Nitrate Reductase A (NarGHI) and DMSO Reductase (DmsABC) |
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Nitrate reductase A (NarGHI) is an excellent model system to study protonmotive force generation by the redox loop mechanism. It has two substrate binding sites, one located in the cytoplasmic membrane, which releases protons into the periplasmic compartment as a result of quinol oxidation, and a nitrate reducing site that consumes cytoplasmically-localized protons as a result of nitrate reduction to nitrite. Catalytic turnover thus contributes to the protonmotive force across the cytoplasmic membrane. We are studying these two substrate binding sites and the prosthetic groups that mediate electron transfer between them. Our efforts recently culminated in a 1.9Å resolution structure of the enzyme (in collaboration with Dr. Natalie Strynadka, UBC). |
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The molybdenum cofactor of NarGHI has a number of novel features: the protein-Mo ligand is an aspartate residue; one of the two pterins is a molybdopterin rather than a pyranopterin; and there is a novel [4Fe-4S] cluster nearby that has a S=3/2 groundstate. |
We use electron paramagnetic resonance (EPR) to study NarGHI and other respiratory chain enzymes. Two spectrometers are located in the laboratory, a Bruker Elexys E500 equiped with a liquid helium cryogenic system is used to record spectra of hemes and [Fe-S] clusters, and a Bruker ESP300E equiped with a liquid nitrogen cryogenic system is used to record spectra of free radicals and molybdenum. |
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The [4Fe-4S] cluster located in the vicinity of the molybdenum cofactor of NarGHI has unusual coordination, comprising three cysteines and one histidine. This coordination results in it having unusual EPR properties in its reduced state. Rather than having an EPR spectrum located in the g=2 region, it has a spectrum comprising two visible peaks around g=5.25. Such spectra arise from paramagnetic species with high spin (S=3/2) ground states. |
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