optiMEET – Optimisation of mediated extracellular electron transfer for Cupriavidus necator

optiMEET â 优化 Cupriavidus necator 介导的细胞外电子转移

• 批准号: 536355345
• 负责人: Professor Dr.-Ing. Dirk Holtmann
• 金额: --
• 依托单位: Arbeitsgruppe Physikalische Chemie/ Biophysikalische Chemie
• 依托单位国家: 德国
• 项目类别: Priority Programmes
• 资助国家: 德国
• 项目状态: 未结题
• 来源: https://gepris.dfg.de/gepris/projekt/536355345?language=en
• 关键词: optiMEET; Optimisation; mediated; extracellular; electron

Cupriavidus necator represents an ideal candidate for electro-microbial production of value-added compounds, as it combines all the advantages necessary for efficient electro-microbial processes: i) As a facultative chemolithoautotrophic bacterium, it grows rapidly to high cell densities on a variety of substrates, including CO2/H2. ii) The availability of of its complete genome sequence as well as genetic tools allow for easy genetic engineering. iii) C. necator performs both aerobic and anaerobic respiration. In the current project the applicants completed the following tasks: • An eight cuvette-based screening tool for electrochemical investigations on microbes has been developed and validated. • Four parallel-controlled electrobioreactors have been established to perform scalable electromicrobial processes. • Anodic respiration, based on a mediated electron transfer, has been established for C. necator, and proteins likely involved in this electron transfer have been identified. • Vibrio natriegens has been identified as another organism capable of utilizing artificial redox mediators for anodic respiration. • The extent of toxicity of artificial redox mediators has been determined in living cell cultures. • The homologous and heterologous overproduction of CymA/NapC-like metalloproteins in the cytoplasmic membrane was studied to enhance electron transfer from the quinone pool to (artificial) periplasmic electron acceptors. • The SpyTag/SpyCatcher system has been engineered for systhetic covalent coupling of periplasmic redox enzymes both in vitro and in vivo. Thus, main objectives of the project have already been achieved, leading to four peer-reviewed publications so far. Another manuscript is under review and two more are in preparation. Based on preliminary work, in the follow-up project the applicants are planning the following tasks: • Generate a solid understanding of mediated electron transfer based on gene knockout and overexpression strains of C. necator. • Computational fluid dynamics-assisted scale-up of electro-bioreactors. • Model-based optimization of the interaction between electrodes, artificial redox mediators and living microorganisms. • Transfer of knowledge of mediated electron transfer and artificial coupling of redox proteins in the periplasm to other organisms (S. oneidensis and V. natriegens). The main objectives of the project "Optimization of mediated electron transfer for C. necator" are to gain a deep understanding of anode respiration, as well as electrosynthetic properties of C. necator, to transfer the knowledge to larger reactors and the application of general principles of artificial (extracellular) electron transfer to other organisms.

钩虫贪铜菌代表了电微生物生产增值化合物的理想候选者，因为它结合了有效电微生物过程所需的所有优点：i）作为兼性化能无机自养细菌，它在各种底物（包括CO2/H2）上快速生长至高细胞密度。ii）其完整基因组序列的可用性以及遗传工具允许容易的遗传工程。iii）C. necator进行有氧和无氧呼吸。在目前的项目中，申请人完成了以下任务：·已经开发并验证了用于微生物电化学研究的基于八个比色皿的筛选工具。·已经建立了四个平行控制的电生物反应器以执行可扩展的电微生物过程。·C.已经建立了基于介导电子转移的阳极呼吸。necator和可能参与这种电子转移的蛋白质已经被鉴定。·产钠弧菌已被鉴定为另一种能够利用人工氧化还原介体进行阳极呼吸的生物体。·人工氧化还原介质的毒性程度已在活细胞培养物中确定。·研究了细胞质膜中CymA/NapC样金属蛋白的同源和异源过量产生，以增强从醌库到（人工）周质电子受体的电子转移。·SpyTag/SpyCatcher系统已被工程化用于在体外和体内合成周质氧化还原酶的共价偶联。因此，该项目的主要目标已经实现，迄今已出版了四份经同行审查的出版物。另一份手稿正在审查中，另外两份正在编写中。在前期工作的基础上，申请人计划在后续项目中开展以下工作：·对基于C.钩虫·电生物反应器的计算流体动力学辅助放大。·基于模型优化电极、人工氧化还原介质和活微生物之间的相互作用。·将介导的电子转移和周质中氧化还原蛋白的人工偶联的知识转移到其他生物体（S。oneidensis和V. natriegens）。本课题的主要目的是研究C. necator”的研究，以深入了解阳极呼吸以及C. necator，将知识转移到更大的反应器，并将人工（细胞外）电子转移的一般原理应用于其他生物。