title: Differentiation-dependent interpentameric disulfide bond stabilizes native human papillomavirus type 16.
abstract: Genetic and biochemical analyses of human papillomavirus type 16 (HPV16) capsids have shown that certain conserved L1 cysteine residues are critical for capsid assembly, integrity, and maturation. Since previous studies utilized HPV capsids produced in monolayer culture-based protein expression systems, the ascribed roles for these cysteine residues were not placed in the temporal context of the natural host environment for HPV, stratifying and differentiating human tissue. Here we extend upon previous observation, that HPV16 capsids mature and become stabilized over time (10-day to 20-day) in a naturally occurring tissue-spanning redox gradient, by identifying temporal roles for individual L1 cysteine residues. Specifically, the C175S substitution severely undermined wild-type titers of the virus within both 10 and 20-day tissue, while C428S, C185S, and C175,185S substitutions severely undermined wild-type titers only within 20-day tissue. All mutations led to 20-day virions that were less stable than wild-type and failed to form L1 multimers via nonreducing SDS-PAGE. Furthermore, Optiprep-fractionated 20-day C428S, C175S, and C175,185S capsids appeared permeable to endonucleases in comparison to wild-type and C185S capsids. Exposure to an oxidizing environment failed to enhance infectious titers of any of the cysteine mutants over time as with wild-type. Introduction of these cys mutants results in failure of the virus to mature.
authors.affiliation: Department of Microbiology and Immunology, The Pennsylvania State University College of Medicine, Hershey, Pennsylvania, United States of America.
authors.forename: Michael J
authors.initials: MJ
authors.lastname: Conway
authors.forename: Linda
authors.initials: L
authors.lastname: Cruz
authors.forename: Samina
authors.initials: S
authors.lastname: Alam
authors.forename: Neil D
authors.initials: ND
authors.lastname: Christensen
authors.forename: Craig
authors.initials: C
authors.lastname: Meyers
mesh_terms: D015153:Blotting, Western; D002213:Capsid / Q000378:metabolism; D036022:Capsid Proteins / Q000737:chemistry / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D004279:DNA, Viral / Q000378:metabolism; D004220:Disulfides / Q000378:metabolism*; D016679:Genome, Viral / Q000235:genetics; D019803:Glutathione Disulfide / Q000378:metabolism / Q000494:pharmacology; D052162:Human papillomavirus 16 / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D006801:Humans; D007231:Infant, Newborn; D008297:Male; D009154:Mutation / Q000235:genetics; D009856:Oncogene Proteins, Viral / Q000737:chemistry / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D055503:Protein Multimerization; D014771:Virion / Q000235:genetics / Q000378:metabolism / Q000472:pathogenicity
chemical_list: D036022:Capsid Proteins; D004279:DNA, Viral; D004220:Disulfides; D009856:Oncogene Proteins, Viral; C063317:L1 protein, Human papillomavirus type 16; D019803:Glutathione Disulfide