Is Ca2+ antagonists binding protein from cytosolic fraction the precursor of alpha 1-subunit of Ca2+ channel?
The binding of Ca2+ antagonists to soluble proteins obtained by ammonium sulphate precipitation from cytosol fraction of rabbit skeletal muscles was studied. The KD values for 3H D-888 and 3H PN 200-110 binding to soluble proteins were 21.3 +/- 3.1 nmol.l-1 and 28.8 +/- 8.9 nmol.l-1 respectively. Photoaffinity labelling of the soluble proteins with the arylazide 1,4-dihydropyridine probe 3H azidopine resulted in labelling of the 85-95 K protein band as determined by SDS polyacrylamide gel electrophoresis. Partial purification of prelabelled soluble sample by gel filtration on Sephadex G-150 gave a more precise molecular weight of 90 +/- 2.5K. Polyclonal antibodies prepared against Ca2+ channel complex from rabbit muscle T-tubules inhibited the 3H PN 200-110 binding. Our results suggest that the soluble protein with Mr = 90K +/- 2.5K may be a precursor of the large subunit of the membrane bound L-type Ca2+ channel in rabbit skeletal muscle.
Krizanová
O
O
Centre of Physiological Sciences, Slovak Academy of Sciences, Bratislava.
Hudecová
S
S
Orlický
J
J
Zachar
J
J
D000345:Affinity Labels; D000818:Animals; D000906:Antibodies; D001386:Azides / Q000378:metabolism; D001667:Binding, Competitive; D002121:Calcium Channel Blockers / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism*; D003593:Cytoplasm / Q000032:analysis; D004095:Dihydropyridines / Q000378:metabolism; D066298:In Vitro Techniques; D017275:Isradipine; D009132:Muscles / Q000032:analysis; D010069:Oxadiazoles / Q000378:metabolism; D011498:Protein Precursors / Q000302:isolation & purification*; D011817:Rabbits; D011869:Radioligand Assay; D011978:Receptors, Nicotinic / Q000302:isolation & purification*; D014466:Ultraviolet Rays; D014700:Verapamil / Q000031:analogs & derivatives / Q000378:metabolism
D000345:Affinity Labels; D000906:Antibodies; D001386:Azides; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D004095:Dihydropyridines; D010069:Oxadiazoles; D011498:Protein Precursors; D011978:Receptors, Nicotinic; C041130:azidopine; C043184:4-desmethoxyverapamil; D014700:Verapamil; D017275:Isradipine