CORRELATION OF ACTOS1, MYOFIBRILLAR, AND MUSCLE-FIBER ATPASES

Our objective was to determine a good in vitro model for muscle fiber ATPase, and we compared the kinetics of Ca2+-activated myofibrils and cross-linked actoS1 in a buffer of physiological ionic strength. The myofibrils were cross-linked chemically to mimic the isometric condition of fibers or were un-cross-linked (the isotonic condition), and temperature perturbation was used to probe their ATPase mechanisms. At 4-degrees-C, we have already shown that the kinetics of cross-linked actoS1 and myofibrils (cross-linked or not) are similar: there were large P(i) bursts and k(cat) values of about 1 s-1, close to that obtained with fibers [Herrmann, C., Sleep, J., Chaussepied, P., Travers, F. & Barman, T. (1993) Biochemistry 32, 7255-7263]. So, at 4-degrees-C cross-linked actoS1 and myofibrils are equally good as models for fiber ATPase. At 20-degrees-C, this similarity vanishes: progress curves with the myofibrils (cross-linked or not) had large P(i) bursts, but with cross-linked actoS1, bursts could not be discerned. This shows that at 20-degrees-C the predominant steady-state state intermediates are ATP complexes with actoS1 but are products complexes with the myofibrils, as with fibers [Ferenczi, M. A. (1986) Biophys. J. 50, 471-477]. Further, the k(cat) values were different: 15.5 s-1 with cross-linked actoS1, 8.3 s-1 for myofibrils, and 3.5 s-1 for cross-linked myofibrils. With fibers, k(cat) = 3.3 s-1. These results show that cross-linked myofibrillar ATPase is a good model for muscle fibers contracting isometrically. Our results may also help to explain the Fenn effect, namely, that as the load on the system is increased so k(cat) decreases: cross-linked actoS1 --> myofibrils --> cross-linked myofibrils.