Laboratory of Mechanisms of Cell Motility, Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Avenue, St. Petersburg 194064, Russia.
Biochem Biophys Res Commun. 2011 Apr 1;407(1):197-201. doi: 10.1016/j.bbrc.2011.02.139. Epub 2011 Mar 3.
To understand the molecular mechanism by which the hypertrophic cardiomyopathy-causing Asp175Asn and Glu180Gly mutations in α-tropomyosin alter contractile regulation, we labeled recombinant wild type and mutant α-tropomyosins with 5-iodoacetamide-fluorescein and incorporated them into the ghost muscle fibers. The orientation and mobility of the probe were studied by polarized fluorimetry at different stages of the ATPase cycle. Multistep alterations in the position and mobility of wild type tropomyosin on the thin filaments during the ATP cycle were observed. Both mutations were found to shift tropomyosin strands further towards the open position and to change the affinity of tropomyosin for actin, with the effect of the Glu180Gly mutation being greater than Asp175Asn, showing an increase in the binding strong cross-bridges to actin during the ATPase cycle. These structural changes to the thin filament are likely to underlie the observed increased Ca(2+)-sensitivity caused by these mutations which initiates the disease remodeling.
为了理解导致肥厚型心肌病的α-原肌球蛋白 Asp175Asn 和 Glu180Gly 突变改变收缩调节的分子机制，我们用 5-碘乙酰胺-荧光素标记重组野生型和突变型α-原肌球蛋白，并将其掺入幽灵肌纤维中。通过在 ATP 酶循环的不同阶段进行偏振荧光研究，研究了探针的取向和迁移率。在 ATP 循环中观察到野生型原肌球蛋白在细肌丝上的位置和迁移率的多步改变。这两种突变都被发现使原肌球蛋白链进一步向开放位置移动，并改变原肌球蛋白与肌动蛋白的亲和力，Glu180Gly 突变的效果大于 Asp175Asn，表明在 ATP 酶循环中与肌动蛋白结合的强交联桥增加。这些细丝结构的变化可能是这些突变引起的 Ca(2+)敏感性增加的基础，这种敏感性引发了疾病重塑。
