Staff Publications

Staff Publications

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    'Staff publications' is the digital repository of Wageningen University & Research

    'Staff publications' contains references to publications authored by Wageningen University staff from 1976 onward.

    Publications authored by the staff of the Research Institutes are available from 1995 onwards.

    Full text documents are added when available. The database is updated daily and currently holds about 240,000 items, of which 72,000 in open access.

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Record number 367217
Title Laboratory evolution of Pyrococcus furiosus alcohol dehydrogenase to improve the production of (2S,5S)-hexanediol at moderate temperatures
Author(s) Machielsen, M.P.; Leferink, N.G.H.; Hendriks, A.; Brouns, S.J.J.; Hennemann, H.; Daussmann, T.; Oost, J. van der
Source Extremophiles 12 (2008)4. - ISSN 1431-0651 - p. 587 - 594.
DOI https://doi.org/10.1007/s00792-008-0164-8
Department(s) Microbiological Laboratory
Biochemistry
VLAG
Publication type Refereed Article in a scientific journal
Publication year 2008
Keyword(s) directed evolution - escherichia-coli - protein - (2r,5r)-hexanediol - biocatalysis - catalysis - nadp(+) - complex - design - acid
Abstract There is considerable interest in the use of enantioselective alcohol dehydrogenases for the production of enantio- and diastereomerically pure diols, which are important building blocks for pharmaceuticals, agrochemicals and fine chemicals. Due to the need for a stable alcohol dehydrogenase with activity at low-temperature process conditions (30°C) for the production of (2S,5S)-hexanediol, we have improved an alcohol dehydrogenase from the hyperthermophilic archaeon Pyrococcus furiosus (AdhA). A stable S-selective alcohol dehydrogenase with increased activity at 30°C on the substrate 2,5-hexanedione was generated by laboratory evolution on the thermostable alcohol dehydrogenase AdhA. One round of error-prone PCR and screening of ~1,500 mutants was performed. The maximum specific activity of the best performing mutant with 2,5-hexanedione at 30°C was tenfold higher compared to the activity of the wild-type enzyme. A 3D-model of AdhA revealed that this mutant has one mutation in the well-conserved NADP(H)-binding site (R11L), and a second mutation (A180V) near the catalytic and highly conserved threonine at position 183.
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