School of Biosciences, University of Kent, Canterbury, Kent, United Kingdom.
Biochemistry. 2012 Dec 11;51(49):9880-90. doi: 10.1021/bi301323n. Epub 2012 Nov 30.
α-Tropomyosin (Tm) carrying hypertrophic cardiomyopathy mutation D175N or E180G was expressed in Escherichia coli. We have assembled dimers of two polypeptide chains in vitro that carry one (αα*) or two (αα) copies of the mutation. We found that the presence of the mutation has little effect on dimer assembly, thereby predicting that individuals heterozygous for the Tm mutations are likely to express both αα* and αα Tm. Depending on the expression level, the heterodimer may be the predominant form in individuals carrying the mutation. Thus, it is important to define differences in the properties of Tm molecules carrying one or two copies of the mutation. We examined the Tm homo- and heterodimer properties: actin affinity, thermal stability, calcium regulation of myosin subfragment 1 binding, and calcium regulation of myofibril force. We report that the properties of the heterodimer may be similar to those of the wild-type homodimer (actin affinity, thermal stability, D175N αα*), similar to those of the mutant homodimer (calcium sensitivity, D175N αα*), intermediate between the two (actin affinity, E180G αα*), or different from both (thermal stability, E180G αα*). Thus, the properties of the homodimer are not a completely reliable guide to the properties of the heterodimer.
α-原肌球蛋白（Tm）携带肥厚型心肌病突变 D175N 或 E180G 在大肠杆菌中表达。我们已在体外组装了两条多肽链的二聚体，它们携带一个（αα*）或两个（αα）突变拷贝。我们发现突变的存在对二聚体组装的影响很小，从而预测杂合子携带 Tm 突变的个体可能表达αα和αα* Tm。根据表达水平，杂二聚体可能是携带突变的个体中的主要形式。因此，定义携带一个或两个突变拷贝的 Tm 分子的特性差异很重要。我们检查了 Tm 同型和异型二聚体的特性：肌动蛋白亲和力、热稳定性、肌球蛋白亚片段 1 结合的钙调节以及肌原纤维力的钙调节。我们报告说，异二聚体的特性可能与野生型同型二聚体（肌动蛋白亲和力、热稳定性、D175N αα*）相似，与突变型同型二聚体（钙敏感性、D175N αα*）相似，介于两者之间（肌动蛋白亲和力、E180G αα*），或与两者都不同（热稳定性、E180G αα*）。因此，同型二聚体的特性并不是异型二聚体特性的完全可靠指南。
