Purification and properties of dihydroxyacetone kinase from Klebsiella pneumoniae

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Abstract
Dihydroxyacetone (DHA) kinase of K. pneumoniae, a gene product of the dha regulon responsible for fermentative dissimilation of glycerol and DHA, was purified 120-fold to a final specific activity of 10 .mu.mol .times. min-1 .times. mg protein-1 at 30.degree. C. The enzyme, a dimer of a 53,000 .+-. 5000-dalton polypeptide, is highly specific for DHA (Km, .apprx. 4 .mu.M). Glycerol is not a substrate at 1 mM and is not an inhibitor even at 100 mM. The enzyme is not inhibited by 5 mM fructose-1,6-diphosphate. Ca2+ gives a higher enzyme activity than Mg2+ as a cationic cofactor. Escherichia coli glycerol kinase acts on both glycerol and DHA and is allosterically inhibited by fructose-1,6-diphosphate. Antibodies raised against E. coli glycerol kinase cross-reacted with K. pneumoniae glycerol kinase but not with K. pneumoniae DHA kinase.