title: Resolving the contributions of the membrane-bound and periplasmic nitrate reductase systems to nitric oxide and nitrous oxide production in Salmonella enterica serovar Typhimurium.
abstract: The production of cytotoxic nitric oxide (NO) and conversion into the neuropharmacological agent and potent greenhouse gas nitrous oxide (N₂O) is linked with anoxic nitrate catabolism by Salmonella enterica serovar Typhimurium. Salmonella can synthesize two types of nitrate reductase: a membrane-bound form (Nar) and a periplasmic form (Nap). Nitrate catabolism was studied under nitrate-rich and nitrate-limited conditions in chemostat cultures following transition from oxic to anoxic conditions. Intracellular NO production was reported qualitatively by assessing transcription of the NO-regulated genes encoding flavohaemoglobin (Hmp), flavorubredoxin (NorV) and hybrid cluster protein (Hcp). A more quantitative analysis of the extent of NO formation was gained by measuring production of N₂O, the end-product of anoxic NO-detoxification. Under nitrate-rich conditions, the nar, nap, hmp, norV and hcp genes were all induced following transition from the oxic to anoxic state, and 20% of nitrate consumed in steady-state was released as N₂O when nitrite had accumulated to millimolar levels. The kinetics of nitrate consumption, nitrite accumulation and N₂O production were similar to those of wild-type in nitrate-sufficient cultures of a nap mutant. In contrast, in a narG mutant, the steady-state rate of N₂O production was ~30-fold lower than that of the wild-type. Under nitrate-limited conditions, nap, but not nar, was up-regulated following transition from oxic to anoxic metabolism and very little N₂O production was observed. Thus a combination of nitrate-sufficiency, nitrite accumulation and an active Nar-type nitrate reductase leads to NO and thence N₂O production, and this can account for up to 20% of the nitrate catabolized.
authors: School of Biological Sciences, University of East Anglia, Norwich Research Park, Norwich NR4 7TJ, UK. g.rowley@uea.ac.uk
authors: Gary
authors: G
authors: Rowley
authors: Daniela
authors: D
authors: Hensen
authors: Heather
authors: H
authors: Felgate
authors: Anke
authors: A
authors: Arkenberg
authors: Corinne
authors: C
authors: Appia-Ayme
authors: Karen
authors: K
authors: Prior
authors: Carl
authors: C
authors: Harrington
authors: Sarah J
authors: SJ
authors: Field
authors: Julea N
authors: JN
authors: Butt
authors: Elizabeth
authors: E
authors: Baggs
authors: David J
authors: DJ
authors: Richardson
mesh_terms: D000332:Aerobiosis; D000693:Anaerobiosis; D015687:Cell Hypoxia; D002462:Cell Membrane / Q000201:enzymology*; D015964:Gene Expression Regulation, Bacterial; D009565:Nitrate Reductases / Q000378:metabolism*; D009566:Nitrates / Q000378:metabolism; D009572:Nitrite Reductases / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism; D009609:Nitrous Oxide / Q000378:metabolism*; D019897:Periplasm / Q000201:enzymology*; D012486:Salmonella typhimurium / Q000201:enzymology* / Q000378:metabolism
chemical_list: D009566:Nitrates; D009573:Nitrites; D009565:Nitrate Reductases; D009572:Nitrite Reductases; D009609:Nitrous Oxide