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Semi-Rational Design of Geobacillus stearothermophilus L-Lactate Dehydrogenase to Access Various Chiral α-Hydroxy Acids

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Abstract

Chiral α-hydroxy acids (AHAs) are rapidly becoming important synthetic building blocks, in particular for the production of pharmaceuticals and other fine chemicals. Chiral compounds of a variety of functionalities are now often derived using enzymes, and L-lactate dehydrogenase from the thermophilic organism Geobacillus stearothermophilus (bsLDH) has the potential to be employed for the industrial synthesis of chiral α-hydroxy acids. Despite the thorough characterization of this enzyme, generation of variants with high activity on non-natural substrates has remained difficult and therefore limits the use of bsLDH in industry. Here, we present the engineering of bsLDH using semi-rational design as a method of focusing screening in a small and smart library for novel biocatalysts. In this study, six mutant libraries were designed in an effort to expand the substrate range of bsLDH. The eight variants identified as having enhanced activity toward the selected α-keto acids belonged to the same library, which targeted two positions simultaneously. These new variants now may be useful biocatalysts for chiral synthesis of α-hydroxy acids.

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Acknowledgments

This work was supported by a grant from the TUBİTAK (grant number 213S099) and special thanks to COST Action (grant number CM1303).

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Authors and Affiliations

  1. Faculty of Science and Letters, Department of Molecular Biology and Genetics, Istanbul Technical University, 34469, Istanbul, Turkey

    Aşkın Sevinç Aslan & Nevin Gül Karagüler

  2. Molecular Biology-Biotechnology and Genetics Research Center (MOBGAM), Istanbul Technical University, 34469, Istanbul, Turkey

    Aşkın Sevinç Aslan & Nevin Gül Karagüler

  3. School of Chemistry, Manchester Institute of Biotechnology, University of Manchester, 131 Princess Street, Manchester, M1 7DN, UK

    William R. Birmingham & Nicholas J. Turner

  4. Molecular Biology and Genetics, Istanbul AREL University, Tepekent, Büyükçekmece, Istanbul, 34537, Turkey

    Barış Binay

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  1. Aşkın Sevinç Aslan
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  2. William R. Birmingham
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  3. Nevin Gül Karagüler
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  4. Nicholas J. Turner
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  5. Barış Binay
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Correspondence to Barış Binay.

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Aslan, A.S., Birmingham, W.R., Karagüler, N.G. et al. Semi-Rational Design of Geobacillus stearothermophilus L-Lactate Dehydrogenase to Access Various Chiral α-Hydroxy Acids. Appl Biochem Biotechnol 179, 474–484 (2016). https://doi.org/10.1007/s12010-016-2007-x

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  • DOI: https://doi.org/10.1007/s12010-016-2007-x

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