title: Expression in Escherichia coli of seven DNA fragments comprising the complete L1 and L2 open reading frames of human papillomavirus type 6b and localization of the 'common antigen' region.
abstract: Molecular cloning was used to express human papillomavirus type 6b (HPV-6b) antigens in Escherichia coli. Seven genomic DNA fragments of HPV-6b which together comprise the complete L1 and L2 open reading frames, known to code for capsid proteins, were cloned and expressed in E. coli as both beta-galactosidase and TrpE fusion proteins. Western blots of HPV-6b beta-galactosidase fusion proteins using 'genus-specific' antisera produced by immunization of rabbits with disrupted bovine papillomavirus type 1 (BPV-1) showed that polypeptides encoded by two DNA fragments from the mid portion of L1 of HPV-6b were cross-reactive. Only one of these two polypeptides reacted with antisera raised against disrupted HPV-1, directly demonstrating that this polypeptide contains the papillomavirus 'common antigen'. The cross-reactive region was confirmed by reversing antigen and antibody. Polyclonal antisera were raised against the seven HPV-6b beta-galactosidase fusion proteins and tested against BPV-1 virion proteins on Western blots. Only antiserum against the mid portion of L1 of HPV-6b reacted with the BPV-1 major capsid protein. HPV-6b fusion proteins were also used to test human sera for antibodies reactive in Western blots. Serum samples from 38 patients with documented HPV-6 infections and from 22 presumably uninfected controls were tested. Antibodies were not detected in any of the sera to any of the seven fusion proteins. HPV-6b beta-galactosidase fusion proteins are antigenic and can be used on Western blots to localize immunologically reactive sub-regions of proteins by reacting protein fragments with antisera from immunized animals. However, alternative methods will be required to detect anti-HPV antibodies in human sera.
authors: Department of Medicine, University of Rochester School of Medicine and Dentistry, New York 14642.
authors: D G
authors: DG
authors: Strike
authors: W
authors: W
authors: Bonnez
authors: R C
authors: RC
authors: Rose
authors: R C
authors: RC
authors: Reichman
mesh_terms: D000328:Adult; D000818:Animals; D000914:Antibodies, Viral / Q000032:analysis; D000942:Antigens, Bacterial / Q000235:genetics* / Q000276:immunology; D000956:Antigens, Viral / Q000235:genetics* / Q000276:immunology; D015153:Blotting, Western; D052688:Bovine papillomavirus 1 / Q000235:genetics / Q000276:immunology; D003001:Cloning, Molecular; D003062:Codon / Q000235:genetics*; D003429:Cross Reactions; D004279:DNA, Viral / Q000235:genetics*; D000939:Epitopes / Q000276:immunology; D004926:Escherichia coli / Q000235:genetics* / Q000276:immunology; D005786:Gene Expression Regulation; D006801:Humans; D027383:Papillomaviridae / Q000235:genetics* / Q000276:immunology; D012333:RNA, Messenger / Q000235:genetics*; D011817:Rabbits; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D011994:Recombinant Proteins / Q000235:genetics*; D001616:beta-Galactosidase / Q000235:genetics
chemical_list: D000914:Antibodies, Viral; D000942:Antigens, Bacterial; D000956:Antigens, Viral; D003062:Codon; D004279:DNA, Viral; D000939:Epitopes; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D001616:beta-Galactosidase