title: Highly efficient production of rare sugars D-psicose and L-tagatose by two engineered D-tagatose epimerases.
abstract: Rare sugars are monosaccharides that do not occur in nature in large amounts. However, many of them demonstrate high potential as low-calorie sweetener, chiral building blocks or active pharmaceutical ingredients. Their production by enzymatic means from broadly abundant epimers is an attractive alternative to synthesis by traditional organic chemical means, but often suffers from low space-time yields and high enzyme costs due to rapid enzyme degradation. Here we describe the detailed characterization of two variants of d-tagatose epimerase under operational conditions that were engineered for high stability and high catalytic activity towards the epimerization of d-fructose to d-psicose and l-sorbose to l-tagatose, respectively. A variant optimized for the production of d-psicose showed a very high total turnover number (TTN) of up to 10(8) catalytic events over a catalyst's lifetime, determined under operational conditions at high temperatures in an enzyme-membrane reactor (EMR). Maximum space-time yields as high as 10.6 kg L(-1) d(-1) were obtained with a small laboratory-scale EMR, indicating excellent performance. A variant optimized for the production of l-tagatose performed less stable in the same setting, but still showed a very good TTN of 5.8 × 10(5) and space-time yields of up to 478 g L(-1) d(-1) . Together, these results confirm that large-scale enzymatic access to rare sugars is feasible.
authors.affiliation: Bioprocess Lab, Department of Biosystems Science and Engineering D-BSSE, ETH Zurich Mattenstrasse 26, Basel, 4058, Switzerland.
authors.forename: Andreas
authors.initials: A
authors.lastname: Bosshart
authors.affiliation: Bioprocess Lab, Department of Biosystems Science and Engineering D-BSSE, ETH Zurich Mattenstrasse 26, Basel, 4058, Switzerland.
authors.forename: Nina
authors.initials: N
authors.lastname: Wagner
authors.affiliation: Bioprocess Lab, Department of Biosystems Science and Engineering D-BSSE, ETH Zurich Mattenstrasse 26, Basel, 4058, Switzerland.
authors.forename: Lei
authors.initials: L
authors.lastname: Lei
authors.affiliation: Bioprocess Lab, Department of Biosystems Science and Engineering D-BSSE, ETH Zurich Mattenstrasse 26, Basel, 4058, Switzerland.
authors.forename: Sven
authors.initials: S
authors.lastname: Panke
authors.affiliation: Bioprocess Lab, Department of Biosystems Science and Engineering D-BSSE, ETH Zurich Mattenstrasse 26, Basel, 4058, Switzerland. matthias.bechtold@bsse.ethz.ch.
authors.forename: Matthias
authors.initials: M
authors.lastname: Bechtold
mesh_terms: D019020:Directed Molecular Evolution; D005632:Fructose / Q000378:metabolism*; D006601:Hexoses / Q000378:metabolism*; D015202:Protein Engineering; D019998:Racemases and Epimerases / Q000235:genetics* / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism
chemical_list: D006601:Hexoses; D011994:Recombinant Proteins; C003243:psicose; D005632:Fructose; D019998:Racemases and Epimerases; C030192:tagatose
keywords: Directed evolution; d-tagatose epimerase; enzyme engineering; enzyme membrane reactor; improving catalytic activity; rare monosaccharides