Department of Molecular Genetics and Microbiology, Duke University, Durham, North Carolina 27710, United States.
Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, United States.
ACS Chem Biol. 2023 Mar 17;18(3):549-560. doi: 10.1021/acschembio.2c00893. Epub 2023 Feb 15.
Protein aggregation is a hallmark of the polyglutamine diseases. One potential treatment for these diseases is suppression of polyglutamine aggregation. Previous work identified the cellular slime mold as being naturally resistant to polyglutamine aggregation. Further work identified serine-rich chaperone protein 1 (SRCP1) as a protein that is both necessary in and sufficient in human cells to suppress polyglutamine aggregation. Therefore, understanding how SRCP1 suppresses aggregation may be useful for developing therapeutics for the polyglutamine diseases. Here we utilized a protein modeling approach to generate predictions of SRCP1's structure. Using our best-fit model, we generated mutants that were predicted to alter the stability of SRCP1 and tested these mutants' stability in cells. Using these data, we identified top models of SRCP1's structure that are consistent with the C-terminal region of SRCP1 forming a β-hairpin with a highly dynamic N-terminal region. We next generated a series of peptides that mimic the predicted β-hairpin and validated that they inhibit aggregation of a polyglutamine-expanded mutant huntingtin exon 1 fragment . To further assess mechanistic details of how SRCP1 inhibits polyglutamine aggregation, we utilized biochemical assays to determine that SRCP1 inhibits secondary nucleation in a manner dependent upon the regions flanking the polyglutamine tract. Finally, to determine if SRCP1 more could generally suppress protein aggregation, we confirmed that it was sufficient to inhibit aggregation of polyglutamine-expanded ataxin-3. Together these studies provide details into the structural and mechanistic basis of the inhibition of protein aggregation by SRCP1.
蛋白质聚集是多聚谷氨酰胺疾病的标志。这些疾病的一种潜在治疗方法是抑制多聚谷氨酰胺聚集。先前的工作确定了细胞黏菌作为对多聚谷氨酰胺聚集具有天然抗性的生物。进一步的工作确定富含丝氨酸的伴侣蛋白 1（SRCP1）既是细胞中抑制多聚谷氨酰胺聚集所必需的，也是人类细胞中抑制多聚谷氨酰胺聚集所必需的。因此，了解 SRCP1 如何抑制聚集可能有助于开发多聚谷氨酰胺疾病的治疗方法。在这里，我们利用蛋白质建模方法生成了 SRCP1 结构的预测。使用我们的最佳拟合模型，我们生成了突变体，这些突变体被预测会改变 SRCP1 的稳定性，并在细胞中测试了这些突变体的稳定性。利用这些数据，我们确定了与 SRCP1 的 C 末端区域形成 β-发夹结构且具有高度动态的 N 末端区域的结构的最佳模型。我们接下来生成了一系列模拟预测的 β-发夹的肽，并验证了它们抑制多聚谷氨酰胺扩展的 huntingtin 外显子 1 片段的聚集。为了进一步评估 SRCP1 抑制多聚谷氨酰胺聚集的机制细节，我们利用生化测定来确定 SRCP1 以依赖于多聚谷氨酰胺片段侧翼区域的方式抑制二级成核。最后，为了确定 SRCP1 是否更能普遍抑制蛋白质聚集，我们证实它足以抑制多聚谷氨酰胺扩展的 ataxin-3 的聚集。这些研究共同提供了关于 SRCP1 抑制蛋白质聚集的结构和机制基础的详细信息。
