Kellermayer M S, Hinds T R, Pollack G H
Center for Bioengineering, University of Washington, Seattle 98195, USA.
Physiol Chem Phys Med NMR. 1995;27(3):167-78.
We have previously demonstrated in vitro actin movement at nanomolar adenosine triphosphate (ATP) levels using heavy meromyosin from skeletal muscle. In the present work we tested whether the motility at nonomolar ATP-concentrations could be supported by cardiac myosin as well. Actomyosin (skeletal actin and bovine ventricular myosin) was pretreated in the in vitro motility assay with 1 mM ATP; subsequently, the ATP level was reduced by multiple rigor-solution washes. By the final rigor-solution wash, the ATP concentration, monitored by the luciferin-luciferase assay, dropped to the order of 100 nM. Even at this low ATP level actin-filament movement remained in evidence. This was in marked contrast to the situation where ATP concentration was gradually increased from zero; in the latter, filament movement began only as ATP levels exceeded 1-2 microM. The difference indicates that potential energy is stored during the initial ATP treatment, and utilized later as the free ATP falls below micromolar levels. Although the velocity of cardiac myosin-supported movement was only one fourth of that of skeletal myosin, both myosins supported actin movement down to similar ATP concentrations. The similarity in response of the two myosins to ATP implies a similar degree of potential energy storage. Given the significantly different specific ATPase activities, however, it appears that the mechanism of potential energy storage and release involves factors different from those involved in the release of chemical energy by the myosin ATPase.
我们之前曾利用骨骼肌的重酶解肌球蛋白，在体外纳摩尔浓度的三磷酸腺苷（ATP）水平下证明了肌动蛋白的运动。在本研究中，我们测试了心肌肌球蛋白是否也能在纳摩尔ATP浓度下支持这种运动。在体外运动分析中，将肌动球蛋白（骨骼肌肌动蛋白和牛心室肌球蛋白）用1 mM ATP进行预处理；随后，通过多次用僵直溶液洗涤来降低ATP水平。到最后一次用僵直溶液洗涤时，通过荧光素 - 荧光素酶测定法监测的ATP浓度降至100 nM左右。即使在如此低的ATP水平下，肌动蛋白丝的运动仍然明显。这与ATP浓度从零逐渐增加的情况形成了显著对比；在后者中，只有当ATP水平超过1 - 2 microM时，丝的运动才开始。这种差异表明，势能在最初的ATP处理过程中被储存起来，随后在游离ATP降至微摩尔水平以下时被利用。尽管心肌肌球蛋白支持的运动速度仅为骨骼肌肌球蛋白的四分之一，但两种肌球蛋白都能支持肌动蛋白运动至相似的ATP浓度。两种肌球蛋白对ATP反应的相似性意味着势能储存程度相似。然而，鉴于它们的比ATP酶活性有显著差异，势能储存和释放的机制似乎涉及与肌球蛋白ATP酶释放化学能所涉及的因素不同的因素。
