The anaerobic bacterium Chrysiogenes arsenatis respires using the oxyanion arsenate (AsO4 3-) as the terminal electron acceptor, where it is reduced to arsenite (AsO3 3-) while concomitantly oxidizing various organic (e.g. acetate and formate) or inorganic (e.g. hydrogen) electron donors. This respiratory activity is catalyzed in the periplasm of the bacterium by the enzyme arsenate reductase (Arr), with expression of the enzyme controlled by a sensor histidine kinase (ArrS) and a periplasmic binding protein (PBP), ArrX. Here, we report for the first time, the molecular structure of ArrX in the absence and presence of bound ligand, arsenate. Comparison of the ligand-bound structure of ArrX with other PBPs shows a high level of conservation of critical residues for ligand binding by these proteins, however this suite of PBPs show different structural alterations upon ligand binding. For ArrX and its homologue AioX (from Rhizobium sp. str. NT-26), which specifically binds arsenite, the structures of the substrate binding sites in the vicinity of a conserved and critical cysteine residue, contribute to the discrimination of binding for these chemically similar ligands.

The arsenite oxidase (AioAB) from Pseudorhizobium banfieldiae sp. strain NT-26 catalyzes the oxidation of arsenite to arsenate and transfers electrons to its cognate electron acceptor cytochrome c 552 (cytc 552). This activity underpins the ability of this organism to respire using arsenite present in contaminated environments. The crystal structure of the AioAB/cytc 552 electron transfer complex reveals two A2B2/(cytc 552)2 assemblies per asymmetric unit. Three of the four cytc 552 molecules in the asymmetric unit dock to AioAB in a cleft at the interface between the AioA and AioB subunits, with an edge-to-edge distance of 7.5 Å between the heme of cytc 552 and the [2Fe–2S] Rieske cluster in the AioB subunit. The interface between the AioAB and cytc 552 proteins features electrostatic and nonpolar interactions and is stabilized by two salt bridges. A modest number of hydrogen bonds, salt bridges and relatively small, buried surface areas between protein partners are typical features of transient electron transfer complexes. Interestingly, the fourth cytc 552 molecule is positioned differently between two AioAB heterodimers, with distances between its heme and the AioAB redox active cofactors that are outside the acceptable range for fast electron transfer. This unique cytc 552 molecule appears to be positioned to facilitate crystal packing rather than reflecting a functional complex.

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