Latency studies on rat liver microsomal glucose-6-phosphatase. Correlation of membrane modification and solubilization by Triton X-114 with the enzymatic activity.
Interrelationships between the catalytic properties of glucose-6-phosphatase and the membrane structure of rat liver microsomes were investigated. Membrane modification and solubilization employing the nonionic surfactant Triton X-114 were standardized and analysed by ultracentrifugation, surface tension- and turbidity measurements. The effect of Triton X-114 on the glucose-6-phosphatase activity was studied systematically and the whole magnitude of time- and temperature-dependent inactivation of this enzyme has been demonstrated. The results show that the activity measured is always a resultant of two processes, the beginning of inactivation and the release of latency. Maximal activation of about 600% (83% of apparent latency) was obtained at 0 degree C. A correlation between membrane modification and solubilization and the conditions under preincubation and test incubation reveals that studies on detergent-disrupted microsomes are performed on structures reassembled from solubilizates and this implies a modified microenvironment in the reconstitutes. Kinetic analyses suggest interrelationships between Triton X-114 and the permeability barrier of the glucose-6-phosphatase system. At 0 degree C 2-propanol and ethanol are more potent tools for membrane modification than Triton X-114 and release 88% and 86% latent activity corresponding to an activation of the glucose-6-phosphatase of about 850% and 700%, respectively. These observations suggest that detergent treatment of microsomes could not preserve the functional integrity of the glucose-6-phosphate phosphohydrolase, which is one dogma of the substrate-transport hypothesis developed by Arion and his co-workers (Arion, W.J., et al. (1975) Mol. Cell. Biochem. 6, 75-83).
Schulze
H U
HU
Kannler
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R
Junker
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B
D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003080:Cold Temperature; D005952:Glucose-6-Phosphatase / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D009391:Nephelometry and Turbidimetry; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains; D012995:Solubility; D014461:Ultracentrifugation
D011092:Polyethylene Glycols; D017830:Octoxynol; C010615:Nonidet P-40; D005952:Glucose-6-Phosphatase