DO LIGHT CHAIN EXTENSIONS ENHANCE MUSCLE POWER OUTPUT?

振荡发力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端延伸的存在有关。同源延伸存在于脊椎动物心肌肌球蛋白必需轻链和果蝇跳飞肌调控轻链中。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力量。我们的中心假设是，轻链延伸使分子接触有助于将肌凝蛋白的运动亚基预先定位在肌动蛋白上的靶区附近，以实现最佳的相互作用和发电。轻链结构将在小鼠心肌和苍蝇中创建，以评估通过去除或替换被认为参与细丝相互作用的残留物来减少功率输出的程度。以下假设将被检验：1)缺乏基本轻链n端延伸的完整心室条带在亚最大钙水平下产生的振荡功率输出低于具有全长轻链的条带，2)基本轻链延伸仅在体内晶格间距下对功率产生影响，3)轻链延伸通过与细丝的特定静电相互作用对功率产生影响。4)果蝇飞行和跳跃肌肉中调节轻链n端延伸的类似改变产生的结构和功能表型与小鼠心脏中观察到的相似。在电子显微镜的辅助下，使用低角度x射线衍射测量完整和去膜（剥皮）制备中的丝间间距，晶格顺序和肌球蛋白头部排列指数。等长力、卸载缩短速度和动态刚度（振荡功率输出）将在剥皮制剂中测量，在这种条件下，否则膨胀的晶格通过渗透压缩恢复到其体内间距。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.