Kinetic data from different acetoacetyl-CoA reductases

These folders contain experimental records, calculation datasheets and scripts to simulate or find best-fitted kinetic parameters from certain enzymatically catalyzed reactions. These reactions were catalyzed by four engineered enzymes derived from the acetoacetyl-CoA reductase encoded by the phaB1 gene from Cupriavidus necator. The DNA and amino acid sequences of these enzymes are provided. It is also provided the DNA sequence map of the vector where the amino acid encoding sequences for these engineered enzymes were inserted. Inside the folder named "kinetic data" it is also possible to find a TXT file with a detailed description of the content. The experimental conditions of the enzymatic assays is carefully explained. Briefly, this folder contains (1) the raw experimental data recorded with the software Gen 5 (Biotek). (2) It is also possible to find Microsoft Excel datasheets with the records of raw absorbance in time together with initial substrates, initial enzyme and product concentrations in time (reaction progress curves). Most of these progress curves were analyzed with the software DYNAFIT (Biokin) to obtain the kinetic parameters. The progress curves highlighted in red in the Microsoft Excel files were not considered for the statistical analysis due to experimental errors. The progress curves highlighted in yellow in the Microsoft Excel files were considered after the elimination of some outlier points. (3) Scripts enabling two kind of statistical analyses with DYNAFIT were included: scripts to make model discrimination analyses of the different reaction progress curves obtained in every single experiment, and scripts to calculate the confidence intervals for the kinetic parameters using a Monte Carlo approach. We included the scripting manual of DYNAFIT to understand the synthaxis of these scripts .(4) It is included two MATLAB scripts to make a comparison between the kinetic parameters obtained using the Michaelis-Menten model and the true kinetic parameters of a BiBi reversible reaction. (5) Finally, we included DYNAFIT, MATLAB and Microsoft Excel files enabling the calculation of the relative use of NADH over NADPH by one of the engineered enzymes, and also to calculate the metabolic flux capacity of this enzyme.本数据集文件夹涵盖若干酶促反应的实验记录、计算数据表，以及用于模拟反应过程或拟合最优动力学参数的脚本程序。本次研究涉及的所有酶促反应，均由4种经工程化改造的酶催化——这些酶均源自贪铜菌（Cupriavidus necator）phaB1基因编码的乙酰乙酰辅酶A还原酶（acetoacetyl-CoA reductase）。本数据集同步提供了上述4种工程酶的DNA序列与氨基酸序列，以及插入了这些工程酶编码序列的载体DNA序列图谱。 在名为"kinetic data（动力学数据）"的文件夹内，还存有一份详细说明数据集内容的TXT文件，并对酶活实验的具体实验条件进行了详尽阐释。简言之，该动力学数据文件夹包含以下内容： 1. 使用Gen 5（Biotek）软件记录的原始实验数据； 2. 若干Microsoft Excel格式数据表，记录了随时间变化的原始吸光度值，以及初始底物、初始酶与产物的实时浓度（即反应进程曲线）。其中多数反应进程曲线通过DYNAFIT（Biokin）软件分析，以拟合得到动力学参数。Excel文件中标注为红色的反应进程曲线因存在实验误差，未纳入统计分析；标注为黄色的曲线则在剔除异常值后纳入统计范畴； 3. 两类基于DYNAFIT的统计分析脚本：其一用于对单次实验获取的不同反应进程曲线开展模型判别分析，其二用于通过蒙特卡洛（Monte Carlo）方法计算动力学参数的置信区间。此外，本数据集还附带了DYNAFIT脚本编写手册，以帮助使用者理解上述脚本的语法规范； 4. 两份MATLAB脚本，用于对比米氏方程（Michaelis-Menten）模型拟合得到的动力学参数，与双底物双产物（BiBi）可逆反应的真实动力学参数； 5. 最后，本数据集还包含DYNAFIT、MATLAB与Microsoft Excel文件，用于计算其中一种工程酶对NADH与NADPH的相对利用效率，以及该酶的代谢通量能力。