Kinetic and thermodynamic resolution of the interactions between sulfite and the pentahaem cytochrome NrfA from Escherichia coli

NrfA is a pentahaem cytochrome present in a wide-range of γ-, δ- and ε-proteobacteria. Its nitrite and nitric oxide reductase activities have been studied extensively and contribute to respiratory nitrite ammonification and nitric oxide detoxification respectively. Sulfite is a third substrate for NrfA that may be encountered in the micro-oxic environments where nrfA is expressed. Consequently, we have performed quantitative kinetic and thermodynamic studies of the interactions between sulfite and Escherichia coli NrfA to provide a biochemical framework from which to consider their possible cellular consequences. A combination of voltammetric, spectroscopic and crystallographic analyses define dissociation constants for sulfite binding to NrfA in oxidized (~54 μM), semi-reduced (~145 μM) and reduced (~180 μM) states that are comparable with each other, and the Km (~70 μM) for sulfite reduction at pH 7. Under comparable conditions Km values of ~22 and ~300 μM describe nitrite and nitric oxide reduction respectively, whereas the affinities of nitrate and thiocyanate for NrfA fall more than 50-fold on enzyme reduction. These results are discussed in terms of the nature of sulfite co-ordination within the active site of NrfA and their implications for the cellular activity of NrfA.

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Diversity and evolution of nitric oxide reduction

10.1101/2021.10.15.464467 ◽

2021 ◽

Author(s):

James Hemp ◽

Ranjani Murali ◽

Laura A Pace ◽

Robert A Sanford ◽

Roland Hatzenpichler ◽

...

Keyword(s):

Nitric Oxide ◽

Active Site ◽

Phylogenetic Analyses ◽

Essential Element ◽

Nitric Oxide Reductase ◽

Oxide Reduction ◽

Rhodothermus Marinus ◽

Fixed Nitrogen ◽

Proton Channels ◽

Nitric Oxide Reduction

Nitrogen is an essential element for life, with the availability of fixed nitrogen limiting productivity in many ecosystems. The return of oxidized nitrogen species to the atmospheric N2 pool is predominately catalyzed by microbial denitrification (NO3- → NO2- → NO → N2O → N2). Incomplete denitrification can produce N2O as a terminal product, leading to an increase in atmospheric N2O, a potent greenhouse and ozone depleting gas2. The production of N2O is catalyzed by nitric oxide reductase (NOR) members of the heme-copper oxidoreductase (HCO) superfamily3. Here we propose that a number of uncharacterized HCO families perform nitric oxide reduction and demonstrate that an enzyme from Rhodothermus marinus, belonging to one of these families does perform nitric oxide reduction. These families have novel active-site structures and several have conserved proton channels, suggesting that they might be able to couple nitric oxide reduction to energy conservation. They also exhibit broad phylogenetic and environmental distributions, expanding the diversity of microbes that can perform denitrification. Phylogenetic analyses of the HCO superfamily demonstrate that nitric oxide reductases evolved multiple times independently from oxygen reductases, suggesting that complete denitrification evolved after aerobic respiration.

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Kinetics of electron transfer from NADH to the Escherichia coli nitric oxide reductase flavorubredoxin

FEBS Journal ◽

10.1111/j.1742-4658.2006.05612.x ◽

2006 ◽

Vol 274 (3) ◽

pp. 677-686 ◽

Cited By ~ 14

Author(s):

João B. Vicente ◽

Francesca M. Scandurra ◽

João V. Rodrigues ◽

Maurizio Brunori ◽

Paolo Sarti ◽

...

Keyword(s):

Nitric Oxide ◽

Escherichia Coli ◽

Electron Transfer ◽

Nitric Oxide Reductase ◽

Kinetics Of

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Reaction of Carbon Monoxide with the Reduced Active Site of Bacterial Nitric Oxide Reductase†

Biochemistry ◽

10.1021/bi011428t ◽

2001 ◽

Vol 40 (44) ◽

pp. 13361-13369 ◽

Cited By ~ 27

Author(s):

Janneke H. M. Hendriks ◽

Louise Prior ◽

Adam R. Baker ◽

Andrew J. Thomson ◽

Matti Saraste ◽

...

Keyword(s):

Nitric Oxide ◽

Carbon Monoxide ◽

Active Site ◽

Nitric Oxide Reductase

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Kinetic Characterization of the Escherichia coli Nitric Oxide Reductase Flavorubredoxin

Globins and Other Nitric Oxide-Reactive Proteins, Part B - Methods in Enzymology ◽

10.1016/s0076-6879(07)37003-1 ◽

2008 ◽

pp. 47-62 ◽

Cited By ~ 10

Author(s):

João B. Vicente ◽

Francesca M. Scandurra ◽

Elena Forte ◽

Maurizio Brunori ◽

Paolo Sarti ◽

...

Keyword(s):

Nitric Oxide ◽

Escherichia Coli ◽

Kinetic Characterization ◽

Nitric Oxide Reductase

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The role of Bradyrhizobium japonicum nitric oxide reductase in nitric oxide detoxification in soya bean root nodules

Biochemical Society Transactions ◽

10.1042/bst0340195 ◽

2006 ◽

Vol 34 (1) ◽

pp. 195-196 ◽

Cited By ~ 17

Author(s):

G.E. Meakin ◽

B.J.N. Jepson ◽

D.J. Richardson ◽

E.J. Bedmar ◽

M.J. Delgado

Keyword(s):

Nitric Oxide ◽

Bradyrhizobium Japonicum ◽

Root Nodules ◽

Soya Bean ◽

Nitric Oxide Reductase ◽

Bean Root ◽

Detoxification Systems ◽

Nitric Oxide Detoxification

The identification of nitric oxide-bound leghaemoglobin within soya bean nodules has led to the question of how Bradyrhizobium japonicum bacteroids overcome the toxicity of this nitric oxide. It has previously been shown that one candidate for nitric oxide detoxification, the respiratory nitric oxide reductase, is expressed in soya bean nodules from plants supplied with nitrate [Mesa, de Dios Alché, Bedmar and Delgado (2004) Physiol. Plant. 120, 205–211]. In this paper, the role of this enzyme in nitric oxide detoxification is assessed and discussion is provided on other possible B. japonicum nitric oxide detoxification systems.

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Dioxygen and nitric oxide reactivity of a reduced heme/non-heme diiron(II) complex [(5L)FeII⋯FeIICl]+. Using a tethered tetraarylporphyrin for the development of an active site reactivity model for bacterial nitric oxide reductase

Inorganica Chimica Acta ◽

10.1016/s0020-1693(99)00404-1 ◽

2000 ◽

Vol 297 (1-2) ◽

pp. 362-372 ◽

Cited By ~ 11

Author(s):

Telvin D. Ju ◽

Amina S. Woods ◽

Robert J. Cotter ◽

Pierre Moënne-Loccoz ◽

Kenneth D. Karlin

Keyword(s):

Nitric Oxide ◽

Active Site ◽

Nitric Oxide Reductase

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The nitric oxide reductase of enterohaemorrhagic Escherichia coli plays an important role for the survival within macrophages

Molecular Microbiology ◽

10.1111/j.1365-2958.2012.08122.x ◽

2012 ◽

Vol 85 (3) ◽

pp. 492-512 ◽

Cited By ~ 45

Author(s):

Takeshi Shimizu ◽

Hiroyasu Tsutsuki ◽

Akio Matsumoto ◽

Haruaki Nakaya ◽

Masatoshi Noda

Keyword(s):

Nitric Oxide ◽

Escherichia Coli ◽

Nitric Oxide Reductase ◽

Enterohaemorrhagic Escherichia Coli

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Glyoxalase III from Escherichia coli: a single novel enzyme for the conversion of methylglyoxal into d-lactate without reduced glutathione

Biochemical Journal ◽

10.1042/bj3050999 ◽

1995 ◽

Vol 305 (3) ◽

pp. 999-1003 ◽

Cited By ~ 67

Author(s):

K Misra ◽

A B Banerjee ◽

S Ray ◽

M Ray

Keyword(s):

Escherichia Coli ◽

Active Site ◽

Carbonyl Compounds ◽

Reduced Glutathione ◽

Thiol Group ◽

Glyoxalase I ◽

Ph Optimum ◽

Native Form ◽

Wide Range ◽

Glyoxalase Iii

A single novel enzyme, glyoxalase III, which catalyses the conversion of methylglyoxal into D-lactate without involvement of GSH, has been detected in and purified from Escherichia coli. Of several carbonyl compounds tested, only the alpha-ketoaldehydes methylglyoxal and phenylglyoxal were found to be substrates for this enzyme. Glyoxalase III is active over a wide range of pH with no sharp pH optimum. In its native form it has an M(r) of 82000 +/- 2000, and it is composed of two subunits of equal M(r). Glutathione analogues, which are inhibitors of glyoxalase I, do not inhibit glyoxalase III. Glyoxalase III is found to be sensitive to thiol-blocking reagents. The p-hydroxymercuribenzoate-inactivated enzyme could be almost completely re-activated by dithiothreitol and other thiol-group-containing compounds, indicating the possible involvement of thiol group(s) at or near the active site of the enzyme.

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