Calcium-dependent regulation of caldesmon by an 11-kDa smooth muscle calcium-binding protein, caltropin.
Caldesmon from chicken gizzard muscle has been examined for its ability to interact with caltropin using affinity chromatography and the fluorescent probe acrylodan. The action of caltropin on the inhibitory effect of caldesmon on actomyosin ATPase was also studied. Like calmodulin, caltropin could release the inhibitory effect of caldesmon in the presence of Ca2+. Complete reversal was obtained when 1 mol of caltropin was added per mol of caldesmon. When caldesmon was applied to caltropin-Sepharose in the presence of Ca2+, most of the caldesmon was bound to the column and could be eluted with EGTA, indicating that there is a direct interaction between caldesmon and caltropin. Acrylodan-labeled caldesmon, when excited at 375 nm, had an emission maximum at 504 nm. Addition of caltropin in the presence of Ca2+ resulted in a nearly 50% increase in fluorescence intensity, and this was accompanied by a blue shift in the emission maximum (i.e., lambda em,max 492 nm), suggesting that the probe now occupies a more nonpolar environment. Titration of caltropin with labeled caldesmon indicated a strong affinity for this protein (Kd was in the order of 8 x 10(-8)-2 x 10(-7) M). However, when caltropin was added to labeled caldesmon in the presence of EGTA, there was no indication of any interaction. Caltropin was at least as potent as calmodulin, if not better, in reversing the inhibitory effect of caldesmon in the presence of calcium, making it a potential Ca2+ factor in regulating caldesmon in smooth muscle.
Mani
R S
RS
Department of Biochemistry, University of Alberta, Edmonton, Canada.
McCubbin
W D
WD
Kay
C M
CM
D015081:2-Naphthylamine / Q000031:analogs & derivatives; D000199:Actins / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000494:pharmacology*; D002135:Calcium-Binding Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D002148:Calmodulin-Binding Proteins / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology; D002645:Chickens; D002846:Chromatography, Affinity; D002942:Circular Dichroism; D004533:Egtazic Acid / Q000494:pharmacology; D005456:Fluorescent Dyes; D005895:Gizzard, Avian / Q000737:chemistry; D009130:Muscle, Smooth / Q000737:chemistry*; D009218:Myosins / Q000037:antagonists & inhibitors; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D014335:Tropomyosin / Q000494:pharmacology
D000199:Actins; D002135:Calcium-Binding Proteins; D002148:Calmodulin-Binding Proteins; D005456:Fluorescent Dyes; D014335:Tropomyosin; D004533:Egtazic Acid; C038518:acrylodan; D015081:2-Naphthylamine; D009218:Myosins; D002118:Calcium