Properties of polyphosphate: AMP phosphotransferase of Acinetobacter strain 210A.
Polyphosphate:AMP phosphotransferase, an enzyme which catalyzes the phosphorylation of AMP to ADP at the expense of polyphosphate, was purified more than 1,500-fold from Acinetobacter strain 210A by streptomycin sulfate precipitation and by Mono-Q, Phenyl Superose, and Superose column chromatography. Streptomycin sulfate precipitation appeared to be an effective step in the purification procedure. During the following chromatographic steps, there was a 29-fold increase in specific activity but the yield was low (0.3%). Kinetic studies showed apparent Km values of 0.26 mM for AMP and 0.8 microM for polyphosphate with an average chain length of 35 phosphate groups. The highest activities were found with polyphosphate molecules of 18 to 44 phosphate residues. The polyphosphate chain was degraded completely to ADP. The mechanism of degradation is processive. No activity was obtained with ortho-, pyro-, tri-, and tetraphosphate. The enzyme was inhibited by pyro-, tri-, and tetraphosphate. The inhibition by tri- and tetraphosphate was mixed with polyphosphate as a substrate. The inhibition constants for the dissociation of the enzyme-inhibitor complex and for the enzyme-inhibitor-substrate complex were 0.9 and 6.5 mM, respectively, for triphosphate and 0.7 and 1.5 mM, respectively, for tetraphosphate.
Bonting
C F
CF
Department of Microbiology, Agricultural University Wageningen, The Netherlands.
Kortstee
G J
GJ
Zehnder
A J
AJ
D000150:Acinetobacter / Q000201:enzymology*; D000249:Adenosine Monophosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D007700:Kinetics; D010770:Phosphotransferases / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D017856:Phosphotransferases (Phosphate Group Acceptor); D011122:Polyphosphates / Q000378:metabolism
D001426:Bacterial Proteins; D011122:Polyphosphates; D000249:Adenosine Monophosphate; D010770:Phosphotransferases; D017856:Phosphotransferases (Phosphate Group Acceptor); C059169:polyphosphate AMP phosphotransferase