scholarly journals Utilization of troponin C as a model calcium-binding protein for mapping of the calmodulin-binding sites of caldesmon

1997 ◽  
Vol 321 (3) ◽  
pp. 873-878 ◽  
Author(s):  
Alexei A. POLYAKOV ◽  
Nikolai B. GUSEV
Keyword(s):  
Skeletal Muscle ◽  
Atpase Activity ◽  
Binding Sites ◽  
Calcium Binding ◽  
Binding Protein ◽  
Troponin C ◽  
Rabbit Skeletal Muscle ◽  
Actomyosin Atpase ◽  
Calmodulin Binding ◽  
Central Helix

Troponin C, a structural analogue of calmodulin, was used for mapping the calmodulin-binding sites of caldesmon. The apparent Kd values for the formation of the caldesmonŐcalcium-binding-protein complex as determined by native gel electrophoresis were 0.5, 1.2 and 3.9 ƁM for calmodulin, rabbit skeletal muscle troponin C and bovine cardiac troponin C respectively. Troponin C induced a 4Ő6 nm blue shift of the Trp fluorescence of caldesmon without affecting the amplitude of fluorescence. In the presence of Ca2+, troponin C induced partial displacement of caldesmon from actinŐtropomyosin complexes. Addition of 5,5ƀ-dithiobis(nitrobenzoic) acid to an equimolar complex of caldesmon and troponin C induced disulphide cross-linking between Cys-98 of rabbit skeletal muscle troponin C and the single Cys residue of duck gizzard caldesmon, located in a position analogous to Cys-580 of the chicken gizzard protein. The cross-linked caldesmonŐtroponin C complex was ineffective in inhibiting actomyosin ATPase activity. It is concluded that Cys-580 of caldesmon can be located close to both the central helix of calcium-binding proteins and the C-terminal domain of actin. This may be important for the regulation of actomyosin ATPase activity by caldesmon.

scholarly journals Calcium-Binding Properties of Sarcoplasmic Reticulum As Influenced by ATP, Caffeine, Quinine, and Local Anesthetics

1968 ◽  
Vol 52 (4) ◽  
pp. 622-642 ◽  
Author(s):  
Arselio P. Carvalho
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Binding Sites ◽  
Local Anesthetics ◽  
Calcium Binding ◽  
Rabbit Skeletal Muscle ◽  
Cellular Membranes ◽  
Binding Properties ◽  
45Ca Fluxes

Calcium retained at binding sites of the sarcoplasmic reticulum membranes isolated from rabbit skeletal muscle requires 10-5 - 10-4 M ATP to exchange with 45Ca added to the medium. The ATP requirement for Ca exchangeability was observed with respect to the "intrinsic" Ca of the reticulum membranes and the fraction of Ca that is "actively" bound in the presence of ATP. Furthermore, a concentration of free Ca in the medium higher than 10-8 M is required for ATP to promote Ca exchangeability. This exchangeability is not influenced by caffeine, quinine, procaine, and tetracaine, and Ca that is either nonexchangeable (in the absence of ATP) or exchangeable (in the presence of ATP) is released by 1–5 mM quinine or tetracaine, but neither caffeine (6 mM) nor procaine (2–5 mM) has this effect. Quinine or tetracaine also releases Ca and Mg bound passively to the reticulum membranes. A possible role of ATP in maintaining the integrity of cellular membranes is discussed, and the effects of caffeine, quinine, and of local anesthetics on the binding of Ca by the isolated reticulum are related to the effects of these agents on 45Ca fluxes and on the twitch output observed in whole muscles.

scholarly journals Concerted Action of the High Affinity Calcium Binding Sites in Skeletal Muscle Troponin C

1995 ◽  
Vol 270 (17) ◽  
pp. 9770-9777 ◽  
Author(s):  
Martha M. Sorenson ◽  
Ana C. R. da Silva ◽  
Claudia S. Gouveia ◽  
Valeria P. Sousa ◽  
Wanda Oshima ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Binding Sites ◽  
Calcium Binding ◽  
Troponin C ◽  
Concerted Action ◽  
High Affinity ◽  
Calcium Binding Sites

scholarly journals Calcium-Binding Properties of Sarcoplasmic Reticulum As Influenced by ATP, Caffeine, Quinine, and Local Anesthetics

1968 ◽  
Vol 52 (3) ◽  
pp. 622-642 ◽  
Author(s):  
Arselio P. Carvalho
Keyword(s):  
Skeletal Muscle ◽  
Sarcoplasmic Reticulum ◽  
Binding Sites ◽  
Local Anesthetics ◽  
Calcium Binding ◽  
Rabbit Skeletal Muscle ◽  
Cellular Membranes ◽  
Binding Properties ◽  
45Ca Fluxes

Calcium retained at binding sites of the sarcoplasmic reticulum membranes isolated from rabbit skeletal muscle requires 10-5 – 10-4 M ATP to exchange with 45Ca added to the medium. The ATP requirement for Ca exchangeability was observed with respect to the "intrinsic" Ca of the reticulum membranes and the fraction of Ca that is "actively" bound in the presence of ATP. Furthermore, a concentration of free Ca in the medium higher than 10-8 M is required for ATP to promote Ca exchangeability. This exchangeability is not influenced by caffeine, quinine, procaine, and tetracaine, and Ca that is either nonexchangeable (in the absence of ATP) or exchangeable (in the presence of ATP) is released by 1–5 mM quinine or tetracaine, but neither caffeine (6 mM) nor procaine (2–5 mM) has this effect. Quinine or tetracaine also releases Ca and Mg bound passively to the reticulum membranes. A possible role of ATP in maintaining the integrity of cellular membranes is discussed, and the effects of caffeine, quinine, and of local anesthetics on the binding of Ca by the isolated reticulum are related to the effects of these agents on 45Ca fluxes and on the twitch output observed in whole muscles.

scholarly journals Calmodulin-binding proteins from brain and other tissues

1979 ◽  
Vol 183 (2) ◽  
pp. 285-295 ◽  
Author(s):  
R J A Grand ◽  
S V Perry
Keyword(s):  
Skeletal Muscle ◽  
Binding Proteins ◽  
Binding Protein ◽  
Gel Filtration ◽  
Bovine Brain ◽  
Rabbit Skeletal Muscle ◽  
High Yield ◽  
Rabbit Brain ◽  
Dependent Manner ◽  
Calmodulin Binding

The calmodulin contents of rabbit brain, lung, kidney and liver, of bovine aorta and uterus, and of chicken gizzard have been determined. 2. The calmodulin in all of these tissues has been shown to be present in the form of very stable complexes with several other proteins. 3. A calmodulin-binding protein of mol.wt. 22 000 has been purified in high yield from bovine brain. It has been shown to interact with calmodulin and rabbit skeletal-muscle troponin C in a Ca2+-dependent manner. 4. The 22 000-mol.wt. protein inhibits the activation of bovine brain phosphodiesterase by calmodulin, but has very little affect on the activation of myosin light-chain kinase. 5. Calmodulin-binding proteins of mol.wts. 140000, 77000 and 61000 have also been partially purified from rabbit brain by affinity chromatography and have been shown to interact in a Ca2+-dependent manner with calmodulin. 6. The apparent molecular weights of the calmodulin-calmodulin-binding protein complexes, determined by gel filtration in the presence of 6M-urea, have been shown to be similar for most of the mammalian tissues examined. 7. By using 125I-labelled calmodulin, similar complexes have been demonstrated in rabbit skeletal muscle, although they are present at much lower concentrations.

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