尖孢镰刀菌一种新型苯甲酸羟化酶细胞色素P450的比较功能表征

Comparative functional characterization of a novel benzoate hydroxylase cytochrome P450 of Fusarium oxysporum.

作者信息

Durairaj Pradeepraj, Jung Eunok, Park Hyun Ho, Kim Byung-Gee, Yun Hyungdon

机构信息

School of Biotechnology, Yeungnam University, Gyeongsan, South Korea.

School of Chemical and Biological Engineering, Seoul National University, Seoul, South Korea.

出版信息

Enzyme Microb Technol. 2015 Mar;70:58-65. doi: 10.1016/j.enzmictec.2014.12.013. Epub 2014 Dec 31.

DOI:10.1016/j.enzmictec.2014.12.013

PMID:25659633

Abstract

FoCYP53A19, a novel cytochrome P450 capable of performing benzoate hydroxylation, was identified and characterized from the ascomycete Fusarium oxysporum f.sp. lycopersici. Comparative functional analysis of FoCYP53A19 with the heterologous and homologous cytochrome P450 reductases (CPR) such as Saccharomyces cerevisiae (ScCPR), Candida albicans (CaCPR) and F. oxysporum (FoCPR) revealed novel catalytic properties. The catalytic efficiency and substrate specificity of FoCYP53A19 were significantly influenced and altered by the source of the reductase employed. The yeast reconstitution system of FoCYP53A19 with ScCPR performed the hydroxylation of benzoic acid (BA) and demethylation of 3-methoxybenzoic acid (3-MBA); but when reconstituted with CaCPR, FoCYP53A19 performed only the essential hydroxylation of fungal benzoate catabolism. Remarkably, FoCYP53A19 with its homologous reductase FoCPR, not only demonstrated the improved conversion rates of BA and 3-MBA, but also exhibited activity toward the hydroxylation of 3-hydroxybenzoic acid. The electron transfer compatibility and the coupling efficiency between the homologous FoCYP-FoCPR system are significant and it favored enhanced monooxygenase activity with broader substrate specificity.

摘要

从尖孢镰刀菌番茄专化型（Fusarium oxysporum f.sp. lycopersici）中鉴定并表征了一种新型细胞色素P450——FoCYP53A19，它能够进行苯甲酸羟基化反应。对FoCYP53A19与异源和同源细胞色素P450还原酶（CPR），如酿酒酵母（ScCPR）、白色念珠菌（CaCPR）和尖孢镰刀菌（FoCPR）进行比较功能分析，揭示了其新的催化特性。所使用的还原酶来源对FoCYP53A19的催化效率和底物特异性有显著影响并使其发生改变。FoCYP53A19与ScCPR的酵母重组系统能够进行苯甲酸（BA）的羟基化反应和3-甲氧基苯甲酸（3-MBA）的去甲基化反应；但当与CaCPR重组时，FoCYP53A19仅进行真菌苯甲酸分解代谢的必要羟基化反应。值得注意的是，FoCYP53A19与其同源还原酶FoCPR不仅表现出更高的BA和3-MBA转化率，还对3-羟基苯甲酸的羟基化反应具有活性。同源FoCYP-FoCPR系统之间的电子转移兼容性和偶联效率显著，有利于增强具有更广泛底物特异性的单加氧酶活性。