DO LIGHT CHAIN EXTENSIONS ENHANCE MUSCLE POWER OUTPUT?

振荡功率产生是横纹肌的一个普遍特征。增强的振荡功率输出与肌球蛋白轻链氨基末端的延伸相关。脊椎动物心肌的肌球蛋白基本轻链和果蝇跳跃和飞行肌肉的调节轻链存在同源延伸。这些蛋白质延伸的确切功能尚不清楚，但初步结果表明，它们在收缩过程中增强了动力。我们的中心假设是，轻链延伸进行分子接触，有助于将肌球蛋白的运动亚单位预先定位在肌动蛋白的靶区附近，以实现最佳的相互作用和发电。将在小鼠心肌和苍蝇中创建轻链结构，以评估通过移除或取代被认为参与细丝相互作用的残基而减少功率输出的程度。将检验下列假设：1)缺乏基本轻链N-末端延伸的完整脑室条在亚最大钙水平下产生的振荡功率输出比具有全长轻链的条更低；2)基本轻链延伸仅在体内晶格间距时才显示其对功率的影响；3)轻链延伸通过与细丝的特定的、静电的相互作用而对功率产生影响；以及4)果蝇飞行和跳跃肌肉中调节轻链的N-端延伸的类似变化产生与在小鼠心脏中观察到的结构和功能表型相似的结构和功能表型。在完整的和去膜的(去皮的)制备中，将使用小角X射线衍射，并辅之以电子显微镜，测量肌球蛋白头部排列的丝间距、晶格顺序和指数。在通过渗透压缩恢复体内间隔的条件下，将在皮肤制剂中测量等长力、空载缩短速度和动态刚度(振荡功率输出)。这项研究将有助于理解和治疗能量输出受损的人类肌肉疾病。

Oscillatory power production is a general feature of striated muscle. Enhanced oscillatory power output is correlated with the presence of extensions of the amino terminus of myosin light chains. Homologous extensions exist in myosin essential light chains of vertebrate myocardium and the regulatory light chain of Drosophila jump and flight muscles. The exact function of these protein extensions is unknown, but preliminary results suggest that they augment power during contraction. Our central hypothesis is that the light chain extensions make molecular contacts that help pre-position the motor subunit of myosin near its target zone on actin for optimal interaction and power generation. Light chain constructs will be created in mouse myocardium and flies to assess the extent to which power output is diminished by removing or replacing residues thought to be involved in thin filament interaction. The following hypotheses will be tested: 1) intact ventricular strips lacking the essential light chain N-terminal extension produce lower oscillatory power output at submaximal calcium levels than strips with full length light chains, 2) the essential light chain extension exhibits its effect on power only at in vivo lattice spacing, 3) the light chain extension exerts its effect on power by specific, electrostatic interactions with the thin filament, and 4) comparable alterations of the N-terminal extension of the regulatory light chain in Drosophila flight and jump muscles produce structural and functional phenotypes comparable to those observed in mouse hearts. Interfilament spacing, lattice order, and indices of myosin head alignment will be measured in both intact and demembranated (skinned) preparations using low-angle X-ray diffraction, aided by electron microscopy. Isometric force, unloaded shortening velocity, and dynamic stiffness (oscillatory power output) will be measured in skinned preparations under conditions in which the otherwise swollen lattice is restored by osmotic compression to its in vivo spacing. This research will contribute to understanding and treating human muscle diseases in which power output is compromised.