The regulatory proteins of the myofibril. Characterization and biological activity of the calcium-sensitizing factor (troponin A)

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Abstract
1. Electrophoretically homogeneous calcium-sensitizing factor was prepared from the troponin complex by chromatography successively on sulphoethyl-Sephadex and on diethyl-(2-hydroxypropyl)aminoethyl-Sephadex in 6m-urea. It is a protein containing 53% of polar amino acids, of which a net excess consists of acidic residues. 2. On gel filtration the calcium-sensitizing factor was shown to be the only myofibrillar protein that bound 45Ca2+ tightly in the presence of 2–6m-urea. 3. Calcium-sensitizing factor effectively neutralized the effect of the inhibitory factor on the ATPase activities of actomyosin systems. Tropomyosin was essential for the regulation, by changes in the Ca2+ concentration, of the neutralizing effect of calcium-sensitizing factor on the inhibitory factor. 4. Prolonged exposure to chelators of Ca2+ produced an irreversibly modified form of calcium-sensitizing factor of higher electrophoretic mobility at pH8.6. The modified form neutralized the inhibitory factor action but this property could no longer be controlled by the Ca2+ concentration in the presence of tropomysin. 5. The calcium-sensitizing factor and tropomyosin could be replaced by their carboxymethylated derivatives in the relaxing-protein system.