Extracts of Acetobacter xylinum catalyze the phosphorylation of
glycerol and
dihydroxyacetone (DHA) by
adenosine 5'-triphosphate (
ATP) to form, respectively, L-
alpha-glycerophosphate and DHA
phosphate. The ability to promote phosphorylation of
glycerol and DHA was higher in
glycerol-grown cells than in
glucose- or
succinate-grown cells. The activity of
glycerol kinase in extracts is compatible with the overall rate of
glycerol oxidation in vivo. The
glycerol-DHA
kinase has been purified 210-fold from extracts, and its molecular weight was determined to be 50,000 by gel filtration. The
glycerol kinase to DHA
kinase activity ratio remained essentially constant at 1.6 at all stages of purification. The optimal pH for both reactions was 8.4 to 9.2. Reaction rates with the purified
enzyme were hyperbolic functions of
glycerol, DHA, and
ATP. The Km for
glycerol is 0.5 mM and that for DHA is 5 mM; both are independent of the
ATP concentration. The Km for
ATP in both
kinase reactions is 0.5 mM and is independent of
glycerol and DHA concentrations.
Glycerol and DHA are competitive substrates with Ki values equal to their respective Km values as substrates. D-
Glyceraldehyde and l-
Glyceraldehyde were not phosphorylated and did not inhibit the
enzyme. Among the
nucleotide triphosphates tested, only
ATP was active as the phosphoryl group donor.
Fructose diphosphate (
FDP) inhibited both
kinase activities competitively with respect to
ATP (Ki= 0.02 mM) and noncompetitively with respect to
glycerol and DHA.
Adenosine 5'-diphosphate (
ADP) and
adenosine 5'-monophosphate (
AMP) inhibited both enzymic activities competitively with respect to
ATP (Ki (
ADP) = 0.4 mM; Ki (
AMP) =0.25 mM). A. xylinum cells with a high
FDP content did not grow
on glycerol. Depletion of cellular
FDP by
starvation enabled rapid growth
on glycerol. It is concluded that a single
enzyme from A. xylinum is responsible for the phosphorylation of both
glycerol and DHA. This as well as the sensitivity of the
enzyme to inhibition by
FDP and
AMP suggest that it has a regulatory role in
glycerol metabolism.