title: Achromobacter denitrificans strain YD35 pyruvate dehydrogenase controls NADH production to allow tolerance to extremely high nitrite levels.
abstract: We identified the extremely nitrite-tolerant bacterium Achromobacter denitrificans YD35 that can grow in complex medium containing 100 mM nitrite (NO2(-)) under aerobic conditions. Nitrite induced global proteomic changes and upregulated tricarboxylate (TCA) cycle enzymes as well as antioxidant proteins in YD35. Transposon mutagenesis generated NO2(-)-hypersensitive mutants of YD35 that had mutations at genes for aconitate hydratase and α-ketoglutarate dehydrogenase in the TCA cycle and a pyruvate dehydrogenase (Pdh) E1 component, indicating the importance of TCA cycle metabolism to NO2(-) tolerance. A mutant in which the pdh gene cluster was disrupted (Δpdh mutant) could not grow in the presence of 100 mM NO2(-). Nitrite decreased the cellular NADH/NAD(+) ratio and the cellular ATP level. These defects were more severe in the Δpdh mutant, indicating that Pdh contributes to upregulating cellular NADH and ATP and NO2(-)-tolerant growth. Exogenous acetate, which generates acetyl coenzyme A and then is metabolized by the TCA cycle, compensated for these defects caused by disruption of the pdh gene cluster and those caused by NO2(-). These findings demonstrate a link between NO2(-) tolerance and pyruvate/acetate metabolism through the TCA cycle. The TCA cycle mechanism in YD35 enhances NADH production, and we consider that this contributes to a novel NO2(-)-tolerating mechanism in this strain.
authors.affiliation: Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Doi
authors.forename: Motoyuki
authors.initials: M
authors.lastname: Shimizu
authors.forename: Tomoya
authors.initials: T
authors.lastname: Fujita
authors.forename: Akira
authors.initials: A
authors.lastname: Nakamura
authors.forename: Noboru
authors.initials: N
authors.lastname: Takizawa
authors.forename: Naoki
authors.initials: N
authors.lastname: Takaya
mesh_terms: D042441:Achromobacter denitrificans / Q000187:drug effects* / Q000201:enzymology* / Q000254:growth & development; D000332:Aerobiosis; D004251:DNA Transposable Elements; D024881:Drug Resistance, Bacterial; D017353:Gene Deletion; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D009243:NAD / Q000378:metabolism*; D009573:Nitrites / Q000378:metabolism* / Q000633:toxicity*; D030481:Pyruvate Dehydrogenase (Lipoamide) / Q000378:metabolism*; D017422:Sequence Analysis, DNA
chemical_list: D004251:DNA Transposable Elements; D009573:Nitrites; D009243:NAD; D030481:Pyruvate Dehydrogenase (Lipoamide)