Thermodynamic Stability and Aggregation Kinetics of EF Helix and EF Loop Variants of Transthyretin.
Thermodynamic Stability and Aggregation Kinetics of EF Helix and EF Loop Variants of Transthyretin.
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DOI：
10.1021/acs.biochem.1c00073
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发表时间：
2021-03-16
期刊：
影响因子：
2.9
通讯作者：
Wright PE
中科院分区：
文献类型：
--
作者：
Ferguson JA;Sun X;Dyson HJ;Wright PE
转甲状腺素（TTR）的错误折叠和聚集与淀粉样蛋白疾病有关。淀粉样变性发生时，TTR同聚体解离成易于聚集的单体，自组装成淀粉样蛋白。在家族性甲状腺转蛋白淀粉样变性中，遗传性氨基酸替换破坏TTR的稳定性并促进其聚集。在本研究中，我们利用19F-NMR确定了EF螺旋（Y78F、K80D、K80E和A81T）和EF环（G83R和I84S）突变对TTR四聚体和单体聚集动力学和稳定性的影响。EF区作为一个支架，稳定了四聚体的强二聚体和弱二聚体界面的相互作用，是一簇致病性突变的位点。K80D和K80E是非自然突变体，它们破坏EF螺旋的稳定性，并在中性pH下产生四聚体和单体的平衡混合物，这为在非变性条件下确定四聚体组装的热力学参数提供了独特的机会。在研究的致病性突变体中，只有A81T在中性ph下形成可观的单体。实时19F NMR测量显示，致病性Y78F突变通过破坏四聚体和单体物种的稳定来加速聚集。致病突变A81T、G83R和I84S破坏了单体的稳定性，并通过破坏谢尔曼螺旋C-capping基序增加了其聚集率。这些研究为影响四聚体或单体稳定性的相对微妙的突变促进TTR进入解离-聚集途径的机制提供了新的见解。
Misfolding and aggregation of transthyretin (TTR) is linked to amyloid disease. Amyloidosis occurs when the TTR homotetramer dissociates into aggregation-prone monomers that self-assemble into amyloid. In familial transthyretin amyloidosis, hereditary amino acid substitutions destabilize TTR and promote aggregation. In the present work, we used 19F-NMR to determine the effect of mutations, in the EF helix (Y78F, K80D, K80E, and A81T) and EF loop (G83R and I84S) on the aggregation kinetics and stability of the TTR tetramer and monomer. The EF region acts as a scaffold that stabilizes interactions in both the strong and weak dimer interfaces of the tetramer and is the site of a cluster of pathogenic mutations. K80D and K80E are non-natural mutants that destabilize the EF helix and yield an equilibrium mixture of tetramer and monomer at neutral pH, providing a unique opportunity to determine the thermodynamic parameters for tetramer assembly under non-denaturing conditions. Of the pathogenic mutants studied, only A81T formed appreciable monomer at neutral pH. Real-time 19F NMR measurements showed that the pathogenic Y78F mutation accelerates aggregation by destabilizing both the tetrameric and monomeric species. The pathogenic mutations A81T, G83R, and I84S destabilize the monomer and increase its aggregation rate by disrupting a Schellman helix C-capping motif. These studies provide new insights into the mechanism by which relatively subtle mutations that affect tetramer or monomer stability promote entry of TTR into the dissociation-aggregation pathway.
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影响因子：
5.6
作者：
Palaninathan, Satheesh K.;Mohamedmohaideen, Nilofar N.;Sacchettini, James C.
通讯作者：
Sacchettini, James C.
影响因子：
2.9
作者：
COLON, W;KELLY, JW
通讯作者：
KELLY, JW
影响因子：
2.9
作者：
Azevedo, Estefania P. C.;Pereira, Humberto M.;Palhano, Fernando L.
通讯作者：
Palhano, Fernando L.
影响因子：
5.6
作者：
Pasquato, Nicola;Berni, Rodolfo;Zanotti, Giuseppe
通讯作者：
Zanotti, Giuseppe
影响因子：
8
作者：
HUYGHUESDESPOINTES, BMP;SCHOLTZ, JM;BALDWIN, RL
通讯作者：
BALDWIN, RL