title: Hydrophobicity engineering to facilitate surface display of heterologous gene products on Staphylococcus xylosus.
abstract: Protein engineering has been employed to investigate the effect of specific amino acid changes on the targeting of heterologous proteins to the outer cell surface of the Gram-positive bacterium Staphylococcus xylosus. Three different variants, corresponding to a 101 amino acid region of the major glycoprotein (G protein) of human respiratory syncytial virus (RSV), were generated in which multiple hydrophobic phenylalanine residues were either substituted or deleted. The different G protein fragments were expressed as one part of recombinant receptors designed for surface display on S. xylosus cells. The engineered variants of the RSV G protein hybrid receptors were, in contrast to a non-engineered fragment, efficiently targeted to the outer cell surface of recombinant S. xylosus cells as determined by different methods, including fluorescence-activated cell sorting. In addition, immunization of mice with live recombinant S. xylosus demonstrated that surface exposure was required to generate receptor-specific antibodies. The present strategy of hydrophobic engineering should be of general interest in surface-display applications and for secretion of proteins otherwise difficult to translocate through host cell membranes.
authors.affiliation: Centre d'Immunologie Pierre Fabre, Saint-Julien en Genevois, France.
authors.forename: T N
authors.initials: TN
authors.lastname: Nguyen
authors.forename: M H
authors.initials: MH
authors.lastname: Gourdon
authors.forename: M
authors.initials: M
authors.lastname: Hansson
authors.forename: A
authors.initials: A
authors.lastname: Robert
authors.forename: P
authors.initials: P
authors.lastname: Samuelson
authors.forename: C
authors.initials: C
authors.lastname: Libon
authors.forename: C
authors.initials: C
authors.lastname: Andréoni
authors.forename: P A
authors.initials: PA
authors.lastname: Nygren
authors.forename: H
authors.initials: H
authors.lastname: Binz
authors.forename: M
authors.initials: M
authors.lastname: Uhlén
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D003124:Colorimetry; D017931:DNA Primers; D005260:Female; D005434:Flow Cytometry; D005822:Genetic Vectors; D015707:HN Protein; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology; D051379:Mice; D008807:Mice, Inbred BALB C; D008969:Molecular Sequence Data; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics; D012136:Respiratory Syncytial Viruses / Q000235:genetics*; D013210:Staphylococcus / Q000235:genetics*; D014614:Vaccines, Synthetic / Q000235:genetics / Q000276:immunology; D014759:Viral Envelope Proteins; D014764:Viral Proteins / Q000235:genetics* / Q000276:immunology; D014765:Viral Vaccines / Q000235:genetics / Q000276:immunology
chemical_list: D017931:DNA Primers; D015707:HN Protein; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; D014614:Vaccines, Synthetic; D014759:Viral Envelope Proteins; D014764:Viral Proteins; D014765:Viral Vaccines; C116204:attachment protein G