McConnell Mark, Tal Grinspan Lauren, Williams Michael R, Lynn Melissa L, Schwartz Benjamin A, Fass Ofer Z, Schwartz Steven D, Tardiff Jil C
Department of Biomedical Engineering, University of Arizona , Tucson, Arizona 85721, United States.
Department of Medicine, Columbia University Medical Center , New York, New York 10032, United States.
Biochemistry. 2017 Jul 5;56(26):3403-3413. doi: 10.1021/acs.biochem.7b00266. Epub 2017 Jun 21.
The progression of genetically inherited cardiomyopathies from an altered protein structure to clinical presentation of disease is not well understood. One of the main roadblocks to mechanistic insight remains a lack of high-resolution structural information about multiprotein complexes within the cardiac sarcomere. One example is the tropomyosin (Tm) overlap region of the thin filament that is crucial for the function of the cardiac sarcomere. To address this central question, we devised coupled experimental and computational modalities to characterize the baseline function and structure of the Tm overlap, as well as the effects of mutations causing divergent patterns of ventricular remodeling on both structure and function. Because the Tm overlap contributes to the cooperativity of myofilament activation, we hypothesized that mutations that enhance the interactions between overlap proteins result in more cooperativity, and conversely, those that weaken interaction between these elements lower cooperativity. Our results suggest that the Tm overlap region is affected differentially by dilated cardiomyopathy-associated Tm D230N and hypertrophic cardiomyopathy-associated human cardiac troponin T (cTnT) R92L. The Tm D230N mutation compacts the Tm overlap region, increasing the cooperativity of the Tm filament, contributing to a dilated cardiomyopathy phenotype. The cTnT R92L mutation causes weakened interactions closer to the N-terminal end of the overlap, resulting in decreased cooperativity. These studies demonstrate that mutations with differential phenotypes exert opposite effects on the Tm-Tn overlap, and that these effects can be directly correlated to a molecular level understanding of the structure and dynamics of the component proteins.
遗传性心肌病从蛋白质结构改变到疾病临床表现的进展尚未得到充分理解。阻碍深入了解其机制的主要障碍之一仍然是缺乏关于心肌肌节内多蛋白复合物的高分辨率结构信息。一个例子是细肌丝的原肌球蛋白（Tm）重叠区域，它对心肌肌节的功能至关重要。为了解决这个核心问题，我们设计了实验和计算相结合的方法来表征Tm重叠区域的基线功能和结构，以及导致不同心室重塑模式的突变对结构和功能的影响。由于Tm重叠区域有助于肌丝激活的协同性，我们假设增强重叠蛋白之间相互作用的突变会导致更高的协同性，反之，削弱这些元件之间相互作用的突变会降低协同性。我们的结果表明，扩张型心肌病相关的Tm D230N和肥厚型心肌病相关的心型肌钙蛋白T（cTnT）R92L对Tm重叠区域的影响不同。Tm D230N突变使Tm重叠区域紧凑，增加了Tm细丝的协同性，导致扩张型心肌病表型。cTnT R92L突变导致重叠区域N端附近的相互作用减弱，从而降低协同性。这些研究表明，具有不同表型的突变对Tm-Tn重叠区域产生相反的影响，并且这些影响可以直接与对组成蛋白的结构和动力学的分子水平理解相关联。
