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来自Pseudomonas aeruginosa的两个不同的异酸盐脱水酶:酶学和辅酶进化影响

Xuefei Chen¹, Wei Wei¹, Wei Xiong¹

¹Anhui Provincial Key Laboratory of Molecular Enzymology and Mechanism of Major Diseases and Key Laboratory of Biomedicine in Gene Diseases and Health of Anhui Higher Education Institutes, Anhui Normal University, Wuhu 241000, China.

International journal of molecular sciences

|October 14, 2023

概括

Pseudomonas aeruginosa 具有两种依赖NADP+的异酸盐脱酶 (NADP-IDHs),具有不同的特性. PaIDH1的酸化降低了它的活性,突变可以将辅酶特异性从NADP+转变为NADP+.

科学领域:

生物化学生化学
分子生物学分子生物学
微生物学微生物学

背景情况:

Pseudomonas aeruginosa 携带两个依赖NADP+的同位素脱酶 (NADP-IDHs),PaIDH1和PaIDH2,它们是从依赖NAD的对应物进化而来的.
了解这些酶对于理解细菌代谢和病原性至关重要.

研究的目的:

为了克隆,过度表达,并净化PaIDH1和PaIDH2从P. aeruginosa.
描述它们的生物化学特性,包括基质亲和力,最佳条件和调节机制.
研究突变对辅酶特异性和酶活性的影响.

主要方法:

在大肠杆菌中基因克隆和蛋白质过度表达.
酶净化和表征 (Km值,温度和pH容忍度).
位点定向的突变发生和酶活性和辅酶特异性的分析.

主要成果:

与PaIDH2.2相比,PaIDH1对NADP+和异酸盐具有更高的亲和力,更高的热稳定性和更广泛的pH耐受性.
通过PaIDH K/P对PaIDH1的酸化显著降低了其活性.
突变导致两种PaIDH的协酶特异性从NADP+转变为NAD+,而不会改变它们的寡合状态.

结论:

PaIDH1和PaIDH2具有不同的生化特性和调节机制.
酸化作为PaIDH1活动的调节机制.
工程突变可以改变这些酶的辅酶特异性,为它们的进化轨迹提供了洞察力.

关键词:

伪omonas aeruginosa 这种类型的病毒.同酶的特异性进化演化演化演化演化演化演化演化异酸盐脱基酶是什么酸化的方法是:光化.

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