漆酶/多酚介体协同催化交联天然胶原及其调控机理研究

采用漆酶催化交联增强蛋白质结构稳定性以制备高性能的蛋白基功能材料是生物化工和食品领域的研究热点，但目前关于漆酶对天然胶原的催化交联机理以及多酚介体对漆酶催化交联胶原的调控机制尚不清楚。本项目基于胶原分子在溶液中的聚集行为特征，将溶液中胶原的聚集态结构与漆酶/多酚催化交联胶原的结构性能相关联，以此实现采用漆酶/多酚对胶原的协同催化交联过程的调控。拟采用氨基酸分析、氧气消耗测定等技术分别研究酪氨酸分布和多酚介体对漆酶催化交联胶原的影响；采用荧光、电泳等方法研究交联溶液中胶原的聚集态结构；采用热分析、力学测试等手段研究漆酶/多酚催化交联胶原的结构与性能；阐明溶液中胶原的聚集态结构与漆酶/多酚催化交联胶原结构性能的关系，揭示漆酶-多酚-胶原的相互作用规律及多酚对漆酶催化交联胶原的调控机制。本项目研究对采用酶交联制备高性能胶原基生物材料具有重要理论意义和应用价值，也为制革鞣制理论提供新的实验依据。

The use of laccase-catalyzed cross-linking to enhance the structural stability of proteins to prepare high-performance protein-based functional materials is a research hot-spot in the fields of biochemical engineering and food. However, the mechanism of catalytic cross-linking of laccase on natural collagen and the regulation mechanism of polyphenol mediator on laccase-catalyzed cross-linked collagen are still unclear. Based on the aggregation behavior of collagen molecules in solution, this project is to correlate the aggregate structure of collagen in solution with the structural properties of laccase/polyphenol-catalyzed cross-linked collagen, and to achieve the regulation of the catalytic crosslinking process for collagen by laccase/polyphenols. The effects of tyrosine distribution and polyphenol mediator on laccase-catalyzed cross-linked collagen were studied by means of amino acid analysis and oxygen consumption measurement etc. Fluorescence, electrophoresis and other methods were used to study the aggregation structure of collagen in the cross-linking solution. Thermal analysis, mechanical test and other means were used to study the structure and properties of laccase/polyphenol catalyzed cross-linked collagen. The relationship between the aggregation structure of collagen in solution and the structure and performance of laccase/polyphenol-catalyzed cross-linked collagen was elucidated. The interactions of laccase-polyphenol-collagen and the regulation mechanism of polyphenols on laccase-catalyzed cross-linked collagen were revealed. This project has important theoretical significance and application value for the preparation of high-performance collagen-based biomaterials by enzyme cross-linking, and provides a new experimental basis for the tanning theory of leather manufacture.