STABILIZATION OF PHOSPHO-INTERMEDIATES OF RAT-LIVER PLASMA-MEMBRANE ALKALINE-PHOSPHATASE BY UNCOMPETITIVE INHIBITION - RELATION WITH PHOSPHATE-UPTAKE INTO HEPATOCYTES

Rat liver plasma membrane alkaline phosphatase (ALP) phospho-intermedi ates, which have molecular masses of 151 and 135 kDa bands, were label led at physiological pH with either (gamma-P-32) ATP or 32Pi. This lab eling was stabilized by a potent enzyme inhibitor, bromolevamisole (BL ), and not by bromodexamisole (BD). BL augmented the rate and extent o f autophosphorylation and slowed down the rate of autodephosphorylatio n of ALP. The phospho-intermediates labeling presented nearly the same kinetic behaviour with either (gamma-P-32) ATP or 32Pi. In the presen ce of BL a marked decrease of the phosphorylation state of many protei ns was observed in hepatocytes. BL also produced a decrease of the 32P i uptake into hepatocytes and a decrease of the specific radioactivity of cellular ATP. BD had nearly the same effect as BL on protein phosp horylation and 32Pi uptake. These results argued against a direct invo lvement of ALP in Pi transport across hepatocyte plasma membrane.