Role of alkyl hydroperoxide reductase subunit C (AhpC) as a peroxiredoxin in Leptospira interro-gans against oxidative stress
10.3760/cma.j.issn.0254-5101.2018.03.006
- VernacularTitle:问号钩端螺旋体过氧化物还原酶AhpC在氧化应激中的功能研究
- Author:
Yihui LUO
1
;
Yifei WU
;
Munyampundu JEAN-PIERRE
Author Information
1. 浙江大学医学院病原生物学系
- Keywords:
Leptospira interrogans;
ahpC gene;
Oxidative stress;
Survival
- From:
Chinese Journal of Microbiology and Immunology
2018;38(3):193-198
- CountryChina
- Language:Chinese
-
Abstract:
Objective To analyze the enzymatic properties of alkyl hydroperoxide reductase sub-unit C (AhpC) from Leptospira interrogans (L.interrogans) and to elucidate its physiological roles in host-pathogen interactions in macrophages during Leptospira infection. Methods A prokaryotic expression system for ahpC gene of L.interrogans serogroup Icterohaemorrhagiae serovar Lai strain Lai was established to ex-press the recombinant AhpC(rAhpC). After purified by Ni-NTA affinity chromatography,the enzymatic ac-tivity of the rAhpC and its role in protecting DNA from oxidation were analyzed. The importance of each cys-teine in its molecule was evaluated through site-directed mutation. L.interrogans strains were pretreated with or without Conoidin A, a covalent inhibitor of peroxiredoxin, and then were used to infect macrophages. Changes in oxidative status in leptospires and survival rates of L.interrogans strains were analyzed by fluores-cence-activated cell sorting and colony counting method. Results The rAhpC was successfully expressed in the established prokaryotic expression system. It had peroxiredoxin activity that was able to catalyze the re-duction of hydrogen peroxide. Its ability of reducing hydrogen peroxide depended on the thioredoxin/thiore-doxin reductase system. Cys47 (a peroxidatic cysteine) and Cys167 (a resolving cysteine) were critical to maintaining the enzymatic activity of AhpC. AhpC could protect DNA from hydrogen peroxide induced-oxida-tive damage. When L.interrogans strains were pretreated with Conoidin A,the oxidative status in leptospires was elevated and the survival of L.interrogans in macrophages was significantly reduced in a dose-dependent manner. Conclusion The AhpC of L.interrogans is a thioredoxin-dependent peroxiredoxin that plays an im-portant role in protecting L.interrogans against oxidative stress in macrophages.
