Abstract
The gene epsN of Bacillus subtilis 168 was cloned and overexpressed in Escherichia coli. Purified recombinant EpsN is shown to be a pyridoxal 5'-phosphate (PLP)-dependent aminotransferase by absorption spectroscopy, l-cycloserine inhibition and reverse phase HPLC studies. EpsN catalyzes the conversion of UDP-2,6-dideoxy 2-acetamido 4-keto glucose to UDP-2,6-dideoxy 2-acetamido 4-amino glucose. Lys190 was found by sequence comparison and site-directed mutagenesis to form Schiff base with PLP. Mutagenesis studies showed that, in addition to Lys190, Ser185, Glu164, Gly58 and Thr59 are essential for aminotransferase activity.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Bacillus subtilis / enzymology*
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Bacillus subtilis / metabolism
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Bacterial Proteins / metabolism*
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Biocatalysis
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Glucose / analogs & derivatives*
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Glucose / chemistry
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Glucose / metabolism
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Molecular Structure
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Mutagenesis, Site-Directed
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Polysaccharides, Bacterial / genetics
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Polysaccharides, Bacterial / isolation & purification
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Polysaccharides, Bacterial / metabolism*
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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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Transaminases / metabolism*
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Uridine Diphosphate / chemistry
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Uridine Diphosphate / metabolism*
Substances
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Bacterial Proteins
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Polysaccharides, Bacterial
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Recombinant Proteins
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UDP-2,6-dideoxy 2-acetamido 4-keto glucose
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Uridine Diphosphate
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Transaminases
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Glucose