REDUCED POSITIVE FEEDBACK-REGULATION BETWEEN MYOSIN CROSSBRIDGE AND CARDIAC TROPONIN-C IN FAST SKELETAL MYOFIBRILS

Several studies have shown that substitution of cardiac troponin C int o fast skeletal muscle causes a marked reduction in cooperativity of C a2+-activation of both myofibrillar ATPase and tension development. To clarify the underlying mechanisms, in the present study, Ca2+ binding to cardiac troponin C inserted into fast skeletal myofibrils was meas ured. Two classes of binding sites with different affinities (classes 1 and 2) were clearly identified, which were equivalent stoichiometric ally to the two high-affinity sites (sites III and IV) and a single lo w-affinity site (site II) of troponin C, respectively, Ca2+ binding to class-2 sites and Ca2+-activation of myofibrillar ATPase occurred in roughly the same Ca2+ concentration range, indicating that site II is responsible for Ca2+-regulation. Myosin crossbridge interactions with actin, both in the presence and absence of ATP, enhanced the Ca2+ bind ing affinity of only class-2 sites. These effects of myosin crossbridg es, however, were much smaller than the effects on the Ca2+ binding to the low-affinity sites of fast skeletal troponin C, which are respons ible for regulating fast skeletal myofibrillar ATPase. These findings provide strong evidence that the reduction in the cooperative response to Ca2+ upon substituting cardiac troponin C into fast skeletal myofi brils is due to a decrease in the positive feedback interaction betwee n myosin crossbridge attachment and Ca2+ binding to the regulatory sit e of troponin C.