Auto-immobilization for demanding enzyme catalysis using lipase-functionalized Bacillus endospores
使用脂肪酶功能化芽孢杆菌内生孢子进行自动固定以实现高要求的酶催化
基本信息
- 批准号:417423160
- 负责人:
- 金额:--
- 依托单位:
- 依托单位国家:德国
- 项目类别:Research Grants
- 财政年份:2019
- 资助国家:德国
- 起止时间:2018-12-31 至 2022-12-31
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
Enzyme-catalyzed reactions are indispensible in nowadays chemical synthesis, due to the ability of enzymes to perform difficult conversions with unrivaled selectivity at ambient reaction conditions. However, most commercially exploited enzymes require immobilization of these biocatalysts to enhance their activity, robustness and long-term stability. Current immobilization methods are challenged by the unpredictable success of immobilization, the retention of the enzymes on their respective carrier, and the influence that immobilization has on the enzymatic activity. Novel approaches that expand the current portfolio of immobilization strategies are therefore in high demand.This project aims at addressing these challenges by developing a novel and sustainable biology-based auto-immobilization approach for catalytically active enzymes. It is based on the endospore crust of the biotechnological workhorse Bacillus subtilis, which has recently been described as a novel protein display platform (so-called SporoBeads). It shall be established and optimized for conducting demanding enzyme-catalyzed reactions in technical media. Additionally, it shall be evaluated as a device for screening the catalytic performance of enzyme variants directly under technical (immobilized) conditions. For proper practical relevance, this study will focus on the currently hardly investigated, but synthetically very attractive lipase-catalyzed conversion of silyl ether bonds as a model system Reaching these goals requires combining expertise in lipase-mediated technical biocatalysis with the knowledge on applying endospore differentiation and the ability to efficiently genetically engineering the Gram-positive model organism B. subtilis, as reflected by combined efforts of the two PI’s of this joint proposal.Initially, enabling technologies for this project will be explored and established. The lipase portfolio for conversion of silyl ethers will be explored and SporoBeads will be established as an auto-immobilization platform for lipases of varying structural complexities. The knowledge gained will be combined to evaluate the potential for meeting specific requirements for synthetic application of lipase-functionalized SporoBeads. Towards establishment of SporoBeads as a screening platform for protein design, structure-guided improvements of lipases for silyl ether cleavage and formation will first be implemented, and the potential of applying SporoBeads as genetic screening tools will be evaluated. Once these prerequisites have been met, SporoBeads displaying improved lipases for performance under demanding biocatalysis conditions will be optimized.Ultimately, the experiences gained in this project shall also provide a workflow and strategy for interactively and efficiently optimizing and adjusting both the reaction conditions and the spore-display, when applying SporoBeads for demanding biocatalysis with other enzymes and for screening.
酶催化反应是当今化学合成中不可或缺的,因为酶能够在环境反应条件下以无与伦比的选择性进行困难的转化。然而,大多数商业开发的酶需要这些生物催化剂的固定化,以提高其活性,鲁棒性和长期稳定性。目前的固定化方法受到固定化的不可预测的成功、酶在其各自载体上的保留以及固定化对酶活性的影响的挑战。本项目旨在通过开发一种新的、可持续的、基于生物学的催化活性酶自动固定化方法来应对这些挑战。它基于生物技术主力枯草芽孢杆菌的内生孢子外壳,最近被描述为一种新型蛋白质展示平台(所谓的SporoBeads)。应建立并优化该系统,以便在技术介质中进行要求严格的酶催化反应。此外,应将其作为直接在技术(固定化)条件下筛选酶变体催化性能的装置进行评价。为了适当的实际相关性,本研究将重点关注目前几乎没有研究过的、但在合成上非常有吸引力的脂肪酶催化的甲硅烷基醚键转化作为模型系统。要实现这些目标,需要将脂肪酶介导的技术生物催化的专业知识与应用内生孢子分化的知识相结合,并能够有效地对革兰氏阳性模式生物B进行基因工程改造。枯草芽孢杆菌,这反映了联合提案的两个PI的共同努力。最初,将探索和建立该项目的技术。将探索用于甲硅烷基醚转化的脂肪酶组合,并将SporoBeads建立为不同结构复杂性的脂肪酶的自动固定化平台。将结合所获得的知识,以评价满足脂肪酶功能化SporoBeads合成应用特定要求的潜力。为了建立SporoBeads作为蛋白质设计的筛选平台,将首先实施用于甲硅烷基醚切割和形成的脂肪酶的结构导向改进,并将评估将SporoBeads应用于遗传筛选工具的潜力。一旦这些先决条件得到满足,SporoBeads将在苛刻的生物催化条件下显示出更好的脂肪酶性能。最终,在该项目中获得的经验还将提供一个工作流程和策略,用于交互式和有效地优化和调整反应条件和孢子展示,当应用SporoBeads与其他酶一起进行苛刻的生物催化和筛选时。
项目成果
期刊论文数量(0)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)
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Professorin Dr. Marion Ansorge-Schumacher其他文献
Professorin Dr. Marion Ansorge-Schumacher的其他文献
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{{ truncateString('Professorin Dr. Marion Ansorge-Schumacher', 18)}}的其他基金
Particle-stabilized adsorptive bubble separation of tagged enzymes – a new way to efficient downstream processing?
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407649267 - 财政年份:2019
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Model-assisted assessment and reduction of selectivity constraints in the enzyme-catalysed synthesis of chiral hydroxy ketones from prochiral diketones
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- 批准号:
187378434 - 财政年份:2011
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Biokatalysatoren für die asymmetrische Reduktion von Iminen
用于亚胺不对称还原的生物催化剂
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5452932 - 财政年份:2005
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Klonierung, Expression und Immobilisierung der Carbonyl-Reduktase aus Candida parapsilosis als vielseitiger Biokatalysator für die Synthese schwer wasserlöslicher Verbindungen
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