title: Characteristics of a water-forming NADH oxidase from Methanobrevibacter smithii, an archaeon in the human gut.
abstract: NADH oxidases (NOXs) catalysing the oxidation of NADH to yield NAD+and H2O, H2O2, or both play an important role in protecting organisms from oxidative stress and maintaining the balance of NAD+/NADH. A gene encoding NOX was identified from Methanobrevibacter smithii (NOX-ms), the predominant archaeon in the human gut ecosystem. Subsequent analyses showed that it is an FAD-containing protein with a subunit molecular mass of 48 kDa. NOX-ms was purified to homogeneity after expression in Escherichia coli NOX-ms catalysed the oxidization of NADH and converted O2to H2O with an optimal pH of 7.5 and a temperature optimum of approximately 37°C. The Vmaxand Kmvalues were 42.6-44.1 unit/mg and 47.8-54.6 μM for NADH. The apparent Vmaxand Kmfor oxygen were 189.5-196.1 unit/mg and 14.6-16.8 μM. The mutation analysis suggests that Cys42in NOX-ms plays a key role in the four-electron reduction of O2to H2O. Quantitative reverse transcription-PCR (RT-qPCR) revealed that transcription of NOX-ms was also up-regulated after exposing the cells to oxidative stress and glucose. Finally, the potential of NOX-ms as a target to control colonization of M. smithii and its possible applications are discussed.
authors.affiliation: Department of Clinical Laboratory, The First People's Hospital of Shangqiu City, Shangqiu 476000, China.
authors.forename: Mingguang
authors.initials: M
authors.lastname: Yan
authors.affiliation: Department of Clinical Laboratory, The First People's Hospital of Shangqiu City, Shangqiu 476000, China.
authors.forename: Weibing
authors.initials: W
authors.lastname: Yin
authors.affiliation: Department of Clinical Laboratory, The First People's Hospital of Shangqiu City, Shangqiu 476000, China.
authors.forename: Xiao
authors.initials: X
authors.lastname: Fang
authors.affiliation: Department of Clinical Laboratory, The First People's Hospital of Shangqiu City, Shangqiu 476000, China.
authors.forename: Jianjun
authors.initials: J
authors.lastname: Guo
authors.affiliation: Department of Infectious Diseases, The Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou 510630, China shihong1969@gmail.com.
authors.forename: Hong
authors.initials: H
authors.lastname: Shi
mesh_terms: D000595:Amino Acid Sequence; D001105:Archaea / Q000235:genetics* / Q000378:metabolism; D003001:Cloning, Molecular / Q000379:methods; D004926:Escherichia coli / Q000235:genetics; D006863:Hydrogen-Ion Concentration; D041581:Methanobrevibacter / Q000235:genetics* / Q000378:metabolism; D009097:Multienzyme Complexes / Q000235:genetics* / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics* / Q000378:metabolism*; D010084:Oxidation-Reduction; D010100:Oxygen / Q000378:metabolism; D016415:Sequence Alignment; D013696:Temperature; D014867:Water / Q000378:metabolism*
chemical_list: D009097:Multienzyme Complexes; D014867:Water; C023107:NADH oxidase; D009247:NADH, NADPH Oxidoreductases; D010100:Oxygen
keywords: Methanobrevibacter smithii; NADH oxidase; enzyme kinetics; protein function