A study of the kinetics of the fibrillar adenosine triphosphatase of rabbit skeletal muscle

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Abstract
The time course of the hydrolysis of adenosine triphosphate by rabbit myofibrils has been studied at temperatures of 0[degree], 18.5[degree] and 35[degree] and at ionic strengths (I) between 0.04 and 0.25, with Ca2+ ions, Mg2+ ions or a mixture of both as activators. It has been found that the products inhibit the reaction, the inhibition being more pronounced in the calcium than in the magnesium system. The combined relative inhibition constant, Ks/Kpi + Ks/KADP, varies in the former system from [image] 1.0 to 0[degree] to about 2.2 at 35[degree] for I 0.042, and from [image] 0.8 to [image] 0.7 for the same temperature range at I 0-250. In the magnesium system, no inhibition by products occurs at I [greater than or equ 0*150 at the two lower temperatures, but the inhibition constant at I[less than or equal 0.150 reaches values of[image] 1.00 at 35[degree]. When allowance is made for inhibition by the products in the calcium system, the so-called early explosive phase of the reaction is either absent or restricted to < 2 sec.; in the magnesium system a short explosive phase is always detectable, lasting for < 5 sec. at 18[degree] or above, but is either absent or much reduced in fibrils supercontracted by pretreatment with adenosine triphosphate. It is suggested that the declining velocity during the explosive phase in this system is entirely due to reduction in the number of active enzyme sites during the process of supercontraction. The progress curves of the calcium system and the product inhibition constants characteristic of it can be entirely converted into those characteristic of the magnesium system by addition of Mg2+ ions to one-seventh of the concentration of the Ca2+ ions. The explosive phase of the magnesium system and the characteristic supercontraction appear concomitantly. It is deduced that the Mg2+ ion is always the dominant ion when both ions are added together. It follows that it is the characteristics of the magnesium system which must be taken into account in vivo, where most of the Mg2+ ions are free and most of the Ca2+ ions are bound to protein. The energy of activation of the hydrolysis process is [image] 11 kcal./mole in the calcium system, whereas it always exceeds 15 kcal./mole in the magnesium system, and in vivo is > 20 kcal./mole. It is suggested that these large differences are due to participation of sites on both the actin and the myosin filaments in the magnesium-activated process, but of only myosin sites in the calcium-activated.