Abstract
The gene coding for D-psicose 3-epimerase (DPEase) from Clostridium sp. BNL1100 was cloned and expressed in Escherichia coli. The recombinant enzyme was purified by Ni-affinity chromatography. It was a metal-dependent enzyme and required Co(2+) as optimum cofactor. It displayed catalytic activity maximally at pH 8.0 and 65 °C (as measured over 5 min). The optimum substrate was D-psicose, and the K m, turnover number (k cat), and catalytic efficiency (k cat/K m) for D-psicose were 227 mM, 32,185 min(-1), and 141 min(-1 )mM(-1), respectively. At pH 8.0 and 55 °C, 120 g D-psicose l(-1) was produced from 500 g D-fructose l(-1) after 5 h.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Cations, Divalent / metabolism
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Chromatography, Affinity
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Cloning, Molecular
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Clostridium / enzymology*
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Clostridium / genetics
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Cobalt / metabolism
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Coenzymes / metabolism
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Enzyme Stability
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Escherichia coli / genetics
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Fructose / metabolism*
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Gene Expression
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Hydrogen-Ion Concentration
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Kinetics
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Racemases and Epimerases / chemistry
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Racemases and Epimerases / genetics
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Racemases and Epimerases / isolation & purification
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Racemases and Epimerases / metabolism*
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Recombinant Proteins / chemistry
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Recombinant Proteins / genetics
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Recombinant Proteins / isolation & purification
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Recombinant Proteins / metabolism
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Temperature
Substances
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Cations, Divalent
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Coenzymes
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Recombinant Proteins
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psicose
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Fructose
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Cobalt
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Racemases and Epimerases