Low-Km mannose-6-phosphatase as a criterion for microsomal integrity.
The low-Km activity of mannose-6-phosphatase (Man-6-Pase) has been used for many years to measure the structural integrity of microsomes. Recently histone II-A has been shown to activate glucose-6-phosphatase (Glc-6-Pase) and Man-6-Pase activities. However, in contrast to detergents, this compound appears to activate without disrupting microsomal vesicles (J.-F. St-Denis, B. Annabi, H. Khoury, and G. van de Werve. 1995. 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-Km Man-6-Pase activity without being "leaky." We have studied the relationship of Man-6-Pase with microsomal integrity further by measuring the latency of several enzymes reported to reside within the lumen of endoplasmic reticulum. We have also correlated this latency with the microsomal permeability of substrates for these enzymes. We found that (i) lumenal enzymes have different degrees of latency when compared with each other, (ii) permeability, as determined via osmotically induced changes in light scattering, is not always consistent with enzymatic latency, (iii) increases in the hydrolysis of Glc-6-P and Man-6-P were not parallel when microsomes were treated with low but increasing concentrations of detergent, and (iv) kinetic studies suggest that mannose-6-phosphate is hydrolyzed by untreated microsomes by more than a single mechanism. We propose that Man-6-Pase is not a reliable index of the integrity of microsomes.
Pederson
B A
BA
Department of Biochemistry and Molecular Biology, University of North Dakota School of Medicine and Health Sciences, Grand Forks 58202, USA.
Foster
J D
JD
Nordlie
R C
RC
D017766:Acid Anhydride Hydrolases / Q000378:metabolism; D000217:Acyltransferases / Q000378:metabolism; D000818:Animals; D015415:Biomarkers; D004492:Edetic Acid / Q000493:pharmacokinetics; D042843:Glucose 1-Dehydrogenase; D005948:Glucose Dehydrogenases / Q000378:metabolism; D006868:Hydrolysis; D007425:Intracellular Membranes / Q000201:enzymology* / Q000648:ultrastructure; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology* / Q000648:ultrastructure; D010539:Permeability; D010744:Phosphoric Monoester Hydrolases / Q000378:metabolism*; D051381:Rats
D015415:Biomarkers; D004492:Edetic Acid; D005948:Glucose Dehydrogenases; D042843:Glucose 1-Dehydrogenase; D000217:Acyltransferases; C020124:ethanol acyltransferase; C023553:mannose-6-phosphatase; D010744:Phosphoric Monoester Hydrolases; D017766:Acid Anhydride Hydrolases; C023837:nucleoside-diphosphatase