Otis James Brandt, Sharpe Simon
Molecular Medicine, Hospital for Sick Children, 686 Bay St, Toronto, ONM5G 0A4, Canada.
Department of Biochemistry, University of Toronto, 1 King's College Circle, Toronto, ONM5S 1A8, Canada.
Biomacromolecules. 2022 Dec 12;23(12):5225-5238. doi: 10.1021/acs.biomac.2c01027. Epub 2022 Nov 15.
Resilin is an elastic material found in insects with exceptional durability, resilience, and extensibility, making it a promising biomaterial for tissue engineering. The monomeric precursor, pro-resilin, undergoes thermo-responsive self-assembly through liquid-liquid phase separation (LLPS). Understanding the molecular details of this assembly process is critical to developing complex biomaterials. The present study investigates the interplay between the solvent, sequence syntax, structure, and dynamics in promoting LLPS of resilin-like-polypeptides (RLPs) derived from domains 1 and 3 of pro-resilin. NMR, UV-vis, and microscopy data demonstrate that while kosmotropic salts and low pH promote LLPS, the effects of chaotropic salts with increasing pH are more complex. Subtle variations between the repeating amino acid motifs of resilin domain 1 and domain 3 lead to significantly different salt and pH dependence of LLPS, with domain 3 sequence motifs more strongly favoring phase separation under most conditions. These findings provide new insight into the molecular drivers of RLP phase separation and the complex roles of both RLP sequence and solution composition in fine-tuning assembly conditions.
弹性蛋白是一种存在于昆虫体内的弹性材料，具有卓越的耐久性、弹性和延展性，使其成为组织工程中有前景的生物材料。单体前体，即前弹性蛋白，通过液-液相分离（LLPS）进行热响应性自组装。了解这一组装过程的分子细节对于开发复杂生物材料至关重要。本研究调查了溶剂、序列句法、结构和动力学在促进源自前弹性蛋白结构域1和3的类弹性蛋白多肽（RLP）的液-液相分离中的相互作用。核磁共振（NMR）、紫外可见光谱（UV-vis）和显微镜数据表明，尽管促溶剂盐和低pH值促进液-液相分离，但随着pH值升高，离液盐的影响更为复杂。弹性蛋白结构域1和结构域3的重复氨基酸基序之间的细微差异导致液-液相分离对盐和pH值的依赖性显著不同，在大多数条件下，结构域3的序列基序更有利于相分离。这些发现为RLP相分离的分子驱动因素以及RLP序列和溶液组成在微调组装条件中的复杂作用提供了新的见解。
