Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet, S-17165 Solna, Sweden.
Department of Biosciences and Nutrition, Karolinska Institutet, S-14157 Huddinge, Sweden.
Nano Lett. 2023 Jun 28;23(12):5836-5841. doi: 10.1021/acs.nanolett.3c00773. Epub 2023 Apr 21.
Many protein condensates can convert to fibrillar aggregates, but the underlying mechanisms are unclear. Liquid-liquid phase separation (LLPS) of spider silk proteins, spidroins, suggests a regulatory switch between both states. Here, we combine microscopy and native mass spectrometry to investigate the influence of protein sequence, ions, and regulatory domains on spidroin LLPS. We find that salting out-effects drive LLPS via low-affinity stickers in the repeat domains. Interestingly, conditions that enable LLPS simultaneously cause dissociation of the dimeric C-terminal domain (CTD), priming it for aggregation. Since the CTD enhances LLPS of spidroins but is also required for their conversion into amyloid-like fibers, we expand the stickers and spacers-model of phase separation with the concept of folded domains as conditional stickers that represent regulatory units.
许多蛋白质凝聚物可以转化为纤维状聚集体，但潜在的机制尚不清楚。蜘蛛丝蛋白丝氨酸的液-液相分离（LLPS）表明两种状态之间存在调节开关。在这里，我们结合显微镜和天然质谱法来研究蛋白质序列、离子和调节结构域对丝氨酸蛋白 LLPS 的影响。我们发现盐析效应通过重复结构域中的低亲和力粘性物质驱动 LLPS。有趣的是，能够进行 LLPS 的条件会同时导致二聚体 C 端结构域（CTD）的解离，使其易于聚集。由于 CTD 增强了丝氨酸蛋白的 LLPS，但也需要其转化为类淀粉样纤维，因此我们用折叠结构域作为条件粘性物质的概念扩展了相分离的粘性物质和间隔子模型，该概念代表了调节单元。
