Effect of Organic Solvents on the Activity, Stability and Secondary Structure of asclepain cI, Using FTIR and Molecular Dynamics Simulations

IF 1.9 4区生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY

The Protein Journal Pub Date : 2024-03-07 DOI:10.1007/s10930-024-10182-4

Anabella L. Origone, Esteban G. Vega Hissi, Constanza S. Liggieri, Gerardo E. Camí, Andrés Illanes, Sonia E. Barberis

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Abstract

The present study aims at understanding the effect of organic solvents on the specific proteolytic activity and operational stability of asclepain cI in aqueous-organic media, using correlations between geometrical and structural parameters of asclepain cI. These correlations were determined by molecular dynamics (MD) simulations and the secondary structure of the enzyme validated by Fourier-transform Infrared (FTIR) spectroscopy. Asclepain cI exhibited significantly higher catalytic potential in 29 of the 42 aqueous-organic media tested, composed by 0.1 mM TRIS hydrochloride buffer pH 8 (TCB) and an organic solvent, than in buffer alone. Asclepain cI in water-organic miscible systems showed high FTIR spectral similarity with that obtained in TCB, while in immiscible systems the enzyme acquired different secondary structures than in buffer. Among the conditions studied, asclepain cI showed the highest catalytic potential in 50% v/v ethyl acetate in TCB. According to MD simulations, that medium elicited solvation and flexibility changes around the active center of asclepain cI and conducted to a new secondary structure with the active center preserved. These results provide valuable insights into the elucidation of the molecular mechanism of asclepain cI tolerance to organic solvents and pave the way for its future application for the synthesis of peptides in aqueous-organic media.

Graphical Abstract

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利用傅立叶变换红外光谱和分子动力学模拟，研究有机溶剂对 asclepain cI 的活性、稳定性和二级结构的影响。

本研究旨在利用 Asclepain cI 的几何和结构参数之间的相关性，了解有机溶剂对 asclepain cI 在水有机介质中的特定蛋白水解活性和操作稳定性的影响。这些相关性由分子动力学（MD）模拟确定，酶的二级结构由傅立叶变换红外光谱（FTIR）验证。在由 0.1 mM TRIS hydrochloride buffer pH 8 (TCB) 和有机溶剂组成的 42 种测试水有机介质中，Asclepain cI 在 29 种介质中表现出的催化潜能明显高于仅在缓冲液中的催化潜能。水-有机混溶体系中的 Asclepain cI 与在 TCB 中获得的傅立叶变换红外光谱具有很高的相似性，而在不混溶体系中，酶获得的二级结构与缓冲液中的不同。在所研究的条件中，asclepain cI 在 TCB 中的 50% v/v 乙酸乙酯中显示出最高的催化潜能。根据 MD 模拟，该介质引起了 asclepain cI 活性中心周围的溶解和柔性变化，并在保留活性中心的情况下形成了新的二级结构。这些结果为阐明 Asclepain cI 耐有机溶剂的分子机理提供了有价值的见解，并为其今后在水有机介质中合成多肽的应用铺平了道路。

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来源期刊

The Protein Journal 生物-生化与分子生物学

CiteScore

5.20

自引率

0.00%

发文量

审稿时长

12 months

期刊介绍： The Protein Journal (formerly the Journal of Protein Chemistry) publishes original research work on all aspects of proteins and peptides. These include studies concerned with covalent or three-dimensional structure determination (X-ray, NMR, cryoEM, EPR/ESR, optical methods, etc.), computational aspects of protein structure and function, protein folding and misfolding, assembly, genetics, evolution, proteomics, molecular biology, protein engineering, protein nanotechnology, protein purification and analysis and peptide synthesis, as well as the elucidation and interpretation of the molecular bases of biological activities of proteins and peptides. We accept original research papers, reviews, mini-reviews, hypotheses, opinion papers, and letters to the editor.