Destabilization of human transthyretin leads to its aggregation into amyloid fibrils, which causes a rare, progressive and fatal systemic disorder called ATTR amyloidosis. By contrast, murine transthyretin is known to be very stable and therefore does not aggregate into amyloid fibrils in vivo or in vitro. We examined the hydrophobic residues responsible for the high-stability and low-aggregation properties of murine transthyretin using site-directed mutagenesis. Urea-induced unfolding and thioflavin T fluorescence aggregation assay revealed that Leu73 of murine transthyretin largely contributes to its high stability and low aggregation properties: the I73L mutation stabilized human transthyretin, while the L73I mutation destabilized murine transthyretin. In addition, the I26V/I73L mutation stabilized the amyloidogenic V30M mutant of human transthyretin to the same degree as the suppressor mutation T119M, which protects transthyretin against amyloid fibril aggregation. The I73L mutation resulted in no significant differences in the overall structure of the transthyretin tetramer or the contacts of side-chains in the hydrophobic core of the monomer. We also found that Leu73 of murine transthyretin is conserved in many mammals, while Ile73 of human transthyretin is conserved in monkeys and cats. These studies will provide new insights into the stability and aggregation properties of transthyretin from various mammals.

中文翻译：

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疏水残基 Leu73 对鼠甲状腺素运载蛋白的高稳定性和低聚集特性至关重要

人转甲状腺素蛋白的不稳定导致其聚集成淀粉样蛋白原纤维，从而导致一种罕见的、进行性和致命的全身性疾病，称为 ATTR 淀粉样变性。相比之下，已知鼠运甲状腺素蛋白非常稳定，因此不会在体内或体外聚集成淀粉样蛋白原纤维。我们使用定点诱变检查了负责鼠甲状腺素运载蛋白的高稳定性和低聚集特性的疏水残基。尿素诱导的去折叠和硫黄素 T 荧光聚集测定表明，鼠甲状腺素运载蛋白的 Leu73 很大程度上有助于其高稳定性和低聚集特性：I73L 突变稳定了人运甲状腺素蛋白，而 L73I 突变使鼠运甲状腺素蛋白不稳定。此外，I26V/I73L 突变使人转甲状腺素蛋白的淀粉样蛋白生成 V30M 突变体稳定到与抑制突变 T119M 相同的程度，T119M 保护转甲状腺素蛋白免受淀粉样蛋白原纤维聚集。I73L 突变导致转甲状腺素蛋白四聚体的整体结构或单体疏水核心中侧链的接触没有显着差异。我们还发现鼠甲状腺素运载蛋白的 Leu73 在许多哺乳动物中是保守的，而人甲状腺素运载蛋白的 Ile73 在猴子和猫中是保守的。这些研究将为来自各种哺乳动物的转甲状腺素蛋白的稳定性和聚集特性提供新的见解。I73L 突变导致转甲状腺素蛋白四聚体的整体结构或单体疏水核心中侧链的接触没有显着差异。我们还发现鼠甲状腺素运载蛋白的 Leu73 在许多哺乳动物中是保守的，而人甲状腺素运载蛋白的 Ile73 在猴子和猫中是保守的。这些研究将为来自各种哺乳动物的转甲状腺素蛋白的稳定性和聚集特性提供新的见解。I73L 突变导致转甲状腺素蛋白四聚体的整体结构或单体疏水核心中侧链的接触没有显着差异。我们还发现鼠甲状腺素运载蛋白的 Leu73 在许多哺乳动物中是保守的，而人甲状腺素运载蛋白的 Ile73 在猴子和猫中是保守的。这些研究将为来自各种哺乳动物的转甲状腺素蛋白的稳定性和聚集特性提供新的见解。