PNNL

Abstract

Brucella melitensis is the etiological agent responsible for brucellosis. Present in the B. melitensis genome is a 116-residue protein related to arsenate reductases (YffB – BR0369). Arsenate reductases (ArsC) convert arsenate ion (H2AsO4-), a compound toxic to bacteria, to arsenite ion (As2-), a product that may be efficiently exported out of the cell. Consequently, Bm-YffB is a potential drug target because if arsenate reduction is the protein's major biological function, disabling the cell’s ability to reduce arsenate would make these cells more sensitive to the deleterious effects of arsenate. Size exclusion chromatography and NMR spectroscopy indicate that Bm-YffB is a monomer in solution. The solution structure for Bm-YffB (PDB ID 2KOK) shows the protein consists of two domains: a four-stranded mixed ß-sheet flanked by two a-helices on one side and an a-helical bundle. The a/ß domain is characteristic of the fold in thioredoxin-like proteins and the overall structure is generally similar to known arsenate reductases dispite the marginal sequence similarity. Chemical shift perturbation studies with 15N-labeled Bm-YffB show that the protein binds reduced glutathione at a site adjacent to a region similar to the HX3CX3R catalytic sequence motif that is important for the arsenic detoxification activity in the classical arsenate reductase family of proteins. The latter observation supports the hypothesis that the ArsC-YffB family of proteins may function as glutathione-dependent thiol reductases. Comparison of the structure of Bm-YffB to the structure of proteins in the classical ArsC family suggest the mechanism, and possibly the function, of Bm-YffB and other related proteins (ArsC-YffB) may differ from the ArsC family of proteins.