DO LIGHT CHAIN EXTENSIONS ENHANCE MUSCLE POWER OUTPUT?

振荡功率产生是横纹肌的一般特征。 增强的振荡功率输出与肌球蛋白轻链氨基末端延伸的存在相关。 同源延伸存在于脊椎动物心肌的肌球蛋白必需轻链和果蝇跳跃和飞行肌肉的调节轻链中。 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加了力量。 我们的中心假设是，轻链延伸的分子接触，有助于预先定位肌球蛋白的马达亚基附近的肌动蛋白的目标区域的最佳相互作用和发电。 将在小鼠心肌和果蝇中创建轻链构建体，以评估通过去除或替换被认为参与细丝相互作用的残基而降低功率输出的程度。 将检验以下假设：1）缺乏必需轻链N-末端延伸的完整心室条在次最大钙水平下比具有全长轻链的条产生更低的振荡功率输出，2）必需轻链延伸仅在体内晶格间距下表现出其对功率的影响，3）轻链延伸通过特异性，静电相互作用的细丝，和4）在果蝇的飞行和跳跃肌肉的调节轻链的N-末端延伸的类似的改变产生的结构和功能表型的小鼠心脏中观察到的。 在电子显微镜辅助下，使用低角X射线衍射，在完整和脱膜（去皮）制剂中测量肌丝间距、晶格有序和肌球蛋白头部排列指数。 将在经皮制剂中测量等距力、无载荷缩短速度和动态刚度（振荡功率输出），测量条件为通过渗透压缩将其他溶胀晶格恢复至其体内间距。 这项研究将有助于理解和治疗功率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.