title: Expression of recombinant proteins on the surface of the coagulase-negative bacterium Staphylococcus xylosus.
abstract: An expression system to allow targeting of heterologous proteins to the cell surface of Staphylococcus xylosus, a coagulase-negative gram-positive bacterium, is described. The expression of recombinant gene fragments, fused between gene fragments encoding the signal peptide and the cell surface-binding regions of staphylococcal protein A, targets the resulting fusion proteins to the outer bacterial cell surface via the membrane-anchoring region and the highly charged cell wall-spanning region of staphylococcal protein A. The expression system was used to secrete fusion proteins containing sequences from a malaria blood-stage antigen and a streptococcal albumin-binding receptor to the cell surface of S. xylosus. Analysis of the recombinant cells by immunogold staining and immunofluorescence revealed that both the receptor and the malaria peptide were properly processed and exposed on the surface of the host cells. However, only approximately 40 to 50% of the recombinant cells were strongly stained with antiserum reactive with the albumin-binding receptor, while approximately 10 to 15% of the cells were stained with antiserum reactive with the malaria peptide. The incomplete staining of some of the cells suggests steric effects that make the recombinant fusion proteins inaccessible to the reactive antibodies because of variable cell wall structures. However, the results demonstrate for the first time that recombinant techniques can be used to express heterologous receptors and immunogens on the surface of gram-positive cells.
authors.affiliation: Department of Biochemistry and Biotechnology, Royal Institute of Technology, Stockholm, Sweden.
authors.forename: M
authors.initials: M
authors.lastname: Hansson
authors.forename: S
authors.initials: S
authors.lastname: Ståhl
authors.forename: T N
authors.initials: TN
authors.lastname: Nguyen
authors.forename: T
authors.initials: T
authors.lastname: Bächi
authors.forename: A
authors.initials: A
authors.lastname: Robert
authors.forename: H
authors.initials: H
authors.lastname: Binz
authors.forename: A
authors.initials: A
authors.lastname: Sjölander
authors.forename: M
authors.initials: M
authors.lastname: Uhlén
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000953:Antigens, Protozoan / Q000096:biosynthesis / Q000235:genetics*; D001483:Base Sequence; D002462:Cell Membrane / Q000378:metabolism*; D003030:Coagulase / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics; D005798:Genes, Bacterial; D005822:Genetic Vectors; D016253:Microscopy, Immunoelectron; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides; D010957:Plasmids; D010963:Plasmodium falciparum / Q000235:genetics*; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals / Q000096:biosynthesis / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis*; D013205:Staphylococcal Protein A / Q000096:biosynthesis / Q000235:genetics*; D013210:Staphylococcus / Q000201:enzymology / Q000235:genetics* / Q000648:ultrastructure
chemical_list: D000953:Antigens, Protozoan; D003030:Coagulase; D009838:Oligodeoxyribonucleotides; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; D013205:Staphylococcal Protein A