Specific inactivation of the phosphohydrolase component of the hepatic microsomal glucose-6-phosphatase system by diethyl pyrocarbonate.
We have examined the interactions of the histidine-specific reagent diethyl pyrocarbonate (DEPC) with the components of the rat hepatic glucose-6-phosphatase system (EC 3.1.3.9). DEPC is the first known reagent that satisfies the criteria of an active-site-specific label for the phosphohydrolase component. (a) It inactivates through formation of a stable covalent bond. (b) It is effective at reasonably low concentrations (2-4 mM) under relatively mild conditions (e.g. 30 degrees C at neutral pH). (c) Inactivation is substantially blocked by glucose 6-phosphate, Pi and NaF, compounds which are known to interact quite specifically with the phosphohydrolase. (d) Under conditions where glucose 6-phosphate and NaF protect the enzyme, no protection is provided against DEPC-mediated inactivation of two other functional components of the membrane, the glucose 6-phosphate translocase and UDP-glucuronyltransferase. DEPC also shows potential for use at 0 degree C as a label for UDP-glucuronyltransferase.
D000818:Animals; D001665:Binding Sites; D004047:Diethyl Pyrocarbonate / Q000494:pharmacology*; D005561:Formates / Q000494:pharmacology*; D005952:Glucose-6-Phosphatase / Q000037:antagonists & inhibitors*; D019298:Glucose-6-Phosphate; D005958:Glucosephosphates / Q000494:pharmacology; D014453:Glucuronosyltransferase / Q000037:antagonists & inhibitors; D066298:In Vitro Techniques; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000187:drug effects / Q000201:enzymology*; D010710:Phosphates / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D012969:Sodium Fluoride / Q000494:pharmacology
D005561:Formates; D005958:Glucosephosphates; D010710:Phosphates; D019298:Glucose-6-Phosphate; D012969:Sodium Fluoride; D014453:Glucuronosyltransferase; D005952:Glucose-6-Phosphatase; D004047:Diethyl Pyrocarbonate