Institute of Cytology, Russian Academy of Sciences, 4 Tikhoretsky Av., 194064 St. Petersburg, Russia.
Department of Biophysics, Faculty of Biology, Saint Petersburg State University, 7/9 Universitetskaya Emb., 199034 St. Petersburg, Russia.
Int J Mol Sci. 2020 Oct 14;21(20):7590. doi: 10.3390/ijms21207590.
We have used the technique of polarized microfluorimetry to obtain new insight into the pathogenesis of skeletal muscle disease caused by the GlnPro substitution in β-tropomyosin (Tpm2.2). The spatial rearrangements of actin, myosin and tropomyosin in the single muscle fiber containing reconstituted thin filaments were studied during simulation of several stages of ATP hydrolysis cycle. The angular orientation of the fluorescence probes bound to tropomyosin was found to be changed by the substitution and was characteristic for a shift of tropomyosin strands closer to the inner actin domains. It was observed both in the absence and in the presence of troponin, Ca and myosin heads at all simulated stages of the ATPase cycle. The mutant showed higher flexibility. Moreover, the GlnPro substitution disrupted the myosin-induced displacement of tropomyosin over actin. The irregular positioning of the mutant tropomyosin caused premature activation of actin monomers and a tendency to increase the number of myosin cross-bridges in a state of strong binding with actin at low Ca.
我们使用偏振微荧光技术深入了解了β-原肌球蛋白（Tpm2.2）中的 GlnPro 取代引起的骨骼肌疾病的发病机制。在模拟 ATP 水解循环的几个阶段期间，研究了含有重组细肌丝的单个肌纤维中肌动蛋白、肌球蛋白和原肌球蛋白的空间重排。发现与原肌球蛋白结合的荧光探针的角取向因取代而发生变化，并且特征是原肌球蛋白链向更靠近肌动蛋白内部结构域的方向移动。在缺乏肌钙蛋白、Ca 和肌球蛋白头部的情况下以及在模拟 ATP 酶循环的所有阶段都观察到了这种情况。突变体显示出更高的柔韧性。此外，GlnPro 取代破坏了肌球蛋白诱导的原肌球蛋白在肌动蛋白上的位移。突变体原肌球蛋白的不规则定位导致肌动蛋白单体过早激活，并倾向于在低 Ca 下增加与肌动蛋白强烈结合的肌球蛋白交联的数量。
