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Phosphorylase phosphatase from skeletal muscle membranes
E Villa-Moruzzi1, L M Heilmeyer
1Istituto di Patologia Generale, Università di Pisa, Italy.
European Journal of Biochemistry
|December 15, 1987
Summary
Skeletal muscle microsomes contain inhibited phosphorylase phosphatase. Dilution releases and activates the enzyme, which can rebind to membranes via a protein receptor, suggesting a novel regulatory mechanism for phosphatase activity in muscle.
Area of Science:
- Biochemistry
- Molecular Biology
- Skeletal Muscle Physiology
Background:
- Microsomes from skeletal muscle glycogen particles harbor phosphorylase phosphatase.
- This membrane-associated phosphatase is initially in an inhibited state.
Purpose of the Study:
- To investigate the regulation and characteristics of skeletal muscle phosphorylase phosphatase.
- To elucidate the binding mechanism and regulatory role of membrane-associated phosphatase.
Main Methods:
- Glycogen digestion and differential centrifugation to isolate microsomes.
- Enzyme purification using polylysine and FPLC chromatography.
- SDS-gel electrophoresis and activity assays to characterize the enzyme.
Main Results:
- Dilution of microsomes induces dissociation and activation of the phosphatase.
- A protein receptor on membranes mediates phosphatase binding and inhibition.
- Purified catalytic subunit (38 kDa) forms inactive complexes with inhibitor 2, which can be reactivated.
- The same catalytic subunit exists in different conformations in cytosol, glycogen particles, and microsomes.
Conclusions:
- Membrane association and conformational state are key regulators of skeletal muscle phosphorylase phosphatase.
- Enzyme displacement between cellular sites may represent a novel regulatory mechanism.