Characterization of thioredoxin-thioredoxin reductase system in Filifactor alocis.

IF 2.8 3区医学 Q1 DENTISTRY, ORAL SURGERY & MEDICINE

Molecular Oral Microbiology Pub Date : 2024-10-20 DOI:10.1111/omi.12486

Arunima Mishra, Yuetan Dou, Hansel M Fletcher

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引用次数: 0

Abstract

Introduction: Filifactor alocis is a newly appreciated member of the periodontal community with a strong periodontal disease correlation. Little is known about the survival mechanisms by which F. alocis copes with oxidative stress and establishes the infection within the local inflammatory microenvironment of the periodontal pocket. The aim of this study is to investigate if F. alocis putative peroxiredoxin/AhpC protein FA768 may constitute an alkyl hydroperoxide reductase system utilizing putative thioredoxin reductase protein FA608, and putative thioredoxin/glutaredoxin homolog FA1411/FA455.

Methods: FA768, FA608, FA1411 and FA455 proteins from F. alocis were expressed and purified from Escherichia coli. Insulin and 5,5-dithio-bis-2-nitrobenzoic acid (DTNB) reduction assays were performed to determine if purified FA1411 and FA455 proteins could be a substrate for FA608. The peroxidase activity of FA768 was examined by measuring its ability to reduce hydrogen peroxide (H₂O₂) with FA608 and FA1411/FA455 provided as the reducing systems. Further, the hydroperoxide substrate specificity of FA768 was analyzed by monitoring the NADPH oxidation in the presence of different peroxides, including H₂O₂, cumyl hydroperoxide (CHP), and tert-butyl hydroperoxide (t-BHP).

Results: In this study, we have demonstrated the existence of a functioning thioredoxin-dependent alkyl hydroperoxide system in F. alocis. This system is comprised of a thioredoxin reductase (FA608), a thioredoxin/glutaredoxin homolog (FA1411/FA455), and a typical 2-cysteine peroxiredoxin/AhpC (FA768). FA608, together with FA1411/FA455, can function as a thioredoxin reductase system to reduce insulin, DTNB, and FA768. FA455 is a glutaredoxin-like protein with thioredoxin functions in F. alocis. Both the FA768/FA608/FA1411 and FA768/FA608/FA455 reductase systems were NADPH-dependent and exhibited specificity for broad hydroperoxide substrates H₂O₂, CHP, and t-BHP.

Conclusions: This is the first study of a thioredoxin dependent alkyl hydroperoxide system from a periodontal pathogen. This system is proposed to protect F. alocis against oxidative stress due to the likely absence of a catalase or an additional peroxiredoxin homolog.

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Filifactor alocis 中硫氧化还原酶系统的特征。

导言：Filifactor alocis 是牙周病群体中新近受到重视的成员，与牙周病密切相关。人们对 F. alocis 应对氧化应激并在牙周袋局部炎症微环境中建立感染的生存机制知之甚少。本研究旨在探讨 F. alocis 推测的过氧化氢还原酶/AhpC 蛋白 FA768 是否可能利用推测的硫氧还原酶蛋白 FA608 和推测的硫氧还原酶/谷氨酰还原酶同源物 FA1411/FA455，构成一个烷基过氧化氢还原酶系统：方法：从大肠杆菌中表达并纯化了 F. alocis 的 FA768、FA608、FA1411 和 FA455 蛋白。进行了胰岛素和 5,5-二硫双-2-硝基苯甲酸（DTNB）还原试验，以确定纯化的 FA1411 和 FA455 蛋白是否可作为 FA608 的底物。通过测定 FA768 以 FA608 和 FA1411/FA455 为还原体系还原过氧化氢（H2O2）的能力，检验了 FA768 的过氧化物酶活性。此外，通过监测在不同过氧化物（包括 H2O2、积雪草过氧化氢（CHP）和叔丁基过氧化氢（t-BHP））存在下的 NADPH 氧化作用，分析了 FA768 的过氧化氢底物特异性：结果：在这项研究中，我们证明了 F. alocis 中存在一个依赖硫代氧化还蛋白的烷基过氧化氢系统。该系统由一个硫氧还原酶（FA608）、一个硫氧还原酶/谷硫磷同源物（FA1411/FA455）和一个典型的 2-半胱氨酸过氧化还原酶/AhpC（FA768）组成。FA608 与 FA1411/FA455 一起，可作为硫代氧化还原酶系统来还原胰岛素、DTNB 和 FA768。FA455 是一种类似于谷胱甘肽的蛋白质，在 F. alocis 中具有硫代毒素功能。FA768/FA608/FA1411和FA768/FA608/FA455还原酶系统都依赖于NADPH，并对广泛的过氧化氢底物H2O2、CHP和t-BHP表现出特异性：这是首次研究牙周病原体的硫代氧化还原酶依赖性烷基过氧化氢系统。由于可能缺乏过氧化氢酶或额外的过氧化还原酶同源物，该系统可保护 F. alocis 免受氧化应激。

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来源期刊

Molecular Oral Microbiology DENTISTRY, ORAL SURGERY & MEDICINE-MICROBIOLOGY

CiteScore

6.50

自引率

5.40%

发文量

审稿时长

>12 weeks

期刊介绍： Molecular Oral Microbiology publishes high quality research papers and reviews on fundamental or applied molecular studies of microorganisms of the oral cavity and respiratory tract, host-microbe interactions, cellular microbiology, molecular ecology, and immunological studies of oral and respiratory tract infections. Papers describing work in virology, or in immunology unrelated to microbial colonization or infection, will not be acceptable. Studies of the prevalence of organisms or of antimicrobials agents also are not within the scope of the journal. The journal does not publish Short Communications or Letters to the Editor. Molecular Oral Microbiology is published bimonthly.