The thioredoxin Lmo1903 contributes to oxidative stress tolerance of Listeria monocytogenes

Abstract:

[Objective] The thioredoxin family plays a key role in the oxidative stress response of the foodborne bacterial pathogen Listeria monocytogenes (LM) during environmental adaptation. Here, we studied the biological role of the thioredoxin Lmo1903 in oxidative stress tolerance.[Methods] The phylogenetic relationship and key active sites of Lmo1903 were analyzed by bioinformatics tools. The recombinant Lmo1903 protein was expressed and purified. The oxidoreductase activity of the recombination protein was determined with insulin as the substrate and the cellular localization was predicted after preparation of mouse polyclonal antibody with the recombinant protein. The oligonucleotide-directed site-specific deletion of cysteine from Lmo1903 protein was carried out to analyze the key sites of enzyme activity. The lmo1903-deleted strain Δlmo1903 was constructed by homologous recombination, and the complementation strain CΔlmo1903was constructed with the integrated complement plasmid pIMK2. The growth, motility, and oxidative stress tolerance of the strains were examined in vitro. [Results] Lmo1903 had a classical CX₁X₂C motif and was close related to the thioredoxin family member TrxA from Bacillus subtilis. It was mainly located in the cytoplasm of bacteria and possessed strong reductase activity. Cysteine was the key site for the enzymatic activity of Lmo1903. Deletion of lmo1903 did not affect bacterial growth, while it significantly weakened the tolerance to oxidative stress in Cu²⁺ stress environment. Furthermore, the deletion of this gene affected the transcriptional levels of the genes involved in bacterial flagellar formation and reduced the swimming motility. [Conclusion] The thioredoxin family member Lmo1903 exhibiting the reductase activity contributes to the oxidative stress tolerance and motility of L. monocytogenes.