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Hemolysin Gene Expression in the hns Knockout Mutant of Klebsiella pneumoniae UN Strain

Abstract

Hemolysin proteins are exotoxins produced by bacteria that can cause lysis of red blood cells in vitro. Hemolysin E is a pore-forming hemolysin, one of three types of hemolysin. Generally, hemolysin E is not expressed under standard growth conditions such as growth in LB medium at 37°C. However, in E. coli, when the hns gene, which encodes H-NS, a global regulatory DNA binding protein, is knocked out, hemolysin E gene is expressed. Because hns genes in E. coli and K. pneumoniae are similar, K. pneumoniae UN strain hns gene was knocked out in this research. The hns knockout mutant of K. pneumoniae UN is found to produce a 19 kDa hemolysin protein and shows hemolytic activity.

References

  • Albesa I (1989) Klebsiella pneumoniae haemolysin adsorption to red blood cells. J App Bacteriol 67, 263–266.

    Article  CAS  Google Scholar 

  • Bagley, ST, Seidler, RJ, Talbot HWJ, and Morrow JE (1978) Isolation of Klebsiellae from within living wood. Appl Environ Microbiol 36, 178–185.

    CAS  Google Scholar 

  • Barberis LI, Eraso AJ, Pájaro MC, and Albesa I (1986) Molecular weight determination and partial characterization of Klebsiella pneumoniae hemolysins. Can J Microbiol 32, 884–888.

    Article  CAS  Google Scholar 

  • Bertin P, Hommalis F, Krin E, Soutourina O, Tendeng C, Derzella S, and Danchin A (2001) H-NS and H-NS-like proteins in Gram-negative bacteria and their multiple role in the regulation of bacterial metabolism. Biochimie 83, 235–241.

    Article  CAS  Google Scholar 

  • Braun V and Focareta T (1991) Pore-forming bacterial protein hemolysins (cytolysins). Crit Rev Microbiol 18, 115–158.

    Article  CAS  Google Scholar 

  • Burnette WN (1981) Western blotting: Electrophoretic transfer of proteins from sodium dodecyl sulfate-polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem 112, 195–203.

    Article  CAS  Google Scholar 

  • Chuang Y-C, Su J-H, Lin C-N, and Chang M-C (2002) Cloning of a gene encoding a unique hemolysin from Klebsiella pneumoniae and its potential use as a species-specific gene probe. Microb Pathog 33, 1–6.

    Article  CAS  Google Scholar 

  • Datsenko KA and Wanner BL (2000) One-step inactivation of chromosomal genes in Escherichia coli K-12 using PCR products. Proc Natl Acad Sci 97, 6640–6645.

    Article  CAS  Google Scholar 

  • Donato GM and Kawula TH (1999) Phenotypic analysis of random hns mutations differentiate DNA-binding activity from properties of fimA promoter inversion modulation and bacterial motility. J Bacteriol 181, 941–948.

    CAS  Google Scholar 

  • Dorman CJ (2007) H-NS, the genome sentinel. Nature Rev Microbiol 5, 157–161.

    Article  CAS  Google Scholar 

  • Drancourt M, Bollet C, Carta A, and Rousselier P (2001) Phylogenetic analyses of Klebsiella species delineate Klebsiella and Raoultella gen. nov. with description of Raoultella ornithinolytica comb. nov., Raoultella terrigena comb. nov., and Raoultella planticola comb. nov. Int J Syst Evol Microbiol 51, 925–932.

    Article  CAS  Google Scholar 

  • Edberg SC, Piscitelli V, and Cartter M (1986) Phenotypic characteristics of coliform and noncoliform bacteria from a public water supply compared with regional and national clinical species. Appl Environ Microbiol 52, 474–478.

    CAS  Google Scholar 

  • Gomez-Gomez JM, Blazquez J, Baquero F, and Martinez JL (1996) Hns mutant unveils the presence of a latent hemolytic activity in Escherichia coli K-12. Mol Mocrobiol 19, 909–910.

    Article  CAS  Google Scholar 

  • Hanahan D (1983) Studies on transformation of Escherichia coli with plasmids. J Mol Biol 166, 557–580.

    Article  CAS  Google Scholar 

  • Highsmith AK and Jarvis WR (1985) Klebsiella pneumoniae: Selected virulence factors that contribute to pathogenicity. Infect Control 6, 75–77.

    CAS  Google Scholar 

  • Jiang L, He L, and Fountoulakis M (2004) Comparison of protein precipitation methods for sample preparation prior to proteomic analysis. J Chromatogr A 1023, 317–320.

    Article  CAS  Google Scholar 

  • John RT, Johnston HM, Seidman C, Garfinkel D, Gordon JK, Sha VK, and Brill WJ (1975) Biochemistry and genetics of Klebsiella pneumoniae mutant strains unable to fix N2. J Bacteriol 121, 759–765.

    Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during assembly of the head of bacteriophage T4. Nature 227, 680–685.

    Article  CAS  Google Scholar 

  • Matsen, JM, Spindler, JA, and Blosser, RO (1974) Characterization of Klebsiella isolates from natural receiving waters and comparison with human isolates. Appl Microbiol 28, 672–678.

    CAS  Google Scholar 

  • Moon D-K, Kim N, and Kim W-Y (2006) Reactivity of the antibodies against purified carp vitellogenin and a synthetic vitellogenin peptide. J Korean Soc Appl Biol Chem 49, 196–201.

    CAS  Google Scholar 

  • Neil RW, Stapleton MR, Norte VA, Roberts RE, Grafton J, and Green J (2004) Regulation of Escherichia coli hemolysin E expression by H-NS and Salmonella SlyA. J Bateriol 186, 1620–1628.

    Google Scholar 

  • Podschun R and Ullman U (1998) Klebsiella spp. as nosocomial pathogens: Epidemiology, taxonomy, typing methods, and pathogenicity factors. Clin Microbiol Rev 11, 589–603.

    CAS  Google Scholar 

  • Rowe GE and Welch RA (1994) Assays of hemolytic toxins. Methods Enzymol 235, 665–675.

    Google Scholar 

  • Shägger H (2006) Tricine-SDS-PAGE. Nature Protocols 1, 16–23.

    Article  Google Scholar 

  • Shägger H and Jagow VG (1991) Blue native electrophoresis for isolation of membrane protein complexes in enzymatically active form. Anal Biochem 199, 223–231.

    Article  Google Scholar 

  • Shi X and Bennett GN (1994) Plasmids bearing hfq and the hns-like gene stpA complement hns mutants in modulating arginine decarboxylase gene expression in Escherichia coli. J Bacteriol 21, 6769–6775.

    Google Scholar 

  • Tendeng C and Bertin PN (2003) H-NS in Gram-negative bacteria: A family of multifaceted proteins. Trends Microbiol 11, 511–518.

    Article  CAS  Google Scholar 

  • Wallace AJ, Stillman TJ, Atkins A, Jamieson SJ, Bullough PA, Green J, and Artymiuk PJ (2000) E. coli hemolysin E (HlyE, ClyA, SheA): X-ray crystal structure of the toxin and observation of membrane pores. Cell 2, 268–276.

    Google Scholar 

  • Ware DA (1972) Nitrogenase of Klebsiella pneumoniae: Interaction with viologen dyes as measured by acetylene reduction. J Biochem 130, 301–302

    CAS  Google Scholar 

  • Welch RA (1991) Pore-forming cytolysins of Gram-negative bacteria. Mol Mocrobiol 5, 521–528.

    Article  CAS  Google Scholar 

  • Wyborn NR, Clark A, Roberts RE, Jamieson SJ, Tzokov SB, Stillman TJ, Artymiuk PJ, Galen JE, Zhao L, Levine MM, and Green J (2004) Properties of haemolysin E (HlyE) from a pathogenic Escherichia coli avian isolate and studies of HlyE export. Microbiology 150, 1495–1505.

    Article  CAS  Google Scholar 

  • Yoch DC and Pengra RM (1966) Effect of amino acids on the nitrogenase system of Klebsiella pneumoniae. J Bacteriol 92, 618–622.

    CAS  Google Scholar 

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Correspondence to Woo-Yeon Kim.

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Han, J.S., Jang, IY., Ryu, HS. et al. Hemolysin Gene Expression in the hns Knockout Mutant of Klebsiella pneumoniae UN Strain. J. Korean Soc. Appl. Biol. Chem. 53, 401–406 (2010). https://doi.org/10.3839/jksabc.2010.062

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  • DOI: https://doi.org/10.3839/jksabc.2010.062

Key words

  • hemolysin
  • H-NS
  • Klebsiella pneumoniae UN