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Target molecular weight of the gastric (H+ + K+)-ATPase functional and structural molecular size
Abstract:
The state of assembly of the (H+ + K+)-ATPase in purified hog gastric mucosa membranes was studied by target size analysis applied to radiation-induced enzyme inactivation and polypeptide degradation data. Radiation inactivated the Mg2+-ATPase, K+-stimulated ATPase, and p-nitrophenyl phosphatase activities of the membrane preparation with a dose dependence characteristic of a target size of 270,000-daltons. Radiation also bleached the major 100,000-dalton sodium dodecyl sulfate-gel electrophoresis band of this preparation, indicating a radiation-induced degradation. This apparent polypeptide degradation exhibited a dose dependency corresponding to a target size of 250,000 daltons in situ. It is suggested that the gastric ATPase is a trimeric assembly of the 100,000-dalton polypeptides.
Insights
Researchers studied the assembly of gastric proton pump (H+ + K+)-ATPase using radiation inactivation. Findings suggest the enzyme is a trimeric structure composed of 100,000-dalton polypeptide units.
Area of Science:
- Biochemistry
- Membrane protein assembly
- Enzyme kinetics
Background:
- The gastric proton pump, (H+ + K+)-ATPase, is crucial for acid secretion.
- Understanding its assembly is key to comprehending its function and regulation.
- Previous studies on enzyme structure often relied on biochemical methods.
Purpose of the Study:
- To determine the assembly state and target size of the (H+ + K+)-ATPase in purified hog gastric mucosa membranes.
- To investigate the relationship between enzyme activity, polypeptide structure, and radiation inactivation.
- To elucidate the oligomeric nature of the gastric ATPase.
Main Methods:
- Target size analysis using radiation-induced enzyme inactivation.
- Analysis of radiation-induced polypeptide degradation via SDS-gel electrophoresis.
- Measurement of Mg2+-ATPase, K+-stimulated ATPase, and p-nitrophenyl phosphatase activities.
Main Results:
- Enzyme inactivation (Mg2+-ATPase, K+-stimulated ATPase, p-nitrophenyl phosphatase) indicated a target size of 270,000 daltons.
- Radiation-induced degradation of the major 100,000-dalton polypeptide band corresponded to an in situ target size of 250,000 daltons.
- The data suggest a functional enzyme unit size close to the size of the major polypeptide.
Conclusions:
- The (H+ + K+)-ATPase likely exists as a trimeric assembly of 100,000-dalton polypeptides.
- Radiation inactivation is a valuable tool for probing the in situ size and assembly of membrane-bound enzymes.
- This study provides insights into the quaternary structure of the gastric proton pump.