| Summary: | <p>Abstract</p> <p>Background</p> <p>Thermostable enzymes from thermophiles have attracted extensive studies. In this investigation, a nuclease-encoding gene (designated as <it>GBSV1-NSN</it>) was obtained from a thermophilic bacteriophage GBSV1 for the first time.</p> <p>Results</p> <p>After recombinant expression in <it>Escherichia coli</it>, the purified GBSV1-NSN exhibited non-specific nuclease activity, being able to degrade various nucleic acids, including RNA, single-stranded DNA and double-stranded DNA that was circular or linear. Based on sequence analysis, the nuclease shared no homology with any known nucleases, suggesting that it was a novel nuclease. The characterization of the recombinant GBSV1-NSN showed that its optimal temperature and pH were 60°C and 7.5, respectively. The results indicated that the enzymatic activity was inhibited by enzyme inhibitors or detergents, such as ethylene diamine tetraacetic acid, citrate, dithiothreitol, β-mercaptoethanol, guanidine hydrochloride, urea and SDS. In contrast, the nuclease activity was enhanced by TritonX-100, Tween-20 or chaps to approximately 124.5% – 141.6%. The <it>K</it><sub>m </sub>of GBSV1-NSN nuclease was 231, 61 and 92 μM, while its <it>k</it><sub>cat </sub>was 1278, 241 and 300 s<sup>-1 </sup>for the cleavage of dsDNA, ssDNA and RNA, respectively.</p> <p>Conclusion</p> <p>Our study, therefore, presented a novel thermostable non-specific nuclease from thermophilic bacteriophage and its overexpression and purification for scientific research and applications.</p>
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