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Improved activity and thermostability of Bacillus pumilus lipase by directed evolution

Akbulut, N., Ozturk, M. T., Pijning, T., Ozturk, S. I. & Gumusel, F., 10-Mar-2013, In : Journal of Biotechnology. 164, 1, p. 123-129 7 p.

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  • Improved activity and thermostability of Bacillus pumilus lipase by directed evolution

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DOI

  • Nagihan Akbulut
  • Merve Tuzlakoglu Ozturk
  • Tjaard Pijning
  • Saliha Issever Ozturk
  • Fusun Gumusel

To improve enzymatic activity of Bacillus pumilus lipases, DNA shuffling was applied to two lipase genes from local B. pumilus isolates. Using a high-throughput activity assay, the mutant with highest activity was selected. This chimeric mutant (L3-3), carrying two crossover positions and three point mutations, has a specific activity 6.4 and 8.2 times higher than the two parent enzymes. The mutant also is more tolerant to various detergents and organic solvents, and has a 9 times longer half-life at 50 degrees C. Homology modeling of mutant L3-3, based on the highly homologous B. subtilis lipase A, shows that the increased thermostability is likely due to structural rigidification and reduced surface hydrophobicity. Increased specific activity may result from the location of mutations close to the active site. Together, our results show that it is possible to evolve, by DNA shuffling, B. pumilus lipase variants with improved applicability as biocatalysts, even if the two parent enzymes are highly similar. (C) 2013 Elsevier B.V. All rights reserved.

Original languageEnglish
Pages (from-to)123-129
Number of pages7
JournalJournal of Biotechnology
Volume164
Issue number1
Publication statusPublished - 10-Mar-2013

    Keywords

  • Bacillus pumilus lipase, Biocatalyst, Directed evolution, DNA shuffling, Thermostability, 3D homology model, SUBTILIS LIPASE, MICROBIAL LIPASES, RECOMBINATION, ENANTIOSELECTIVITY, FRAGMENTATION, BIOTECHNOLOGY, PURIFICATION, BIOCATALYSTS, SELECTION, FEATURES

ID: 5801857