工程化碳水化合物结合模块以提高几丁质酶在不溶性几丁质底物上的催化效率。

Engineering a carbohydrate binding module to enhance chitinase catalytic efficiency on insoluble chitinous substrate.

机构信息

College of Food Science and Engineering, Ocean University of China, Qingdao 266003, China.

College of Food Science and Engineering, Ocean University of China, Qingdao 266003, China; Laboratory for Marine Drugs and Bioproducts of Qingdao National Laboratory for Marine Science and Technology, Qingdao 266237, China.

出版信息

Food Chem. 2021 Sep 1;355:129462. doi: 10.1016/j.foodchem.2021.129462. Epub 2021 Mar 16.

DOI:10.1016/j.foodchem.2021.129462

PMID:33848938

Abstract

Development of a high-performance chitinase for efficient biotransformation of insoluble chitinous substrate would be highly valuable in industry. In this study, the chitin-binding domains (ChBDs) of chitinase SaChiA4 were successfully modified to improve the enzymatic activity. The engineered substitution variant R-SaChiA4, which had the exogenous ChBD of chitinase ChiA1 from Bacillus circulans WL-12 (ChBD) substituted for its original ChBD, increased its activity by nearly 54% (28.0 U/mg) towards chitin powder, and by 49% towards colloidal chitin, compared with the wild-type. The substrate-binding assay demonstrated that the ChBD could enhance the capacity of enzymatic hydrolysis by promoting substrate affinity, and molecular dynamics simulations indicated that this could be due to hydrophobic interactions in different substrate binding modes. This work advances the understanding of the role of the ChBD, and provides a step towards the achievement of industrial-scale hydrolysis and utilization of insoluble chitin.

摘要

开发高效的几丁质酶用于高效转化不溶性几丁质底物在工业上具有重要价值。在这项研究中，成功地修饰了几丁质酶 SaChiA4 的几丁质结合结构域（ChBD），以提高酶活性。工程替代变体 R-SaChiA4 将外源性几丁质酶 ChiA1 的 ChBD 取代其原始 ChBD，使其对几丁质粉末的活性增加了近 54%（28.0 U/mg），对胶体几丁质的活性增加了 49%，与野生型相比。底物结合试验表明，ChBD 可以通过促进底物亲和力来提高酶水解能力，分子动力学模拟表明，这可能是由于不同底物结合模式中的疏水相互作用。这项工作推进了对 ChBD 作用的理解，并为实现不溶性几丁质的工业规模水解和利用迈出了一步。