In recent years, short peptide self-assembled materials, prepared under the control of the thermolysin catalyst, have been investigated extensively and shown to acquire various morphologies and functions as building blocks for a wide range of biomaterials and device applications. However, the role played by thermolysin in this enzymatically triggered peptide self-assembly is still ambiguous. Herein, we designed a series of Fmoc-dipeptide amphiphiles to explore the catalytic role of thermolysin. The results from our experiments and computational simulations showed that hydrophobicity and amino acid sequences of substrates have a significant correlation with thermolysin actions, including the binding capacity and catalytic efficiency. Specifically, thermolysin favors a specific substrate pattern with a hydrophilic amino acid in the first residue and hydrophobic amino acid in the second residue. Moreover, thermolysin catalyzed reactions are bidirectional and could move toward hydrolysis or condensation based on the design of its diverse substrates (peptides). However, the specificity of the enzyme action lies in the major site of its cleavage, which is the terminal hydrophobic or bulky amino side chains. We designed a two-step reaction, taking advantage of the bidirectional catalytic actions of thermolysin, to modify the sequence of Nα-fluorenylmethoxycarbonyl (Fmoc)-dipeptide from Fmoc-YL–COOH to Fmoc-YY–NH2 and treated with thermolysin, which resulted in the enzyme-catalyzed gel–sol–gel transition. This work has an instructive significance in the regulation of peptide sequences, secondary amino acid structures, morphology, and the mechanical property of self-assembled hydrogels with precise design and control at the molecular level via thermolysin catalysis.

中文翻译：

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嗜热菌素在Fmoc-二肽催化控制的自组装中的作用

近年来，在嗜热菌素催化剂的控制下制备的短肽自组装材料已得到广泛研究，并显示出获得各种形态和功能，可作为多种生物材料和装置应用的基础。然而，嗜热菌素在这种酶促触发的肽自组装中所起的作用仍然是模棱两可的。本文中，我们设计了一系列Fmoc-二肽两亲物以探索嗜热菌素的催化作用。我们的实验和计算模拟结果表明，底物的疏水性和氨基酸序列与热溶素作用（包括结合能力和催化效率）具有显着相关性。特别，嗜热菌蛋白酶有利于特定的底物图案，在第一残基中具有亲水氨基酸，在第二残基中具有疏水氨基酸。此外，热溶酶催化的反应是双向的，并且基于其多种底物（肽）的设计，可能会发生水解或缩合反应。但是，酶作用的特异性在于其裂解的主要位点，即末端疏水或庞大的氨基侧链。我们设计了一个两步反应，利用嗜热菌素的双向催化作用，将Nα-芴基甲氧基羰基（Fmoc）-二肽的序列从Fmoc-YL-COOH转变为Fmoc-YY-NH2，并用嗜热菌素处理，在酶催化的凝胶-溶胶-凝胶转变中。这项工作对肽序列的调控具有指导意义，