Phosphorylation of cardiac native tropomyosin and troponin: inhibitory effect of actomyosin and possible presence of endogenous myofibrillar-located cyclic-AMP-dependent protein kinase

Canine cardiac native tropomyosin which inhibits Mg²⁺-ATPase and superprecipitation of desensitized actomyosin in the presence of EGTA contains closely associated protein kinase, as manifested by phosphorylation and stimulation by cyclic-AMP. Native tropomyosin functions as a substrate for the endogenous protein kinase. Native actomyosin and desensitized actomyosin showed no significant cyclic-AMP-dependent or independent phosphorylation either in the presence or absence of added protein kinase. Native tropomyosin was fractionated into components by using hydroxylapatite column chromatography. Of the three components, Peak II (troponin) bound Ca²⁺; sodium dodecyl sulfate (SDS) gel electrophoresis revealed four subunits; and Peak III (tropomyosin) was a homogeneous protein and did not bind Ca²⁺. Troponin (Peak II) added to tropomyosin (Peak III) in appropriate concentrations inhibited superprecipitation of desensitized actomyosin in the absence of Ca²⁺. Peak II (troponin) phosphorylation was catalyzed by bovine cardiac protein kinase. The phosphorylation was markedly stimulated by cyclic-AMP. Peak III (tropomyosin) was not phosphorylated by protein kinase in the presence or absence of cyclic-AMP.The addition of desensitized actomyosin completely inhibited the phosphorylation of native tropomyosin. The data suggest the possible involvement of cyclic-AMP and protein kinase in modulating cardiac contraction and relaxation through and action on troponin.