Enhancing catalytic efficiency of GAO-5F from Fusarium odoratissimum and its application in development of a polyaldehyde crosslinked gelatin-based edible packaging film

文献类型： 外文期刊

第一作者： Sun, Huihui

作者： Sun, Huihui;Xia, Guangli;Cao, Na;Zhao, Ling;Cao, Rong;Cao, Rong

作者机构：

关键词： Galactose oxidase; Semi-rational design; Edible packaging film

期刊名称：INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES （ 影响因子：8.5； 五年影响因子：8.7 ）

ISSN： 0141-8130

年卷期： 2024 年 283 卷

页码：

收录情况： SCI

摘要： Galactose oxidase has long captured the interest of the biocatalysis and biotechnology communities due to its unique catalytic characteristics and versatility with various substrates. In our previous studies, we demonstrated that galactose oxidase GAO-5F from Fusarium odoratissimum can oxidize agarose to produce a polyaldehyde polymer, which can be further crosslinked with gelatin to produce food packaging films. Despite its commendable catalytic performance, GAO-5F falls short of meeting the requirements for industrial applications. In this study, we employed a combination of multiple sequence comparisons and site-directed mutagenesis to pinpoint key amino acid sites crucial for enhancing the enzyme's catalytic activity, resulting in the creation of the double mutant GAO-5F/AR (D403A/Q484R), showing a six-fold increase in catalytic activity. The catalytic mechanism of mutant was further elucidated through homology modeling and molecular docking. Results highlighted the significance of increased hydrogen bonding interactions between the enzyme and substrate in enhancing catalytic activity. Then, agarose was transformed into a polyaldehyde polymer by oxidation catalyzed by GAO-5F mutant. The resulting polyaldehyde polymer was crosslinked with gelatin to prepare an edible packaging film; the properties and structure of the film were characterized. In this study, we successfully obtained mutants with increased catalytic activity through a semi-rational-driven site-directed mutagenesis strategy. This approach, which combines rational design with targeted mutagenesis, holds promise for furthering our understanding of enzyme function and may find widespread use in comparative functional genomics studies of other natural enzymes. This study provides valuable insights for the improvement of galactose oxidase, and new ideas for the preparation of edible packaging films for use in the food industry.

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