Laboratorio de Fisicoquímica e Ingeniería de Proteínas, Departamento de Bioquímica, Facultad de Medicina, Universidad Nacional Autónoma de México, México.
Centro de Investigación en Dinámica Celular, IICBA, Universidad Autónoma del Estado de Morelos, México.
Biochim Biophys Acta Gen Subj. 2018 Jul;1862(7):1656-1666. doi: 10.1016/j.bbagen.2018.04.014. Epub 2018 Apr 15.
Solvent conditions modulate the expression of the amyloidogenic potential of proteins. In this work the effect of pH on the fibrillogenic behavior and the conformational properties of 6aJL2, a model protein of the highly amyloidogenic variable light chain λ6a gene segment, was examined. Ordered aggregates showing the ultrastructural and spectroscopic properties observed in amyloid fibrils were formed in the 2.0-8.0 pH range. At pH <3.0 a drastic decrease in lag time and an increase in fibril formation rate were found. In the 4.0-8.0 pH range there was no spectroscopic evidence for significant conformational changes in the native state. Likewise, heat capacity measurements showed no evidence for residual structure in the unfolded state. However, at pH <3.0 stability is severely decreased and the protein suffers conformational changes as detected by circular dichroism, tryptophan and ANS fluorescence, as well as by NMR spectroscopy. Molecular dynamics simulations indicate that acid-induced conformational changes involve the exposure of the loop connecting strands E and F. These results are compatible with pH-induced changes in the NMR spectra. Overall, the results indicate that the mechanism involved in the acid-induced increase in the fibrillogenic potential of 6aJL2 is profoundly different to that observed in κ light chains, and is promoted by localized conformational changes in a region of the protein that was previously not known to be involved in acid-induced light chain fibril formation. The identification of this region opens the potential for the design of specific inhibitors.
溶剂条件调节蛋白质的淀粉样形成潜力的表达。在这项工作中，研究了 pH 值对高度淀粉样变的可变轻链 λ6a 基因片段模型蛋白 6aJL2 的原纤维形成行为和构象特性的影响。在 2.0-8.0 pH 范围内形成了具有在淀粉样纤维中观察到的超微结构和光谱特性的有序聚集体。在 pH <3.0 时，发现滞后时间急剧缩短，原纤维形成速率增加。在 4.0-8.0 pH 范围内，在天然状态下没有发现显著构象变化的光谱证据。同样，热容测量没有显示在展开状态下存在残留结构的证据。然而，在 pH <3.0 时，稳定性严重降低，蛋白质发生构象变化，如圆二色性、色氨酸和 ANS 荧光以及 NMR 光谱检测到的变化。分子动力学模拟表明，酸诱导的构象变化涉及连接 E 和 F 链的环的暴露。这些结果与 NMR 光谱中 pH 诱导变化一致。总体而言，结果表明，酸诱导 6aJL2 原纤维形成潜力增加所涉及的机制与 κ 轻链中观察到的机制大不相同，并且由蛋白质中以前未知的区域的局部构象变化促进，该区域与酸诱导轻链原纤维形成有关。该区域的鉴定为设计特定抑制剂提供了潜力。
