Department of Chemical Engineering, Columbia University, New York, New York 10027, United States.
ACS Synth Biol. 2024 Feb 16;13(2):598-612. doi: 10.1021/acssynbio.3c00564. Epub 2024 Feb 3.
Subcellular phase-separated compartments, known as biomolecular condensates, play an important role in the spatiotemporal organization of cells. To understand the sequence-determinants of phase separation in bacteria, we engineered protein-based condensates in using electrostatic interactions as the main driving force. Minimal cationic disordered peptides were used to supercharge negative, neutral, and positive globular model proteins, enabling their phase separation with anionic biomacromolecules in the cell. The phase behavior was governed by the interaction strength between the cationic proteins and anionic biopolymers, in addition to the protein concentration. The interaction strength primarily depended on the overall net charge of the protein, but the distribution of charge between the globular and disordered domains also had an impact. Notably, the protein charge distribution between domains could tune mesoscale attributes such as the size, number, and subcellular localization of condensates within cells. The length and charge density of the disordered peptides had significant effects on protein expression levels, ultimately influencing the formation of condensates. Taken together, charge-patterned disordered peptides provide a platform for understanding the molecular grammar underlying phase separation in bacteria.
亚细胞相分离隔室，也被称为生物分子凝聚物，在细胞的时空组织中起着重要作用。为了理解细菌中相分离的序列决定因素，我们使用静电相互作用作为主要驱动力，在 中设计了基于蛋白质的凝聚物。使用最小阳离子无序肽来超荷负电、中性和正电球状模型蛋白，使它们与细胞中的阴离子生物大分子发生相分离。相行为受阳离子蛋白和阴离子生物聚合物之间的相互作用强度以及蛋白质浓度的控制。相互作用强度主要取决于蛋白质的总净电荷，但球状和无序结构域之间的电荷分布也有影响。值得注意的是，蛋白质在结构域之间的电荷分布可以调节介观属性，例如细胞内凝聚物的大小、数量和亚细胞定位。无序肽的长度和电荷密度对蛋白质表达水平有显著影响，最终影响凝聚物的形成。总之，带电荷模式的无序肽为理解细菌中相分离的分子语法提供了一个平台。
