title: Thiol-reactive reagents inhibits intracellular trafficking of human papillomavirus type 16 pseudovirions by binding to cysteine residues of major capsid protein L1.
abstract: BACKGROUND
  A human papillomavirus (HPV) virion is composed of capsid proteins L1 and L2. Several cysteine residues are located on L1 of various HPVs at markedly similar relative positions, suggesting their important functions. Although the authentic virions cannot be studied with cultured cells, surrogate pseudovirions consisting of capsid and reporter plasmid are available for studies dealing with infectivity.
  
  
  RESULTS
  HPV type16-pseudovirions (16PVs) were found to lose their infectivity after incubation with thiol-reactive reagents [biotin polyethyleneoxide iodoacetamide (BPEOIA), 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB), N-ethylmaleimide (NEM), 4-(N-maleimido)benzyl-trimethylammonium iodide (MBTA), and [2-(trimethylammonium)ethyl] methanethiosulfonate bromide (MTSET)]. A labelled streptavidin was detected to bind to the complex of BPEOIA and L1 of the 16PVs incubated with BPEOIA. The analysis of molecular mass of trypsin-fragments derived from the complex of the BPEOIA and L1 indicated that BPEOIA bound to at least C146, C225, and C229. No appreciable change of the 16PVs carrying DTNB or NEM was detected by sedimentation analysis or electron microscopy. The 16PVs carrying DTNB or NEM were able to bind to and enter HeLa cells but degraded before they reached the perinuclear region.
  
  
  CONCLUSIONS
  HPV16 L1 C146, C225, and C229 have free thiol, which are accessible to BPEOIA, DTNB, NEM, MBTA, and MTSET. Binding of DTNB or NEM to the thiols may cause conformational changes that result in the inhibition of the entry and trafficking of the 16PVs.
authors.affiliation: Center for Pathogen Genomics, National Institute of Infectious Diseases, 1-23-1 Toyama, Shinjuku-ku, Tokyo 162-8640, Japan. yishii@nih.go.jp
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Ishii
authors.forename: Kazunari
authors.initials: K
authors.lastname: Kondo
authors.forename: Tamae
authors.initials: T
authors.lastname: Matsumoto
authors.forename: Keiko
authors.initials: K
authors.lastname: Tanaka
authors.forename: Fumiko
authors.initials: F
authors.lastname: Shinkai-Ouchi
authors.forename: Ken'ichi
authors.initials: K
authors.lastname: Hagiwara
authors.forename: Tadahito
authors.initials: T
authors.lastname: Kanda
mesh_terms: D001692:Biological Transport / Q000187:drug effects; D002213:Capsid; D036022:Capsid Proteins / Q000737:chemistry / Q000378:metabolism*; D002461:Cell Line, Transformed; D003545:Cysteine / Q000378:metabolism; D052162:Human papillomavirus 16 / Q000378:metabolism*; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000737:chemistry / Q000378:metabolism*; D013438:Sulfhydryl Compounds / Q000378:metabolism; D013439:Sulfhydryl Reagents / Q000378:metabolism* / Q000494:pharmacology; D014771:Virion / Q000378:metabolism
chemical_list: D036022:Capsid Proteins; D009856:Oncogene Proteins, Viral; D013438:Sulfhydryl Compounds; D013439:Sulfhydryl Reagents; C063317:L1 protein, Human papillomavirus type 16; D003545:Cysteine