1990 Fiscal Year Final Research Report Summary
Protein-protein interaction of detergent solubilized Ca^<2+>-ATPase during ATP hydrolysis analyzed by low-angle laser light scattering photometry coupled with high-performance gel chromatography.
| Project/Area Number |
01870107
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Research Field |
医学一般
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| Research Institution | Osaka University School of Medicine |
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Principal Investigator |
TADA Michihiko Osaka University School of Medicine, Department of Pathophysiology, Professor, 医学部, 教授 (90093434)
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| Co-Investigator(Kenkyū-buntansha) |
INUI Makoto Osaka University School of Medicine, Department of Neurochemistry and Neuropharm, 医学部, 助手 (70223237)
HOSHIDA Shiro Osaka University Hospital, First Department of Medicine, Clinical Fellow, 医学部附属病院, 医員
KUZUYA Tsunehiko Osaka University School of medicine, First Department of Medicine, Assistant Pro, 医学部, 助手 (80150340)
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| Project Period (FY) |
1989 – 1990
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| Keywords | Ca^<2+> pump ATPase / Muscle contraction / Low angle laser light scattering / Protein-protein interaction / Active transport / Calcium ion / Solubilization / Membrane protein |
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| Research Abstract |
To elucidate the molecular interaction between Ca pump ATPases of sarcoplasmic reticulum (SR) during the enzyme reaction, we analyzed the solubilized ATPase with non-ionic detergent C_<12>E_8 using low-angle laser light scattering photometry coupled with high-performance gel chromatography. Detergent solubilized ATPase emerged as a single peak with an intermediate molecular weight between the monomer and dimer, suggesting a dissociation-association equilibrium of two components. In the presence of 50 ug/ml phosphatidylcholine and 0.3 mg/ml C_<12>E_8 at 0^ﾟC without ATP, the Ca pump ATP emerged as the two distinct components with molecular weights of 125,000 <plus-minus> 2,100 and 211,300 <plus-minus> 7,300, corresponding to the monomer and the dimer, indicating that there was no rapid interconversion between the two forms. Addition of ATP induced fusion of the two components. The apparent molecular weight of the fused peak shifted from the monomer to the dimer as the amount of the protein increased. Addition of ADP or AMPPCP, non-hydrolyzable adenine nucleotides, or the presence of 5 mM CaCl_2, the conditions in which the turn-over of the ATPase enzyme was extremely slow, did not induce the fusion of the peaks. Thus, the solubilized Ca pump ATPase underwent a rapid interconversion between the monomer and the dimer during the reaction cycle of ATP hydrolysis. These results indicate that the protein-protein interaction between ATPase polypeptides may play an important role in Ca translocation across SR membrane.
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