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Engineering a Thermostable Transketolase for Unnatural Conversion of (2 S )‐Hydroxyaldehydes
Author(s) -
Abdoul Zabar Juliane,
Lorillière Marion,
Yi Dong,
Saravanan Thangavelu,
Devamani Titu,
Nauton Lionel,
Charmantray Franck,
Hélaine Virgil,
Fessner WolfDieter,
Hecquet Laurence
Publication year - 2015
Publication title -
advanced synthesis and catalysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.541
H-Index - 155
eISSN - 1615-4169
pISSN - 1615-4150
DOI - 10.1002/adsc.201500207
Subject(s) - transketolase , chemistry , saturated mutagenesis , mutant , mutagenesis , glyceraldehyde , directed evolution , protein engineering , escherichia coli , stereochemistry , yeast , biochemistry , site directed mutagenesis , enzyme , dehydrogenase , gene
Transketolase (TK) from various origins (including Escherichia coli and yeast) has been described to be fully enantiomer specific for (2 R )‐hydroxyaldehyde substrates. A thermostable TK from G eobacillus stearothermophilus (TK gst ) was found to display a minor reactivity for (2 S )‐hydroxylated aldehydes. To improve this activity by directed protein evolution, we have built a library of TK gst variants by site saturation mutagenesis on two key positions L382 and D470. The best TK gst double mutant L382D/D470S shows up to 4‐ and 5‐fold higher activities towards L ‐lactaldehyde and L ‐glyceraldehyde as acceptor substrates, respectively. Preparative utility of this mutant was demonstrated by the one‐step synthesis of valuable L ‐ribulose and its 5‐deoxy analogue with the L ‐ erythro (3 S ,4 S ) configuration, which were previously inaccessible by using common TK sources.
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