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Hsp70 inhibits aggregation of Islet amyloid polypeptide by binding to the heterogeneous prenucleation oligomers
Biophysical Journal ( IF 3.4 ) Pub Date : 2021-02-01 , DOI: 10.1016/j.bpj.2020.12.019
Neeraja Chilukoti 1 , Timir Baran Sil 1 , Bankanidhi Sahoo 1 , S Deepa 1 , Sreelakshmi Cherakara 1 , Mithun Maddheshiya 1 , Kanchan Garai 1
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Molecular chaperone Hsp70 plays important roles in the pathology of amyloid diseases by inhibiting aberrant aggregation of proteins. However, biophysical mechanism of the interaction of Hsp70 with the intrinsically disordered proteins (IDPs) is unclear. Here, we report that Hsp70 inhibits aggregation of Islet amyloid polypeptide (IAPP) at substoichiometric concentrations under diverse solutions conditions, including in the absence of ATP. The inhibitory effect is strongest if Hsp70 is added in the beginning of aggregation but progressively less if added later, indicating role of Hsp70 in preventing nucleation of IAPP. However, ensemble measurement of the binding affinity suggests poor interactions between Hsp70 and IAPP. Therefore, we hypothesize that the interaction must involve a rare species, e.g., the oligomeric intermediates of IAPP. Size exclusion chromatography (SEC) and field flow fractionation are then used to fractionate the constituent species. Multiangle light scattering and fluorescence correlation spectroscopy measurements indicate that the dominant fraction in SEC contains a few nanomolar of Hsp70-IAPP complexes amidst several μmolar of free Hsp70. Using single particle two-color coincidence detection measurements, we detected a minor fraction that exhibits fluorescence bursts arising from heterogeneous oligomeric complexes of IAPP and Hsp70. Taken together, our results indicate that Hsp70 interacts poorly with the monomers but strongly with oligomers of IAPP. This is likely a generic feature of the interactions of Hsp70 chaperones with the amyloidogenic IDPs. While high-affinity interactions with the oligomers prevent aberrant aggregation, poor interaction with the monomers avert interference with the physiological functions of the IDPs.

中文翻译:

Hsp70 通过与异质核前寡聚体结合来抑制胰岛淀粉样蛋白多肽的聚集

分子伴侣 Hsp70 通过抑制蛋白质的异常聚集在淀粉样蛋白疾病的病理学中发挥重要作用。然而,Hsp70 与内在无序蛋白 (IDP) 相互作用的生物物理机制尚不清楚。在这里,我们报告 Hsp70 在各种溶液条件下(包括在没有 ATP 的情况下)以亚化学计量浓度抑制胰岛淀粉样蛋白多肽 (IAPP) 的聚集。如果在聚集开始时添加 Hsp70,则抑制作用最强,但如果稍后添加则逐渐减少,表明 Hsp70 在阻止 IAPP 成核中的作用。然而,结合亲和力的整体测量表明 Hsp70 和 IAPP 之间的相互作用较差。因此,我们假设相互作用必须涉及稀有物种,例如 IAPP 的寡聚中间体。然后使用尺寸排阻色谱 (SEC) 和场流分级分离组分物质。多角度光散射和荧光相关光谱测量表明,SEC 中的主要部分在几微摩尔的游离 Hsp70 中含有几纳摩尔的 Hsp70-IAPP 复合物。使用单粒子双色符合检测测量,我们检测到一小部分表现出由 IAPP 和 Hsp70 的异质寡聚复合物引起的荧光爆发。总之,我们的结果表明 Hsp70 与单体的相互作用很差,但与 IAPP 的低聚物相互作用很强。这可能是 Hsp70 伴侣与淀粉样蛋白 IDP 相互作用的一般特征。虽然与低聚物的高亲和力相互作用可防止异常聚集,
更新日期:2021-02-01
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