LOW-K-M MANNOSE-6-PHOSPHATASE AS A CRITERION FOR MICROSOMAL INTEGRITY

The low-K-m activity of mannose-6-phosphatase (Man-6-Pase) has been us ed for many years to measure the structural integrity of microsomes. R ecently histone II-A has been shown to activate glucose-6-phosphatase (Glc-6-Pase) and Man-6-Pase activities. However, in contrast to deterg ents, this compound appears to activate without disrupting microsomal vesicles (J.-F. St-Denis, B. Annabi, H. Khoury, and G. van de Werve. 1 995. Biochem. J. 310: 221-224). This suggests that Man-6-Pase latency can be abolished without disrupting microsomal integrity and that even normally microsomes may manifest some low-K-n Man-6-Pase activity wit hout being ''leaky.'' We have studied the relationship of Man-6-Pase w ith microsomal integrity further by measuring the;latency of several e nzymes reported to reside within the lumen of endoplasmic reticulum. W e have also correlated this latency with the microsomal permeability o f substrates for these enzymes. We found that (i) lumenal enzymes have different degrees of latency when compared with each other, (ii) perm eability, as determined via osmotically induced changes in light scatt ering, is not always consistent with enzymatic latency, (iii) increase s in the hydrolysis of Glc-6-P and Man-B-P were not parallel when micr osomes were treated with low but increasing concentrations of detergen t, and (iv) kinetic studies suggest that mannose-6-phosphate is hydrol yzed by untreated microsomes by more than a single mechanism. We propo se that Man-6-Pase is not a reliable index of the integrity of microso mes.