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Expanding the selectivity and activity of galactose oxidase towards bioactive saccharide

扩大半乳糖氧化酶对生物活性糖的选择性和活性

基本信息

批准号：
1960946
负责人：
金额：
--
依托单位：
University of Manchester
依托单位国家：
英国
项目类别：
Studentship
财政年份：
2017
资助国家：
英国
起止时间：
2017 至无数据
项目状态：
已结题

来源：
https://gtr.ukri.org/projects?ref=studentship-1960946
关键词：
Expanding selectivity activity galactose oxidase

项目摘要

This project aims to generate a library of sugar oxidases with novel and highly specific substrate selectivity based on parent galactose oxidase enzymes. We will use a combination of homology searches of genomic databases (Prozomix Limited),directed evolution (The University of Manchester), high-throughput enzyme expression in bacterial hosts (Prozomix) and fast high-resolution analytical screening techniques (Bio-Shape Ltd and The University of Manchester) for enzyme discovery. Background Biocatalytic oxidation reactions have the potential to substitute many chemically catalysed oxidations in the pharmaceutical and fine chemical industry due to their superior regio- and stereoselectivity and low environmental impact.[1] Furthermore, oxidative enzymatic cross-linking of proteins is an important method to functionalising foods.[2] Particularly attractive tools for such bioengineering methods are carbohydrate modifying enzymes such as oxidases, which can be highly selective as demonstrated by the widely used glucose and galactose oxidases. Despite their plethora of applications, a surprisingly narrow range of oxidases in terms of substrate specificity is currently available, which highlights the need for carbohydrates-modifying enzymes with broader substrate scopes.[3] This project aims to address this gap in available biocatalysts by developing a range of oxidases with distinct and diverse substrates specificity towards common glycans including galactosides, mannosides, lactose, fructose, sialic acids and amino sugars. We will select for biocatalysts that can be easily produced in gram quantities and larger in bacterial expression systems and thus could find wide applications in biotechnology including modification and remodelling of oligo- and polysaccharides (lactose, starch, cellulose), glycolipids and glycoproteins. Aims This project aims to now generate a designer library of active, robust and more diverse oxidases using a combination of key methods provided by project p

该项目旨在基于亲本半乳糖氧化酶产生具有新型和高度特异性底物选择性的糖氧化酶库。我们将使用基因组数据库的同源性搜索（Prozomix有限公司），定向进化（曼彻斯特大学），细菌宿主中的高通量酶表达（Prozomix）和快速高分辨率分析筛选技术（Bio-Shape Ltd和曼彻斯特大学）的组合进行酶的发现。背景生物催化氧化反应由于其上级区域和立体选择性以及低环境影响而具有在制药和精细化学工业中替代许多化学催化氧化的潜力。[1]此外，蛋白质的氧化酶交联是食品功能化的重要方法。[2]用于这种生物工程方法的特别有吸引力的工具是碳水化合物修饰酶，例如氧化酶，其可以是高度选择性的，如广泛使用的葡萄糖和半乳糖氧化酶所证明的。尽管它们有大量的应用，但目前可获得的氧化酶在底物特异性方面的范围令人惊讶地窄，这突出了对具有更宽底物范围的碳水化合物修饰酶的需求。[3]该项目旨在通过开发一系列对常见聚糖（包括半乳糖苷、甘露糖苷、乳糖、果糖、唾液酸和氨基糖）具有不同和多样化底物特异性的氧化酶来解决现有生物催化剂中的这一差距。我们将选择生物催化剂，可以很容易地产生克量和更大的细菌表达系统，从而可以在生物技术中找到广泛的应用，包括修改和改造的寡糖和多糖（乳糖，淀粉，纤维素），糖脂和糖蛋白。本项目的目标是现在使用项目p提供的关键方法的组合来生成活性的、稳健的和更多样化的氧化酶的设计者库。