Abstract
Reductive dehalogenases are responsible for the reductive cleavage of carbon-halogen bonds during organohalide respiration. A variety of mechanisms have been proposed for these cobalamin and [4Fe-4S] containing enzymes, including organocobalt, radical, or cobalt-halide adduct based catalysis. The latter was proposed for the oxygen-tolerant Nitratireductor pacificus pht-3B catabolic reductive dehalogenase (NpRdhA). Here, we present the first substrate bound NpRdhA crystal structures, confirming a direct cobalt–halogen interaction is established and providing a rationale for substrate preference. Product formation is observed in crystallo due to X-ray photoreduction. Protein engineering enables rational alteration of substrate preference, providing a future blue print for the application of this and related enzymes in bioremediation.
Highlights
The reductive dehalogenases (RDases) are key enzymes in organohalide respiration, which is performed by a unique subset of bacteria [1,2]
The RDases can be split into two classes: (i) the canonical respiratory dehalogenases, which use a halogenated compound as a final electron acceptor during organohalide respiration and (ii) the catabolic respiratory dehalogenases that occur in the catabolic pathways of non-organohalide respiring bacteria [7,8,9]
The structure of the 35-DB-4-OH: Nitratireductor pacificus pht-3B catabolic reductive dehalogenase (NpRdhA) complex reveals a network of polar interactions occurs between S422, K488 and R552 and the substrate hydroxyl group
Summary
The reductive dehalogenases (RDases) are key enzymes in organohalide respiration, which is performed by a unique subset of bacteria [1,2]. RDase containing organisms have been targeted as a potential bioremediation tool for the clean-up of contaminated anaerobic sites where organohalides have accumulated, often due to improper disposal [3,4,5,6]. The oxygen-sensitive respiratory RDases contain a twin arginine-translocation signal (TAT), allowing transport across the periplasmic membrane followed by association with a membrane anchor RdhB [10]. The catabolic RDase enzymes lack the TAT signal and tend to be oxygen-tolerant, suggesting these might provide robust future bioremediation catalysts [7,10,11,12,13]
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