Isolation and characterization of genes encoding thermoactive and thermostable dextranases from two thermotolerant soil bacteria

Patrick M. Finnegan, Stevens M. Brumbley, Michael G. O'Shea, Helena Nevalainen, Peter L. Bergquist*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

19 Citations (Scopus)

Abstract

Thermotolerant Paenibacillus strain Dex70-1B and unidentified strain Dex70-34 produce thermoactive dextran-degrading enzymes. Plasmid-based genomic DNA libraries constructed from mixed bacterial cultures containing Dex70-1B or Dex70-34 were screened for the ability to confer dextranolytic activity at 70°C onto Escherichia coli. One gene, designated dex1, was isolated from each strain. The Dex70-1B and Dex70-34 dex1 gene sequences were non-identical, and encoded proteins containing 597 (Mr 68.6 kDa) and 600 amino acids (Mr 69.2 kDa), respectively. The Dex1 amino acid sequences were most similar to one another, and formed a new clade among the family 66 glycosyl hydrolase sequences. Expression of the Dex1 proteins in E. coli produced dextranolytic activity that converted ethanol-insoluble blue dextran into an ethanol-soluble form, suggestive of endodextranases (EC 3.2.1.11). Both enzymes were most active at about 60°C and pH 5.5, and retained more than 70% maximal activity after incubation at 57°C for 9.5 h in the absence of substrate.

Original languageEnglish
Pages (from-to)327-333
Number of pages7
JournalCurrent Microbiology
Volume49
Issue number5
DOIs
Publication statusPublished - Nov 2004

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