1887

Abstract

(previously named ) is a Gram-negative, anaerobic, fusiform bacterium that is a primary or secondary aetiological agent in periodontal disease in humans. expresses several putative virulence factors, including a sialidase; however, there has been no molecular genetic characterization of this enzyme. A sialidase clone (pHI-1) was screened from a total of 455 recombinant clones of a genomic DNA library using the 2′- (4-methylumbelliferyl)-α---acetylneuraminic acid (MUNeuAc) filter-paper spot assay. The sialidase gene ORF () consists of a 1395 bp coding sequence and encodes a protein with 465 amino acids with an overall molecular mass of 52 kDa. The sialidase does not have sequence similarity to any other bacterial sialidase. The entire sialidase ORF was expressed in . Furthermore, the sialidase was purified from the type strain of and from a recombinant clone, pHI-1 : 1, and was analysed using a non-denaturing gel, revealing that the enzyme preparations were respectively separated as two major bands and as a single band. Southern blot hybridization analysis revealed similar patterns of -hybridizing bands among clinical isolates of from periodontitis patients. This is the first study on the cloning and expression of a sialidase gene and the purification of the SiaHI enzyme from ATCC 43037 and recombinant .

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2003-12-01
2024-03-28
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References

  1. Arakawa S, Nakajima T, Ishikura H, Ichinose S, Ishikawa I, Tsuchida N. 2000; Novel apoptosis-inducing activity in Bacteroides forsythus : a comparative study with three serotypes of Actinobacillus actinomycetemcomitans . Infect Immun 68:4611–4615 [CrossRef]
    [Google Scholar]
  2. Berry A. M, Paton J. C, Glare E. M, Hansman D, Catcheside D. E. A. 1988; Cloning and expression of the pneumococcal neuraminidase gene in Escherichia coli . Gene 71:299–305 [CrossRef]
    [Google Scholar]
  3. Berry A. M, Lock R. A, Paton J. C. 1996; Cloning and characterization of nanB , a second Streptococcus pneumoniae neuraminidase gene, and purification of the NanB enzyme from recombinant Escherichia coli . J Bacteriol 178:4854–4860
    [Google Scholar]
  4. Bhown A. S, Bennett J. C. 1983; High-sensitivity sequence analysis of proteins recovered from sodium dodecyl sulfate gels. Methods Enzymol 91:450–455
    [Google Scholar]
  5. Briselden A. M, Moncla B. J, Stevens C. E, Hillier S. L. 1992; Sialidases (neuraminidases) in bacterial vaginosis and bacterial vaginosis-associated microflora. J Clin Microbiol 30:663–666
    [Google Scholar]
  6. Byers H. L, Homer K. A, Tarelli E, Beighton D. 1999; N -Acetylneuraminic acid transport by Streptococcus oralis strain AR3. J Med Microbiol 48:375–381 [CrossRef]
    [Google Scholar]
  7. Grossi S. G, Zambon J. J, Ho A. W, Koch G, Dunford R. G, Machtei E. E, Norderyd O. M, Genco R. J. 1994; Assessment of risk for periodontal disease.I. Risk indicators for attachment loss. . J Periodontol 65:260–267 [CrossRef]
    [Google Scholar]
  8. Grossi S. G, Genco R. J, Machtei E. E, Ho A. W, Koch G, Dunford R. G, Zambon J. J, Hausmann E. 1995; Assessment of risk for periodontal disease.II. Risk indicators for alveolar bone loss. J Periodontol 66:23–29 [CrossRef]
    [Google Scholar]
  9. Gyles C. L. 1986; Erysipelothrix . In Pathogenesis of Bacterial Infections in Animals p– 56 Edited by Gyles C. L., Thoen C. O. Ames, IA: Iowa State University Press;
    [Google Scholar]
  10. Holt S. C, Bramanti T. E. 1991; Factors in virulence expression and their role in periodontal disease pathogenesis. Crit Rev Oral Biol Med 2:177–281
    [Google Scholar]
  11. Honma K, Kuramitsu H. K, Genco R. J, Sharma A. 2001; Development of a gene inactivation system for Bacteroides forsythus : construction and characterization of a BspA mutant. Infect Immun 69:4686–4690 [CrossRef]
    [Google Scholar]
  12. Marmur J. A. 1961; A procedure for isolation of deoxyribonucleic acid from microorganisms. J Mol Biol 3:208–218 [CrossRef]
    [Google Scholar]
  13. McGregor J. A, French J. I, Richter R, Franco-Buff A, Johnson A, Hillier S, Judson F. N, Todd J. K. 1990; Antenatal microbiologic and maternal risk factors associated with prematurity. Am J Obstet Gynecol 163:1465–1473 [CrossRef]
    [Google Scholar]
  14. Moncla B. J, Braham P. 1989; Detection of sialidase (neuraminidase) activity in Actinomyces species by using 2′-(4-methylumbelliferyl)-α-d- N -acetylneuraminic acid in a filter paper spot test. J Clin Microbiol 27:182–184
    [Google Scholar]
  15. Moncla B. J, Braham P, Hillier S. L. 1990; Sialidase (neuraminidase) activity among Gram-negative anaerobic and capnophilic bacteria. J Clin Microbiol 28:422–425
    [Google Scholar]
  16. Myers R. W, Lee R. T, Lee Y. C, Thomas G. H, Reynolds L. W, Uchida Y. 1980; The synthesis of 4-methylumbelliferyl α-ketoside of N -acetylneuraminic acid and its use in a fluorometric assay for neuraminidase. Anal Biochem 101:166–174 [CrossRef]
    [Google Scholar]
  17. Roggentin P, Rothe B, Kaper J. B, Galen J, Lawrisuk L, Vimr E. R, Schauer R. 1989; Conserved sequences in bacterial and viral sialidases. Glycoconj J 6:349–353 [CrossRef]
    [Google Scholar]
  18. Saito T, Ishihara K, Kato T, Okuda K. 1997; Cloning, expression, and sequencing of a protease gene from Bacteroides forsythus ATCC 43037 in Escherichia coli . Infect Immun 65:4888–4891
    [Google Scholar]
  19. Sanger F, Nicklen S, Coulson A. R. 1977; DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A 74:5463–5467 [CrossRef]
    [Google Scholar]
  20. Sharma A, Sojar H. T, Glurich I, Honma K, Kuramitsu H. K, Genco R. J. 1998; Cloning, expression, and sequencing of a cell surface antigen containing a leucine-rich repeat motif from Bacteroides forsythus ATCC 43037. Infect Immun 66:5703–5710
    [Google Scholar]
  21. Southern E. M. 1975; Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol 98:503–517 [CrossRef]
    [Google Scholar]
  22. Tanner A. C, Strzempko M. N, Belsky C. A, McKinley G. A. 1985; API ZYM and API An-Ident reactions of fastidious oral Gram-negative species. J Clin Microbiol 22:333–335
    [Google Scholar]
  23. Tanner A. C. R, Listgarten M. A, Ebersole J. L, Strzempko M. N. 1986; Bacteroides forsythus sp.nov., a slow-growing, fusiform Bacteroides sp. from the human oral cavity. Int J Syst Bacteriol 36:213–221 [CrossRef]
    [Google Scholar]
  24. Vimr E. R, Troy F. A. 1985; Regulation of sialic acid metabolism in Escherichia coli : role of N -acetylneuraminate pyruvate-lyase. J Bacteriol 164:854–860
    [Google Scholar]
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