Department of Biomedical Sciences, Florida State University, Tallahassee, FL, 32306-4300, USA.
Program in Molecular Biophysics, Florida State University, Tallahassee, FL, 32306-4380, USA.
Protein Sci. 2018 Feb;27(2):431-440. doi: 10.1002/pro.3332. Epub 2017 Nov 21.
An efficient protein-folding pathway leading to target structure, and the avoidance of aggregation, is essential to protein evolution and de novo design; however, design details to achieve efficient folding and avoid aggregation are poorly understood. We report characterization of the thermally-induced aggregate of fibroblast growth factor-1 (FGF-1), a small globular protein, by solid-state NMR. NMR spectra are consistent with residual structure in the aggregate and provide evidence of a structured region that corresponds to the region of the folding nucleus. NMR data on aggregated FGF-1 also indicate the presence of unstructured regions that exhibit hydration-dependent dynamics and suggest that unstructured regions of aggregated FGF-1 lie outside the folding nucleus. Since it is known that regions outside the folding nucleus fold late in the folding pathway, we postulate that these regions unfold early in the unfolding pathway and that the partially folded state is more prone to intermolecular aggregation. This interpretation is further supported by comparison with a designed protein that shares the same FGF-1 folding nucleus sequence, but has different 1° structure outside the folding nucleus, and does not thermally aggregate. The results suggest that design of an efficient folding nucleus, and the avoidance of aggregation in the folding pathway, are potentially separable design criteria - the latter of which could principally focus upon the physicochemical properties of 1° structure outside the folding nucleus.
一种能够形成目标结构且避免聚集的有效蛋白质折叠途径，对蛋白质的进化和从头设计至关重要；然而，实现高效折叠和避免聚集的设计细节仍知之甚少。我们通过固态 NMR 对成纤维细胞生长因子-1（FGF-1）的热诱导聚集物进行了表征。NMR 谱与聚集物中的残留结构一致，并提供了结构区域对应于折叠核区域的证据。聚集 FGF-1 的 NMR 数据还表明存在无规卷曲区域，其表现出依赖于水合作用的动力学，表明聚集 FGF-1 的无规卷曲区域位于折叠核之外。由于已知折叠核之外的区域在折叠途径中折叠较晚，我们推测这些区域在解折叠途径中较早展开，并且部分折叠状态更容易发生分子间聚集。与共享相同 FGF-1 折叠核序列但具有不同折叠核外 1°结构且不热聚集的设计蛋白进行比较，进一步支持了这一解释。结果表明，设计高效的折叠核以及避免折叠途径中的聚集可能是可分离的设计标准 - 后者主要可以关注折叠核外 1°结构的物理化学性质。
