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Detecting cellulase and esterase enzyme activities encoded by novel genes present in environmental DNA libraries

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Abstract

A genomic DNA library was made from the alkaliphilic cellulase-producing Bacillus agaradhaerans in order to prove our technologies for gene isolation prior to using them with samples of DNA isolated directly from environmental samples. Clones expressing a cellulase activity were identified and sequenced. A new cellulase gene was identified. Genomic DNA libraries were then made from DNA isolated directly from the Kenyan soda lakes, Lake Elmenteita and Crater Lake. Crater Lake clones expressing a cellulase activity and Lake Elmenteita clones expressing a lipase/esterase activity were identified and sequenced. These were encoded by novel genes as judged by DNA sequence comparisons. Genomic DNA libraries were also made from laboratory enrichment cultures of Lake Nakuru and Lake Elmenteita samples. Selective enrichment cultures were grown in the presence of carboxymethylcellulose (CMC) and olive oil. A number of new cellulase and lipase/esterase genes were discovered in these libraries. Cellulase-positive clones from Lake Nakuru were isolated at a frequency of 1 in 15,000 from a library made from a CMC enrichment as compared to 1 in 60,000 from a minimal medium enrichment. Esterase/lipase-positive clones from Lake Elmenteita were isolated with a frequency of 1 in 30,000 from a library made from an olive-oil enrichment as compared to 1 in 100,000 from an environmental library.

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References

  • Aguilar A, Ingemansson T, Magnien E (1998) Extremophile microorganisms as cell factories: support from the European Union. Extremophiles 2:367–373

    Article  CAS  PubMed  Google Scholar 

  • Amann RI, Ludwig W, Schleifer KH (1995) Phylogenetic identification and in situ detection of individual microbial cells without cultivation. Microbiol Rev 59:143–169

    CAS  PubMed  Google Scholar 

  • Chen WP, Kuo TT (1993) A simple and rapid method for the preparation of gram-negative bacterial genomic DNA. Nucleic Acids Res 21:2260

    CAS  PubMed  Google Scholar 

  • Davies GJ, Dauter M, Brzozowski AM, Bjoernvad ME, Andersen KV, Schuelein M (1998) Structure of the Bacillus agaradherans family 5 endoglucanase at 1.6-A and its cellobiose complex at 2.0-A resolution. Biochemistry 37:1926–1932

    Article  CAS  PubMed  Google Scholar 

  • Duckworth AW, Grant WD, Jones BE, Steenbergen R (1996) Phylogenetic diversity of soda lake alkaliphiles. FEMS Microbiol Ecol 19:181–191

    Article  CAS  Google Scholar 

  • Grant S, Grant WD, Jones BE, Kato C, Li L (1999) Novel archaeal phylotypes from an East African saltern. Extremophiles 3:139–145

    Article  CAS  PubMed  Google Scholar 

  • Henne A, Schmitz RA, Bömeke M, Gottschalk G, Daniel R (2000) Screening of environmental DNA libraries for the presence of genes conferring lipolytic activity on Escherichia coli. Appl Environ Microbiol 66:3113–3116

    CAS  PubMed  Google Scholar 

  • Horikoshi K (1999) Alkaliphiles: some applications of their products for biotechnology. Microbiol Mol Biol Revs 63:735–750

    CAS  Google Scholar 

  • Jones BE, Grant WD, Collins NC, Mwatha WE (1994) Alkaliphiles: diversity and identification. In: Priest FG, Ramos-Cormenzana A, Tindal BJ (eds) Bacterial diversity and systematics. Plenum, New York, pp195–230

  • Jones BE, Grant WD, Duckworth AW, Owenson GG (1998) Microbial diversity of soda lakes. Extremophiles 2:191–200

    Article  CAS  PubMed  Google Scholar 

  • Margesin R, Schinner F (2001) Potential of halotolerant and halophilic microorganisms for biotechnology. Extremophiles 5:73–83

    Article  CAS  PubMed  Google Scholar 

  • Marrs B, Delagrave S, Murphy D (1999) Novel appproaches for discovering industrial enzymes. Curr Opin Microbiol 2:241–245

    Article  CAS  PubMed  Google Scholar 

  • Pitcher DG, Saunders NA, Owen RJ (1989) Rapid extraction of bacterial genomic DNA with guanidium thiocyanate. Lett Appl Microbiol 8:151–156

    CAS  Google Scholar 

  • Rees H (2002) Environmental gene screening. PhD thesis, University of Leicester

  • Takami H, Nakasone K, Takaki Y, Maeno G, Sasaki R, Masui N, Fuji F, Hirama C, Nakamura Y, Ogasawara N, Kuhara S, Horikoshi K (2000) Complete genome sequence of the alkaliphilic bacterium Bacillus halodurans and genomic sequence comparison with Bacillus subtilis. Nucleic Acids Res 28:4317–4331

    CAS  PubMed  Google Scholar 

  • Teather RM, Wood PJ (1982) Use of Congo red-polysaccharide interactions in enumeration and characterisation of cellulolytic bacteria from the bovine rumen. Appl Environ Microbiol 43:777–780

    CAS  PubMed  Google Scholar 

  • Upton C, Buckley JT (1995) A new family of lipolytic enzymes? Trends Biochem Sci 20:178–179

    Google Scholar 

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Correspondence to Shaun Heaphy.

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Communicated by K. Horikoshi

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Rees, H.C., Grant, S., Jones, B. et al. Detecting cellulase and esterase enzyme activities encoded by novel genes present in environmental DNA libraries. Extremophiles 7, 415–421 (2003). https://doi.org/10.1007/s00792-003-0339-2

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  • DOI: https://doi.org/10.1007/s00792-003-0339-2

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