Regulation and inactivation of brain phosphocholine-phosphatase activity

Regulation of phosphocholine-hydrolyzing phosphatase (phosphocholine-phosphatase) activity, purified from bovine brain, was examined under physiological conditions. Various endogenous phosphomonoesters, which were utilized as substrate, inhibited the phosphocholine-phosphatase activity competitively (Ki, 5.5-82.0 microM); among phosphomonoesters tested, there was a similar order of capability between the binding affinity of substrate and the inhibitory potency. In addition, phosphate ions also inhibited the phosphatase activity competitively with a Ki value of approximately 167 microM. Although leucine or theophylline inhibited the phosphatase activity at pH 9.0, their inhibitory action decreased greatly at pH 7.4. The pH-Km and pH-Vm profiles indicate that ionizable amino acids are involved in substrate binding as well as catalysis, alluding that the phosphatase activity may be highly dependent on the intracellular pH. Amino acid modification study supports the existence of tyrosine, arginine or lysine residue in the active site, and the participation of tyrosine residue in the catalytic action may be suggested positively from the susceptibiliy to the action of tetranitromethane or HOI-generator. Separately, the oxidative inactivation of phosphocholine-phosphatase activity was investigated. Of oxidants tested, HOONO, HOCl, HOI and ascorbate/Cu2+ system were effective to inactivate the phosphatase activity. Noteworthy, a remarkable inactivation was accomplished by 30 microM HOCl in combination with 1 mM KI. In addition, Cu2+ (3 microM) in combination with ascorbate at concentrations as low as 0.1-0.3 mM reduced the phosphatase activity to a great extent. From these results, it is proposed that the phosphocholine-phosphatase activity may be regulated endogenously and susceptible to the various oxidant systems in vivo.