Comparison of calmodulin and troponin C with and without its amino-terminal helix (residues 1-11) in the activation of erythrocyte Ca(2+)-ATPase.
Troponin C can replace calmodulin in the activation of the Ca(2+)-ATPase of pig erythrocytes provided that the reaction medium contains relatively high free Ca2+ concentrations (> 0.5 microM). In the presence of 10 microM free Ca2+, the troponin C-activated ATPase reaches a maximal velocity of approximately 70% of that attained with calmodulin. The half-maximal concentration for troponin C activation is about 200 times greater than for calmodulin. Troponin C displaces the half-maximal concentration for activation by Ca2+ to pCa 5.46 and the cooperativity between the Ca2+ binding sites to nH 1.1, compared with pCa 6.14 and nH 1.72 when calmodulin is used. Both EF-hand proteins also elicit activation by ATP at a nucleotide regulatory site, as well as a Ca(2+)-dependent p-nitrophenyl phosphatase activity. Troponin I prevents activation of the enzyme by troponin C. A mutant of troponin C with the amino-terminal helix deleted (NHdel) activates the Ca(2+)-ATPase to the same extent and with the same Ca2+ dependence as wild-type troponin C (rTnC); the half-maximal concentration for activation by NHdel is 2.5 times smaller than that for rTnC. We conclude that the structural features that distinguish the two EF-hand proteins affect their binding to the target enzyme more than their ability to transform the enzyme's response to Ca2+ or ATP. The differences in the amino-terminal domains of troponin C and calmodulin cannot account for the differences in ability of these proteins to activate the target system used as a model.
da Silva
E F
EF
Departamento de Bioquímica Médica, Universidade Federal do Rio de Janeiro, Brazil.
Sorenson
M M
MM
Smillie
L B
LB
Barrabin
H
H
Scofano
H M
HM
D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism; D000252:Calcium-Transporting ATPases / Q000378:metabolism*; D002147:Calmodulin / Q000378:metabolism*; D002384:Catalysis; D002417:Cattle; D002645:Chickens; D004789:Enzyme Activation; D004912:Erythrocytes / Q000201:enzymology*; D010446:Peptide Fragments / Q000378:metabolism; D011485:Protein Binding; D015398:Signal Transduction; D013552:Swine; D014336:Troponin / Q000737:chemistry / Q000378:metabolism*; D019209:Troponin C
D002147:Calmodulin; D010446:Peptide Fragments; D014336:Troponin; D019209:Troponin C; D000252:Calcium-Transporting ATPases; D002118:Calcium