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    • Keywords


      Geobacillus; restriction endonuclease; subtype IIS, IIC, IIG; thermostability

    • Abstract


      Screening of extreme environments in search for novel microorganisms may lead to the discovery of robust enzymes with either new substrate specificities or thermostable equivalents of those already found in mesophiles, better suited for biotechnology applications. Isolates from Iceland geysers’ biofilms, exposed to a broad range of temperatures, from ambient to close to water boiling point, were analysed for the presence of DNA-interacting proteins, including restriction endonucleases (REases). GeoICI, a member of atypical Type IIS REases, is the most thermostable isoschizomer of the prototype BbvI, recognizing/cleaving 5′-GCAGC(N8/12)-3′ DNA sequences. As opposed to the unstable prototype, which cleaves DNA at 30°C, GeoICI is highly active at elevated temperatures, up to 73°C and over a very wide salt concentration range. Recognition/cleavage sites were determined by: (i) digestion of plasmid and bacteriophage lambda DNA (λ); (ii) cleavage of custom PCR substrates, (iii) run-off sequencing of GeoICI cleavage products and (iv) shotgun cloning and sequencing of λ DNA fragmented with GeoICI. Geobacillus sp. genomic DNA was PCR-screened for the presence of other specialized REases-MTases and as a result, another putative REase-MTase, GeoICII, related to the Thermus sp. family of bifunctional REases-methyltransferases (MTases) was detected.

    • Author Affiliations


      Joanna Zebrowska1 Olga Zołnierkiewicz1 Marta A Skowron2 Agnieszka Zylicz-Stachula1 Joanna Jezewska-Frackowiak1 Piotr M Skowron1

      1. Department of Molecular Biotechnology, Institute for Environment and Human Health Protection, Faculty of Chemistry,University of Gdansk, Wita Stwosza 63, 80-308 Gdansk, Poland
      2. Department of Molecular Biology, Faculty of Biology, University of Gdansk, Wita Stwosza 63, 80-308 Gdansk, Poland
    • Dates

  • Journal of Biosciences | News

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      Posted on July 25, 2019

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