Effect of pH on the conformational structure of cytochrome c and subsequent enzymatic cross-linking catalyzed by laccase

被引：0

作者：

Li D.-X. ^{[1
]}

Qi Z.-Y. ^{[1
]}

Liu J.-Y. ^{[1
]}

Zhou J.-Q. ^{[1
]}

机构：

[1] College of Pharmaceutical Sciences, Soochow University, Suzhou

来源：

Electronic Journal of Biotechnology | 2022年 / 60卷

基金：

中国国家自然科学基金;

关键词：

Conformational structure; Cytochrome c; High-performance size exclusion chromatography; Isothermal titration calorimetry; Laccase; Laser particle size analysis; pH value; Protein cross-linking; Substrate requirement;

D O I：

10.1016/j.ejbt.2022.07.002

中图分类号：

学科分类号：

摘要：

Background: The aim of the present study was to investigate the effect of substrate conformational structure changes on the laccase-induced protein cross-linking. The effects of laccase amount, pH, and ferulic acid (FA) on the enzymatic cross-linking of substrate, Cyt C, were determined by sodium dodecyl sulfate–polyacrylamide gel electrophoresis. High-performance size exclusion chromatography, laser particle size analysis and isothermal titration calorimetry (ITC) were also applied to investigate the cross-linking product and enthalpy changes. Structural changes of Cyt C at different pH values were analyzed by ultraviolet–visible (UV–vis), fluorescence, and circular dichroism (CD) measurements. Results: Complete cross-linking, partial cross-linking, minute cross-linking, and no cross-linking occurred at pH 2.0, 4.0, 6.0, and 8.0, respectively. ITC analysis demonstrated that the enzymatic cross-linking of Cyt C was an endothermic process. The UV–vis, fluorescence, and CD measurements exhibited that the tertiary structure of Cyt C was disrupted, and part of the α-helical polypeptide region unfolded at pH 2.0. The structural flexibilities decreased, and the tertiary structure of Cyt C became increasingly compact with the increase in pH values from 4.0 to 8.0. The gradual changes in the structure of Cyt C at different pH values were in accordance with the cross-linking results of Cyt C catalyzed by laccase. Conclusions: The results demonstrated that minute structure changes of substrate had a remarkable effect on the laccase-induced cross-linking. The findings promote the understanding of the substrate requirement of laccase in protein cross-linking and are instructive for the modulation of laccase-induced protein cross-linking. How to cite: Li D-X, Qi Z-Y, Liu J-Y, et al. Effect of pH on the conformational structure of cytochrome c and subsequent enzymatic cross-linking catalyzed by laccase. Electron J Biotechnol 2022;60. https://doi.org/10.1016/j.ejbt.2022.07.002. © 2022

引用

页码：1 / 10

页数：9

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