High-level production and purification of a fully active recombinant dextransucrase from Leuconostoc mesenteroides NRRL B-512F |
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Authors: | Moulis Claire Arcache Audrey Escalier Pierre-Claude Rinaudo Marguerite Monsan Pierre Remaud-Simeon Magali Potocki-Veronese Gabrielle |
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Affiliation: | Laboratoire de Biotechnologies - Bioprocédés, UMR CNRS 5504, UMR INRA 792, INSA, Toulouse, France. |
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Abstract: | Recombinant expression of the dextransucrase dsrS gene by Escherichia coli was optimized to produce 5850 U L(-1) (culture) of DSR-S, corresponding to a 30-fold increase compared with previous studies. Rational deletions of the signal peptide, the beginning of the variable region and the last four repeats of the C-terminal end caused no loss of activity. This new variant successfully purified was remarkably stable. With a k(cat) of 584 s(-1), it is the most efficient recombinant glucansucrase described to date. The synthesized polymer possesses more than 95% of alpha-1,6 links, like the dextran produced by the native enzyme, and innovative gel properties were obtained. |
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Keywords: | glucansucrase dextransucrase recombinant expression protein design dextran |
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