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Infect Immun. May 1995; 63(5): 1914–1920.
PMCID: PMC173243

Cloning, expression, and nucleotide sequence of a Staphylococcus aureus gene (fbpA) encoding a fibrinogen-binding protein.

Abstract

Septicemia due to Staphylococcus aureus often begins as a focal infection (e.g., colonized wounds or catheters) from which the organism gains access to the bloodstream. On the basis of recent data from this laboratory, it is likely that S. aureus colonizes catheters and endothelium by using a fibrinogen-binding protein to mediate adhesion to fibrinogen-coated surfaces. To characterize the fibrinogen-reactive protein, we screened a lambda Zap library of S. aureus DB, a clinical isolate, for clones that were reactive with fibrinogen. Of 100,000 plaques screened, 3 were found to react with fibrinogen on immunoblots. Plasmid DNA prepared from clones 14, 30, and 36, upon digestion with EcoR1, which released the insert, revealed fragments of 4.6, 3.6, and 3.2 kb, respectively. To identify the cloned protein expressed in E. coli, cells were fractionated into periplasmic, membrane, and cytoplasmic fractions. Expression studies of clone 14, which comprised approximately two-thirds of the mature molecule, including the C terminus, revealed a 34-kDa fibrinogen-reactive protein in both the periplasmic and membrane fractions. This protein, designated FbpA, could be partially purified on a fibrinogen column. By using both clones 14 and 36 as templates, the complete DNA sequence of the fibrinogen-binding protein was obtained, yielding a molecule with a predicted size of 69,991 Da. Although sequence analysis revealed a high degree of homology with coagulase, there is a unique sequence of 11 amino acids that is not found in three known coagulases as well as two recently cloned fibrinogen-binding proteins. This unique sequence shares homology with a cell wall anchor motif found in other gram-positive surface proteins.

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Selected References

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  • Blake MS, Johnston KH, Russell-Jones GJ, Gotschlich EC. A rapid, sensitive method for detection of alkaline phosphatase-conjugated anti-antibody on Western blots. Anal Biochem. 1984 Jan;136(1):175–179. [PubMed]
  • Bodén MK, Flock JI. Evidence for three different fibrinogen-binding proteins with unique properties from Staphylococcus aureus strain Newman. Microb Pathog. 1992 Apr;12(4):289–298. [PubMed]
  • Bodén MK, Flock JI. Cloning and characterization of a gene for a 19 kDa fibrinogen-binding protein from Staphylococcus aureus. Mol Microbiol. 1994 May;12(4):599–606. [PubMed]
  • Bodén MK, Flock JI. Fibrinogen-binding protein/clumping factor from Staphylococcus aureus. Infect Immun. 1989 Aug;57(8):2358–2363. [PMC free article] [PubMed]
  • Cheung AL, Fischetti VA. Role of surface proteins in staphylococcal adherence to fibers in vitro. J Clin Invest. 1989 Jun;83(6):2041–2049. [PMC free article] [PubMed]
  • Cheung AL, Fischetti VA. The role of fibrinogen in staphylococcal adherence to catheters in vitro. J Infect Dis. 1990 Jun;161(6):1177–1186. [PubMed]
  • Cheung AL, Koomey JM, Butler CA, Projan SJ, Fischetti VA. Regulation of exoprotein expression in Staphylococcus aureus by a locus (sar) distinct from agr. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6462–6466. [PMC free article] [PubMed]
  • Cheung AL, Koomey JM, Lee S, Jaffe EA, Fischetti VA. Recombinant human tumor necrosis factor alpha promotes adherence of Staphylococcus aureus to cultured human endothelial cells. Infect Immun. 1991 Oct;59(10):3827–3831. [PMC free article] [PubMed]
  • Cheung AL, Krishnan M, Jaffe EA, Fischetti VA. Fibrinogen acts as a bridging molecule in the adherence of Staphylococcus aureus to cultured human endothelial cells. J Clin Invest. 1991 Jun;87(6):2236–2245. [PMC free article] [PubMed]
  • Cohen ML. Epidemiology of drug resistance: implications for a post-antimicrobial era. Science. 1992 Aug 21;257(5073):1050–1055. [PubMed]
  • Devereux J, Haeberli P, Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. [PMC free article] [PubMed]
  • Espersen F, Clemmensen I, Barkholt V. Isolation of Staphylococcus aureus clumping factor. Infect Immun. 1985 Sep;49(3):700–708. [PMC free article] [PubMed]
  • Feinberg AP, Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. [PubMed]
  • Fischetti VA, Jones KF, Manjula BN, Scott JR. Streptococcal M6 protein expressed in Escherichia coli. Localization, purification, and comparison with streptococcal-derived M protein. J Exp Med. 1984 Apr 1;159(4):1083–1095. [PMC free article] [PubMed]
  • Fischetti VA, Pancholi V, Schneewind O. Conservation of a hexapeptide sequence in the anchor region of surface proteins from gram-positive cocci. Mol Microbiol. 1990 Sep;4(9):1603–1605. [PubMed]
  • Garnier J, Osguthorpe DJ, Robson B. Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins. J Mol Biol. 1978 Mar 25;120(1):97–120. [PubMed]
  • Perlman D, Halvorson HO. A putative signal peptidase recognition site and sequence in eukaryotic and prokaryotic signal peptides. J Mol Biol. 1983 Jun 25;167(2):391–409. [PubMed]
  • Kaida S, Miyata T, Yoshizawa Y, Igarashi H, Iwanaga S. Nucleotide and deduced amino acid sequences of staphylocoagulase gene from Staphylococcus aureus strain 213. Nucleic Acids Res. 1989 Nov 11;17(21):8871–8871. [PMC free article] [PubMed]
  • Kaida S, Miyata T, Yoshizawa Y, Kawabata S, Morita T, Igarashi H, Iwanaga S. Nucleotide sequence of the staphylocoagulase gene: its unique COOH-terminal 8 tandem repeats. J Biochem. 1987 Nov;102(5):1177–1186. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • McDevitt D, Francois P, Vaudaux P, Foster TJ. Molecular characterization of the clumping factor (fibrinogen receptor) of Staphylococcus aureus. Mol Microbiol. 1994 Jan;11(2):237–248. [PubMed]
  • Neu HC. The crisis in antibiotic resistance. Science. 1992 Aug 21;257(5073):1064–1073. [PubMed]
  • Phonimdaeng P, O'Reilly M, Nowlan P, Bramley AJ, Foster TJ. The coagulase of Staphylococcus aureus 8325-4. Sequence analysis and virulence of site-specific coagulase-deficient mutants. Mol Microbiol. 1990 Mar;4(3):393–404. [PubMed]
  • Phonimdaeng P, O'Reilly M, O'Toole PW, Foster TJ. Molecular cloning and expression of the coagulase gene of Staphylococcus aureus 8325-4. J Gen Microbiol. 1988 Jan;134(1):75–83. [PubMed]
  • Sanger F, Nicklen S, Coulson AR. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. [PMC free article] [PubMed]
  • Schneewind O, Model P, Fischetti VA. Sorting of protein A to the staphylococcal cell wall. Cell. 1992 Jul 24;70(2):267–281. [PubMed]
  • So M, Crosa JH, Falkow S. Polynucleotide sequence relationships among Ent plasmids and the relationship between Ent and other plasmids. J Bacteriol. 1975 Jan;121(1):234–238. [PMC free article] [PubMed]
  • Towbin H, Staehelin T, Gordon J. Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Natl Acad Sci U S A. 1979 Sep;76(9):4350–4354. [PMC free article] [PubMed]
  • Usui Y. Biochemical properties of fibrinogen binding protein (clumping factor) of the staphylococcal cell surface. Zentralbl Bakteriol Mikrobiol Hyg A. 1986 Sep;262(3):287–297. [PubMed]
  • Voller A, Bidwell DE, Bartlett A. Enzyme immunoassays in diagnostic medicine. Theory and practice. Bull World Health Organ. 1976;53(1):55–65. [PMC free article] [PubMed]
  • Watanakunakorn C, Baird IM. Staphylococcus aureus bacteremia and endocarditis associated with a removable infected intravenous device. Am J Med. 1977 Aug;63(2):253–256. [PubMed]
  • Yother J, Briles DE. Structural properties and evolutionary relationships of PspA, a surface protein of Streptococcus pneumoniae, as revealed by sequence analysis. J Bacteriol. 1992 Jan;174(2):601–609. [PMC free article] [PubMed]
  • Yother J, White JM. Novel surface attachment mechanism of the Streptococcus pneumoniae protein PspA. J Bacteriol. 1994 May;176(10):2976–2985. [PMC free article] [PubMed]

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