Department of Applied Chemistry, Faculty of Fundamental Engineering, Nippon Institute of Technology, Miyashiro, Saitama 345-8501, Japan.
Graduate School of Environmental Symbiotic System Major, Nippon Institute of Technology, Miyashiro, Saitama 345-8501, Japan.
Biochemistry. 2020 Jun 16;59(23):2194-2202. doi: 10.1021/acs.biochem.0c00203. Epub 2020 Jun 5.
Tropomyosin (Tpm) is a two-stranded parallel α-helical coiled-coil protein, and studying its structure is crucial for understanding the nature of coiled coils. Previously, we found that the N-terminal half of the human skeletal muscle α-Tpm (α-Tpm 140) was less structurally stable in the presence of phosphate ions than the coiled-coil protein carrier (CCPC) 140 variant with 18 mutated residues, in which all amino acid residues located at the interface between the two α-helices were completely conserved. A classical hypothesis explains that interhelical interactions stabilize the coiled-coil structure. In this study, we tested the hypothesis that the structural stability of Tpm and its variant is governed by the binding of multivalent ions that form a bridge between charged side chains located at positions , , and of the heptad repeat on a single α-helical chain. We found that the structural stability of α-Tpm 140 and CCPC 140 markedly increased upon addition of divalent cations and divalent anions, respectively. We also clarified that the structural stability of the α-Tpm 140/CCPC 140 heteromeric coiled-coil molecule was governed by the stability of a less stable α-helical chain. These results demonstrated that the entire structural stability of Tpm is determined by the stability of a single α-helix. Our findings provide new insights into the study of the structure of coiled-coil proteins.
原肌球蛋白（Tropomyosin，Tpm）是一种双股平行α-螺旋卷曲螺旋蛋白，研究其结构对于理解卷曲螺旋的本质至关重要。我们之前发现，人骨骼肌α-Tpm（α-Tpm 140）的 N 端半段在磷酸盐离子存在下的结构稳定性不如含有 18 个突变残基的卷曲螺旋蛋白载体（CCPC）140 变体，其中位于两个α-螺旋之间界面的所有氨基酸残基都完全保守。一个经典假说解释说，螺旋间相互作用稳定卷曲螺旋结构。在这项研究中，我们检验了这样一个假设，即 Tpm 及其变体的结构稳定性受多价离子结合的控制，这些离子在单条α-螺旋链上的位置 、 和 处的带电侧链之间形成桥。我们发现，α-Tpm 140 和 CCPC 140 的结构稳定性分别在添加二价阳离子和二价阴离子后显著增加。我们还阐明了，α-Tpm 140/CCPC 140 异源卷曲螺旋分子的结构稳定性受不稳定α-螺旋链稳定性的控制。这些结果表明，Tpm 的整个结构稳定性由单个α-螺旋的稳定性决定。我们的研究结果为卷曲螺旋蛋白结构的研究提供了新的见解。
