Reaction kinetics identification for the analysis of reaction mechanisms of ThDP-dependent enzymes

反应动力学鉴定用于分析 ThDP 依赖性酶的反应机制

• 批准号: 172084740
• 负责人: Professorin Dr.-Ing. Antje Spieß
• 金额: --
• 依托单位: Institut für Bioverfahrenstechnik
• 依托单位国家: 德国
• 项目类别: Research Units
• 财政年份: 2010
• 资助国家: 德国
• 起止时间: 2009-12-31 至 2015-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/172084740?language=en
• 关键词: Reaction; kinetics; identification; analysis; reaction

The elucidation of reaction mechanisms and kinetics for thiamine diphosphat (ThDP)- dependent enzymes in synthetic applications is by far not closed. Although mechanistic postulates exist for a range of enzymes and their respective reactions, there is yet no conclusive information available to deduce and predict the catalytic abilities of different enzymes and correlate them with structural information. The analysis of steady-statereaction kinetics allows suggesting suitable reaction kinetic expressions and the related kinetic parameters and thus the determination of the rate-limiting step. An important feature to achieve kinetic identification is the planning of optimal experiments for model discrimination and parameter estimation. Reaction kinetic identification becomes increasingly complex with increasing number of reactants. In case of 2-hydroxy ketones, two different substrates result in the formation of up to four distinct products, each in two enantiomeric forms. For the identification of reactions of that complexity, suitable experimental and identification strategies will be implemented and verified. The suggested reaction mechanisms are verified using NMR methods and molecular modeling. Objective of this project is to support the differentiation and classification of ThDPdependent enzymes in terms of their ability to transfer the acyl functionalities to different acceptors and thus contribute to the development of an integrated understanding of ThDP reaction range and its application in biocatalytic synthesis.

合成应用中二磷酸硫胺素 (ThDP) 依赖性酶的反应机制和动力学的阐明尚未结束。尽管一系列酶及其各自的反应存在机械假设，但尚无确凿的信息可用于推断和预测不同酶的催化能力并将其与结构信息相关联。稳态反应动力学分析可以提出合适的反应动力学表达式和相关动力学参数，从而确定限速步骤。实现动力学识别的一个重要特征是规划模型判别和参数估计的最佳实验。随着反应物数量的增加，反应动力学识别变得越来越复杂。对于 2-羟基酮，两种不同的底物会导致形成多达四种不同的产物，每种产物有两种对映体形式。为了识别如此复杂的反应，将实施和验证合适的实验和识别策略。使用核磁共振方法和分子模型验证了所建议的反应机制。该项目的目的是支持 ThDP 依赖性酶根据其将酰基官能团转移到不同受体的能力进行区分和分类，从而有助于全面了解 ThDP 反应范围及其在生物催化合成中的应用。

项目成果

Systematic determination of intrinsic reaction parameters in enzyme immobilizates

酶固定化物中固有反应参数的系统测定

• DOI: 10.1016/j.ces.2009.12.026
• 发表时间: 2010
• 期刊: Chemical Engineering Science
• 影响因子: 4.7
• 作者: Zavrel M;Michalik C;Schwendt T;Schmidt T;Ansorge-Schumacher M;Janzen C;Marquardt W;Büchs J;Spiess AC
• 通讯作者: Spiess AC

Simultaneous identification of reaction and inactivation kinetics of an enzyme‐catalyzed carboligation

同时鉴定酶催化碳化反应的反应和失活动力学

• DOI: 10.1002/btpr.2656
• 发表时间: 2018
• 期刊: Biotechnology Progress
• 影响因子: 2.9
• 作者: Leipnitz M. Schöpping M;Spiess AC
• 通讯作者: Spiess AC

How graphical analysis helps interpreting optimal experimental designs for nonlinear enzyme kinetic models

• DOI: 10.1002/aic.15814
• 发表时间: 2017-11
• 期刊: Aiche Journal
• 影响因子: 3.7
• 作者: Rüdiger Ohs;Jan Wendlandt;A. Spiess