVp1s were derived from the newly synthesized Vp1ΔC in the same time frame as Vp1's folding and oligomerization. The apparent covalent modifications occurred in the cytoplasm within the core region of Vp1 and depended on the coexpression of the SV40 large T antigen (LT) in the cells. Analogous covalently modified species were found with the expression of recombinant polyomavirus Vp1s and human papillomavirus L1s in COS-7 cells. Furthermore, the mdVp1s formed multiprotein complexes with LT, Hsp70, and Hsp40, and a fraction of the largest mdVp1, md4, was disulfide linked to the unmodified Vp1ΔC. Both mdVp1 formation and most of the multiprotein complex formation were blocked by a Vp1 folding mutation, C87A-C254A. Our observations are consistent with a role for LT in facilitating the folding process of SV40 Vp1 by stimulating certain covalent modifications of Vp1 or by recruiting certain cellular proteins.
authors: Department of Molecular, Cell and Developmental Biology and Molecular Biology Institute, University of California at Los Angeles, Los Angeles, California, USA.
authors: Marika
authors: M
authors: Watanabe
authors: Ellen
authors: E
authors: Phamduong
authors: Chu-Han
authors: CH
authors: Huang
authors: Noriko
authors: N
authors: Itoh
authors: Janie
authors: J
authors: Bernal
authors: Akira
authors: A
authors: Nakanishi
authors: Kathleen
authors: K
authors: Rundell
authors: Ole
authors: O
authors: Gjoerup
authors: Harumi
authors: H
authors: Kasamatsu
mesh_terms: D000818:Animals; D000957:Antigens, Viral, Tumor / Q000235:genetics / Q000378:metabolism*; D019556:COS Cells; D036022:Capsid Proteins / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D002460:Cell Line; D002522:Chlorocebus aethiops; D006801:Humans; D027601:Polyomavirus Infections / Q000821:virology*; D017510:Protein Folding; D013539:Simian virus 40 / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000957:Antigens, Viral, Tumor; D036022:Capsid Proteins; C062481:VP1 protein, polyomavirus