Now, Ana S. Pina and colleagues report that LLPS can significantly increase the catalytic activity of peptides. Specifically, the researchers created single peptide-based coacervates, whereby the spatial confinement led to a more folded structure of the peptides (pictured). The peptide P7 (KVYFSIPWRVPM-NH₂), which was used as a model system, has a high affinity for phosphorylated assemblies, can hydrolyse phosphate ester molecules, and contains arginine, lysine, serine and proline residues that are recognized to be beneficial for phase separation. Screening of peptide and salt concentrations and different temperatures led to suitable conditions to induce the formation of P7 coacervates and further optimization of the experimental design reduced unwanted reactions such as aggregation and precipitation of the peptides and division of the coacervates. Spectroscopic methods were then used to investigate the effect of LLPS on peptide conformation. It was shown that the compartmentalization via LLPS led to a stabilization of the secondary structure of P7. Specifically, a fully folded β-hairpin structure was detected compared to a flexible hairpin-like peptide in solution. Moreover, charge and hydrophobicity of the peptide influenced the partitioning of guest molecules by the coacervates. It was further shown that phosphorylation had an even more pronounced effect, leading to preferential uptake compared to non-phosphorylated versions of protein molecules. Subsequently, the coacervates were tested for catalytic phosphate ester hydrolysis of p-nitrophenyl phosphate (pNPP) as the substrate and catalytic parameters of k_cat = (4.9 ± 0.6) × 10^–3 s⁻¹ and K_M = (8.2 ± 3.2) × 10^–4 M and a catalytic efficiency of k_cat/K_M = 5.9 ± 0.2 were determined. Impressively, this constitutes a 15,000-fold improvement in catalytic efficiency over the reaction mediated by the free peptides in solution determined in a prior work.

The demonstration that simple, small peptides can enable self-coacervation, selectively recruit substrates and accelerate catalysis will likely be of interest to origin of life researchers and might also serve as an inspiration for the future design of systems for drug delivery and sensing.

现在，Ana S. Pina 及其同事报告说，LLPS 可以显着提高肽的催化活性。具体来说，研究人员创建了基于单个肽的凝聚物，从而空间限制导致肽的结构更加折叠（如图）。肽 P7 （KVYFSIPWRVPM-NH₂） 用作模型系统，对磷酸化组装体具有高亲和力，可以水解磷酸酯分子，并包含被认为有利于相分离的精氨酸、赖氨酸、丝氨酸和脯氨酸残基。肽和盐浓度的筛选以及不同的温度导致了诱导 P7 凝聚物形成的合适条件，并且进一步优化了实验设计，减少了不需要的反应，例如肽的聚集和沉淀以及凝聚物的分裂。然后使用光谱方法研究 LLPS 对肽构象的影响。结果表明，通过 LLPS 的区室化导致 P7 二级结构的稳定。具体来说，与溶液中的柔性发夹状肽相比，检测到完全折叠的 β 发夹结构。此外，肽的电荷和疏水性影响了凝聚物对客体分子的分配。进一步表明，磷酸化具有更明显的效果，与非磷酸化蛋白质分子相比，磷酸化导致优先摄取。随后，测试凝聚物中以对硝基苯磷酸盐 （pNPP） 为底物的催化磷酸酯水解以及 k_cat = （4.9 ± 0.6） × ^{10-3 s-1} 和 K_M = （8.2 ± 3） 的催化参数。2） × ^10-4 M 和 k_cat/K_M = 5.9 ± 0.2 的催化效率测定。令人印象深刻的是，与先前工作中测定的溶液中游离肽介导的反应相比，这构成了催化效率的 15,000 倍的提高。

简单、小的肽可以实现自凝聚、选择性募集底物和加速催化的证明可能会引起 Origin of Life 研究人员的兴趣，也可能为未来药物递送和传感系统的设计提供灵感。