Glasser, NR;
Oyala, PH;
Osborne, TH;
Santini, JM;
Newman, DK;
(2018)
Structural and mechanistic analysis of the arsenate respiratory reductase provides insight into environmental arsenic transformations.
Proceedings of the National Academy of Sciences of the United States of America
, 115
(37)
E8614-E8623.
10.1073/pnas.1807984115.
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Abstract
Arsenate respiration by bacteria was discovered over two decades ago and is catalyzed by diverse organisms using the well-conserved Arr enzyme complex. Until now, the mechanisms underpinning this metabolism have been relatively opaque. Here we report the first structure of an Arr complex (solved by X-ray crystallography to 1.6 Å resolution), which was enabled by an improved Arr expression method in the genetically-tractable arsenate respirer Shewanella sp. ANA-3. We also obtained structures bound with the substrate arsenate (1.8 Å), the product arsenite (1.8 Å), and the natural inhibitor phosphate (1.7 Å). The structures reveal a conserved active-site motif that distinguishes Arr ((R/K)GRY) from the closely-related arsenite respiratory oxidase (Arx) complex (XGRGWG). Arr activity assays using methyl viologen as the electron donor and arsenate as the electron acceptor display two-site ping-pong kinetics. A Mo(V) intermediate was detected with electron paramagnetic resonance spectroscopy, which is typical for proteins with a molybdopterin guanine dinucleotide cofactor. Arr is an extraordinarily fast enzyme that approaches the diffusion limit (Km = 44.6 ± 1.6 μM, kcat = 9,810 ± 220 s−1 13 ) and phosphate is a competitive inhibitor of arsenate reduction (Ki = 325 ± 12 μM). These observations, combined with knowledge of typical sedimentary arsenate and phosphate concentrations and known rates of arsenate desorption from minerals in the presence of phosphate, suggest that: (1) arsenate desorption limits microbiologicallyinduced arsenate reductive mobilization and (2) phosphate enhances arsenic mobility by stimulating arsenate desorption rather than by inhibiting it at the enzymatic level.
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