US EPA

7449717 

Journal Article 

Protein phosphatase 2B dephosphorylates twitchin, initiating the catch state of invertebrate smooth muscle 

Yamada, A; Yoshio, M; Nakamura, A; Kohama, K; Oiwa, K 

2004 

Yes 

Journal of Biological Chemistry
ISSN: 0021-9258
EISSN: 1083-351X 

AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC 

BETHESDA 

279 

39 

40762-40768 

English 

15272026 

10.1074/jbc.M405191200 

WOS:000223916800069 

"Catch" is the state where some invertebrate muscles sustain high tension for long periods at low ATP hydrolysis rates. Physiological studies using muscle fibers have not yet fully provided the details of the initiation process of the catch state. The process was extensively studied by using an in vitro reconstitution assay with several phosphatase inhibitors. Actin filaments bound to thick filaments pretreated with the soluble protein fraction of muscle homogenate and Ca2+ (catch treatment) in the presence of MgATP at a low free Ca2+ concentration (the catch state). Catch treatment with > 50 microm okadaic acid, > 1 microm microcystin LR, 1 microm cyclosporin A, 1 microm FK506, or 0.2 mm calcineurin autoinhibitory peptide fragment produced almost no binding of the actin filaments, indicating protein phosphatase 2B (PP2B) was involved. Use of bovine calcineurin (PP2B) and its activator calmodulin instead of the soluble protein fraction initiated the catch state, indicating that only PP2B and calmodulin in the soluble protein fraction are essential for the initiation process. The initiation was reproduced with purified actin, myosin, twitchin, PP2B, and calmodulin. 32P autoradiography showed that only twitchin was dephosphorylated during the catch treatment with either the soluble protein fraction or bovine calcineurin and calmodulin. These results indicate that PP2B directly dephosphorylates twitchin and initiates the catch state and that no other component is required for the initiation process of the catch state. 

Activation analysis; Body fluids; Hydrolysis; Muscle; Invertebrate muscles; Muscle fibres; Protein phosphatase; Adenosinetriphosphate; actin; adenosine triphosphate magnesium; calcineurin; calcineurin inhibitor; calmodulin; cyanoginosin LR; cyclosporin A; muscle protein; okadaic acid; tacrolimus; twitchin; unclassified drug; actin filament; article; autoradiography; binding affinity; catch state; cattle; dephosphorylation; enzyme inhibition; in vitro study; invertebrate; muscle cell; muscle tone; nonhuman; priority journal; process monitoring; protein analysis; protein phosphorylation; protein purification; smooth muscle; Adenosine Triphosphate; Animals; Bivalvia; Caenorhabditis elegans Proteins; Calcineurin; Calcium; Calmodulin; Calmodulin-Binding Proteins; Cattle; Cyclic AMP; Cyclosporine; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Enzyme Inhibitors; Magnesium; Microcystins; Models, Biological; Muscle Proteins; Muscle, Smooth; Okadaic Acid; Peptides, Cyclic; Phosphoric Monoester Hydrolases; Phosphorylation; Tacrolimus; Bos taurus; Bovinae; Invertebrata