J. Biol. Chem., Vol. 258, Issue 6, 3668-3672, Mar, 1983
Platelet AMP deaminase. Regulation by Mg-ATP2- and inorganic phosphate and inhibition by the transition state analog coformycin
B Ashby and H Holmsen
Kinetic studies with platelet AMP deaminase, at pH 7.0 and 100 mM NaC1,
gave cooperative initial velocity curves with AMP as substrate, with Mg-
ATP2- as an activator, and with Pi as an inhibitor. In the absence of
Mg-ATP2-, the s0.5 for AMP was 4.5 mM with a Hill coefficient approaching
2.0. In the presence of saturating Mg-ATP2-, the s0.5 for AMP was reduced
to 0.18 mM, the maximum velocity was increased by about 35%, and the Hill
coefficient was 1.0. The half-activation constant for Mg-ATP2- varied from
0.7 to 0.07 mM as the concentration of AMP was varied from 0.1 to 5.0 mM
and the Hill coefficient for Mg-ATP activation changed from 2.0 to 1.0 over
the same range. Phosphate inhibition was competitive with AMP and with
Mg-ATP2- (Ki = 2.0 mM) and reversed the activation by Mg-ATP2-. Coformycin
inhibited the Mg-ATP- activated enzyme with a Ki less than 0.25 microM.
Coformycin inhibition was slow, with a second order rate constant of 6.0 X
10(4) M-1 min-1, suggesting that the compound acts as a transition state
analog according to Frieden, C., Kurz, L. C., and Gilbert, H. R. (1980)
Biochemistry 19, 5303-5309. The kinetic properties of the enzyme indicate
that substantial regulation can occur through changes in AMP concentration
acting synergistically to enhance Mg-ATP2- binding and displace Pi from a
single type of regulatory site.
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