Increased permeability to Ca2+ in response to a depolarizing membrane potential occurs in a wide variety of excitable cell types. In some cells this permeability increase is either blocked or enhanced by dihydropyridines, leading to the hypothesis that the dihydropyridine binding site is part of or associated with the voltage-dependent calcium channel. We propose to purify the DHP binding protein from bovine ventricular tissue and to characterize its subunit composition. This will be accomplished using the binding radioactive DHPS to follow the purification. Purification procedures will utilize conventional column chromatography, HPLC and FPLC techniques, affinity chromatography, and ultracentrifugation. In a collaborative effort, we will use reconstitution techniques to establish whether the DHP binding protein is also the voltage- dependent calcium channel. Important to the purification and characterization of the cardiac DHP binding protein will be the production of antibody and toxin probes specific for this protein. One approach to the preparation of specific antibodies will be to use the subunits of the skeletal muscle DHP binding protein as antigens. We will also use the partially-purified cardiac preparation. Probes of the DHP binding protein will be used to purify it further, to determine the subunit composition, and to analyze the arrangement of the subunits in the membrane.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Modified Research Career Development Award (K04)
Project #
5K04HL002063-02
Application #
3074258
Study Section
Pharmacology A Study Section (PHRA)
Project Start
1988-03-01
Project End
1993-02-28
Budget Start
1989-03-01
Budget End
1990-02-28
Support Year
2
Fiscal Year
1989
Total Cost
Indirect Cost
Name
Baylor College of Medicine
Department
Type
Schools of Medicine
DUNS #
074615394
City
Houston
State
TX
Country
United States
Zip Code
77030