Catalytic properties of Escherichia coli polyphosphate kinase: an enzyme for ATP regeneration.
Catalytic properties of Escherichia coli polyphosphate kinase (EC 2.7.4.1), a promising enzyme for use in ATP regeneration (Hoffman, et al., 1988, Biotechnol. Appl. Biochem. 10, 107-117), are reported here. E. coli polyphosphate kinase (PPK) is broadly active in the pH range 5.5 to 8.5, having an optimal Vmax at pH 7.2. The Km values for the substrates, ADP and polyphosphate (Pn), change little in the same pH range. The optimal concentration range for the Mg2+ activator is 1-20 mM, with an activity maximum at 10 mM Mg2+. In addition to Mg2+, Mn2+ and Co2+ can serve as activators of E. coli PPK, whereas Zn2+ and Cu2+ are highly inhibitory. E. coli PPK is most active with Pn substrates of chain length greater than 132 phosphoryl units. The enzyme activity decreases with decreasing Pn chain length and approaches zero (less than 1%) at a chain length less than or equal to 5. Equilibrium yields of ATP of greater than 85% are readily attained at substrate concentrations below 1 mM. An operational equilibrium constant for the PPK reaction, defined as [ATP]/[ADP][Pn], was determined to be 7.5 (+/- 3.4) x 10(5) M-1. The data presented here serve as a base of information from which assessments of the suitability of E. coli PPK for specific ATP regeneration applications can be made.
Haeusler
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PA
Department of Chemistry, Lebanon Valley College, Annville, Pennsylvania 17003.
Dieter
L
L
Rittle
K J
KJ
Shepler
L S
LS
Paszkowski
A L
AL
Moe
O A
OA
D000255:Adenosine Triphosphate / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid; D004789:Enzyme Activation; D004926:Escherichia coli / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D010770:Phosphotransferases / Q000378:metabolism*; D017856:Phosphotransferases (Phosphate Group Acceptor); D011122:Polyphosphates / Q000737:chemistry / Q000378:metabolism
D011122:Polyphosphates; D008345:Manganese; D000255:Adenosine Triphosphate; D010770:Phosphotransferases; D017856:Phosphotransferases (Phosphate Group Acceptor); C022424:polyphosphate kinase; D008274:Magnesium; D002118:Calcium