Identification of bacterial dissimilatory antimonate reductase AnrA: genes and proteins involved in antimonate respiration and resistance in Geobacter sp. strain SVR

ABSTRACT Geobacter sp. strain SVR uses antimonate Sb (V) as a terminal electron acceptor for anaerobic respiration. Here, we visualized a possible key enzyme, periplasmic Sb (V) reductase (Anr), via active staining and non-denaturing gel electrophoresis. Liquid chromatography-tandem mass spectrometry analysis revealed that a novel dimethyl sulfoxide (DMSO) reductase family protein, WP₁73201954. 1, is involved in Anr. This protein was closely related with AnrA, a protein suggested to be the catalytic subunit of a respiratory Sb (V) reductase in Desulfuribacillus stibiiarsenatis. The anr genes of strain SVR (anrXSRBAD) formed an operon-like structure, and their transcription was upregulated under Sb (V) -respiring conditions. The expression of anrA gene was induced by more than 1 µM of antimonite Sb (III) ; however, arsenite As (III) did not induce the expression of anrA gene. Tandem mass tag-based proteomic analysis revealed that, in addition to Anr proteins, proteins in the following categories were upregulated under Sb (V) -respiring conditions: (i) Sb (III) efflux systems such as Ant and Ars; (ii) antioxidizing proteins such as ferritin, rubredoxin, and thioredoxin; (iii) protein quality control systems such as HspA, HslO, and DnaK; and (iv) DNA repair proteins such as UspA and UvrB. These results suggest that strain SVR copes with antimony stress by modulating pleiotropic processes to resist and actively metabolize antimony. To the best of our knowledge, this is the first report to demonstrate the involvement of AnrA in Sb (V) respiration at the protein level. Furthermore, this is the first example to show high expression of the Ant system proteins in the Sb (V) -respiring bacterium. IMPORTANCE Antimony (Sb) exists mainly as antimonite Sb (III) or antimonate Sb (V) in the environment, and Sb (III) is more toxic than Sb (V). Recently, microbial involvement in Sb redox reactions has received attention. Although more than 90 Sb (III) -oxidizing bacteria have been reported, information on Sb (V) -reducing bacteria is limited. Especially, the enzyme involved in dissimilatory Sb (V) reduction, or Sb (V) respiration, is unclear, despite this pathway being very important for the circulation of Sb in nature. In this study, we demonstrated that the Sb (V) reductase (Anr) of an Sb (V) -respiring bacterium (Geobacter sp. SVR) is a novel member of the dimethyl sulfoxide (DMSO) reductase family. In addition, we found that strain SVR copes with Sb stress by modulating pleiotropic processes, including the Ant and Ars systems, and upregulating the antioxidant and quality control protein levels. Considering the abundance and diversity of putative anr genes in the environment, Anr may play a significant role in global Sb cycling in both marine and terrestrial environments.