MODULATION OF MAMMALIAN CARDIAC AMP DEAMINASE BY PROTEIN KINASE-C-MEDIATED PHOSPHORYLATION

Using AMP deaminase (AMP aminohydrolase; EC 3.5.4.6) purified from rab bit left-ventricular heart tissue, we report direct investigation of t he potential for cardiac AMP deaminase activity to be regulated by kin ase-mediated phosphorylation. Rabbit heart AMP deaminase served as a s ubstrate for Ca2+/phospholipid-dependent protein kinase (protein kinas e C; PKC) exclusively; no other mammalian protein kinase phosphorylate d the enzyme. PKC-dependent AMP deaminase phosphorylation was rapid, l inear with respect to time and the concentrations of PKC and AMP deami nase in the reaction, and inhibitable by staurosporine. Upon phosphory lation, the apparent K(m) of cardiac AMP deaminase decreased from 5.6 mM to 1.2 mM, without effect on the V(max). Whether phosphorylated or not, rabbit heart AMP deaminase was inhibited by 1.0 mM GTP, which dec reased the V(max) by approximately 50 % in each case. PKC-dependent ph osphorylation of cardiac AMP deaminase did not alter the enzyme's allo sterism toward millimolar ATP or ADP: both nucleotides at 1.0 mM conce ntration decreased the apparent K(m) to approximately 0.5 mM. Treatmen t of cardiac phospho-AMP deaminase with either the protein phosphatase calcineurin or alkaline phosphatase generated a dephosphorylated form which displayed molecular and kinetic properties identical with those of the originally isolated enzyme. These data raise the possibility t hat a phosphorylation-dephosphorylation mechanism may regulate flux th rough AMP deaminase in the heart under pathological conditions, such a s myocardial ischaemia, characterized by PKC activation and adenylate depletion.