title: Site-directed mutagenesis of cysteine residues of hepatitis B surface antigen. Analysis of two single mutants and the double mutant.
abstract: The structure of hepatitis B surface antigen (HBsAg) is mainly maintained by an intricate disulfide network responsible for most of its structural and antigenic properties. Characterization of three cysteine-replacement mutants of HBsAg has been performed by both structural and immunological methods. Replacement of Cys121 or Cys124 with serine results in mutant proteins that show diminished binding titres to both monoclonal antibodies and to a polyclonal serum, indicating that a structural change has taken place. Circular dichroism analysis shows that the substitution of either of these two residues also diminishes the helical content of the protein. However, the double mutant, in which both cysteine residues have been simultaneously changed, reverts the properties of the single mutations, and shows similar behaviour to the wild-type protein. Both the single and double cysteine mutants are efficiently glycosylated and secreted from Chinese hamster ovary cells and, in all cases, the mutant proteins assemble into spherical particles of similar buoyant density to both the wild-type and serum derived HBsAg.
authors.affiliation: Departamento de Bioquímica y Biología Molecular, Facultad de Ciencias Químicas, Universidad Complutense, Madrid, Spain.
authors.forename: B A
authors.initials: BA
authors.lastname: Antoni
authors.forename: I
authors.initials: I
authors.lastname: Rodríguez-Crespo
authors.forename: J
authors.initials: J
authors.lastname: Gómez-Gutiérrez
authors.forename: M
authors.initials: M
authors.lastname: Nieto
authors.forename: D
authors.initials: D
authors.lastname: Peterson
authors.forename: F
authors.initials: F
authors.lastname: Gavilanes
mesh_terms: D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D002942:Circular Dichroism; D006224:Cricetinae; D003412:Cricetulus; D003545:Cysteine / Q000235:genetics*; D006514:Hepatitis B Surface Antigens / Q000737:chemistry / Q000235:genetics* / Q000276:immunology; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation
chemical_list: D006514:Hepatitis B Surface Antigens; D003545:Cysteine