Insights into the recognition and electron transfer steps in nitric oxide reductase from Marinobacter hydrocarbonoclasticus

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Abstract

Marinobacter hydrocarbonoclasticus nitric oxide reductase, cNOR, is an integral membrane protein composed of two subunits with different roles, NorC (electron transfer) and NorB (catalytic) that receives electrons from the soluble cytochrome c552 and reduces nitric oxide to nitrous oxide in the denitrification pathway. The solvent-exposed domain of NorC, harboring a c-type heme was heterologously produced, along with its physiological electron donor, cytochrome c552. These two proteins were spectroscopically characterized and shown to be similar to the native proteins, both being low-spin and Met-His coordinated, with the soluble domain of NorC presenting some additional features of a high-spin heme, which is consistent with the higher solvent accessibility of its heme and weaker coordination of the methionine axial ligand. The electron transfer complex between the two proteins has a 1:1 stoichiometry, and an upper limit for the dissociation constant was estimated by 1H NMR titration to be 1.2 ± 0.4 μM. Electrochemical techniques were used to characterize the interaction between the proteins, and a model structure of the complex was obtained by molecular docking. The electrochemical observations point to the modulation of the NorC reduction potential by the presence of NorB, tuning its ability to receive electrons from cytochrome c552.

Original languageEnglish
Pages (from-to)402-411
Number of pages10
JournalJournal of Inorganic Biochemistry
Volume177
DOIs
Publication statusPublished - Dec 2017

Fingerprint

Marinobacter
Electrons
Heme
Proteins
Electrochemical Techniques
Denitrification
Nitrous Oxide
Model structures
Titration
Stoichiometry
Methionine
Nitric Oxide
Membrane Proteins
Tuning
Nuclear magnetic resonance
Modulation
nitric-oxide reductase
Ligands
cytochrome C-552

Keywords

  • Cytochrome c heterologous production
  • Denitrification
  • Electron transfer complex
  • Marinobacter
  • Nitric oxide redutase
  • Protein interaction

Cite this

@article{49b60648eda640268de1efdbeb03cb68,
title = "Insights into the recognition and electron transfer steps in nitric oxide reductase from Marinobacter hydrocarbonoclasticus",
abstract = "Marinobacter hydrocarbonoclasticus nitric oxide reductase, cNOR, is an integral membrane protein composed of two subunits with different roles, NorC (electron transfer) and NorB (catalytic) that receives electrons from the soluble cytochrome c552 and reduces nitric oxide to nitrous oxide in the denitrification pathway. The solvent-exposed domain of NorC, harboring a c-type heme was heterologously produced, along with its physiological electron donor, cytochrome c552. These two proteins were spectroscopically characterized and shown to be similar to the native proteins, both being low-spin and Met-His coordinated, with the soluble domain of NorC presenting some additional features of a high-spin heme, which is consistent with the higher solvent accessibility of its heme and weaker coordination of the methionine axial ligand. The electron transfer complex between the two proteins has a 1:1 stoichiometry, and an upper limit for the dissociation constant was estimated by 1H NMR titration to be 1.2 ± 0.4 μM. Electrochemical techniques were used to characterize the interaction between the proteins, and a model structure of the complex was obtained by molecular docking. The electrochemical observations point to the modulation of the NorC reduction potential by the presence of NorB, tuning its ability to receive electrons from cytochrome c552.",
keywords = "Cytochrome c heterologous production, Denitrification, Electron transfer complex, Marinobacter, Nitric oxide redutase, Protein interaction",
author = "Susana Ramos and Almeida, {Rui M.} and Cordas, {Cristina M.} and Moura, {Jos{\'e} J.G.} and Pauleta, {Sofia R.} and Isabel Moura",
note = "Sem PDF. Fundacao para a Ciencia e Tecnologia (FCT) (SFRH/BD/91288/2012; FRH/BPD/80293/2011; PTDC/QUI-BIQ/116481/2010; PTDC/BBB-BQB/0129/2014; PTDC/BIA-MIC/098882/2008; PTDC/BIA-PRO/109796/2009) Unidade de Ciencias Biomoleculares Aplicadas-UCIBIO - FCT/MEC (UID/Multi/04378/2013) ERDF (POCI-01-0145-FEDER-007728) FCT",
year = "2017",
month = "12",
doi = "10.1016/j.jinorgbio.2017.09.001",
language = "English",
volume = "177",
pages = "402--411",
journal = "Journal of Inorganic Biochemistry",
issn = "0162-0134",
publisher = "Elsevier Inc.",

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TY - JOUR

T1 - Insights into the recognition and electron transfer steps in nitric oxide reductase from Marinobacter hydrocarbonoclasticus

AU - Ramos, Susana

AU - Almeida, Rui M.

AU - Cordas, Cristina M.

AU - Moura, José J.G.

AU - Pauleta, Sofia R.

AU - Moura, Isabel

N1 - Sem PDF. Fundacao para a Ciencia e Tecnologia (FCT) (SFRH/BD/91288/2012; FRH/BPD/80293/2011; PTDC/QUI-BIQ/116481/2010; PTDC/BBB-BQB/0129/2014; PTDC/BIA-MIC/098882/2008; PTDC/BIA-PRO/109796/2009) Unidade de Ciencias Biomoleculares Aplicadas-UCIBIO - FCT/MEC (UID/Multi/04378/2013) ERDF (POCI-01-0145-FEDER-007728) FCT

PY - 2017/12

Y1 - 2017/12

N2 - Marinobacter hydrocarbonoclasticus nitric oxide reductase, cNOR, is an integral membrane protein composed of two subunits with different roles, NorC (electron transfer) and NorB (catalytic) that receives electrons from the soluble cytochrome c552 and reduces nitric oxide to nitrous oxide in the denitrification pathway. The solvent-exposed domain of NorC, harboring a c-type heme was heterologously produced, along with its physiological electron donor, cytochrome c552. These two proteins were spectroscopically characterized and shown to be similar to the native proteins, both being low-spin and Met-His coordinated, with the soluble domain of NorC presenting some additional features of a high-spin heme, which is consistent with the higher solvent accessibility of its heme and weaker coordination of the methionine axial ligand. The electron transfer complex between the two proteins has a 1:1 stoichiometry, and an upper limit for the dissociation constant was estimated by 1H NMR titration to be 1.2 ± 0.4 μM. Electrochemical techniques were used to characterize the interaction between the proteins, and a model structure of the complex was obtained by molecular docking. The electrochemical observations point to the modulation of the NorC reduction potential by the presence of NorB, tuning its ability to receive electrons from cytochrome c552.

AB - Marinobacter hydrocarbonoclasticus nitric oxide reductase, cNOR, is an integral membrane protein composed of two subunits with different roles, NorC (electron transfer) and NorB (catalytic) that receives electrons from the soluble cytochrome c552 and reduces nitric oxide to nitrous oxide in the denitrification pathway. The solvent-exposed domain of NorC, harboring a c-type heme was heterologously produced, along with its physiological electron donor, cytochrome c552. These two proteins were spectroscopically characterized and shown to be similar to the native proteins, both being low-spin and Met-His coordinated, with the soluble domain of NorC presenting some additional features of a high-spin heme, which is consistent with the higher solvent accessibility of its heme and weaker coordination of the methionine axial ligand. The electron transfer complex between the two proteins has a 1:1 stoichiometry, and an upper limit for the dissociation constant was estimated by 1H NMR titration to be 1.2 ± 0.4 μM. Electrochemical techniques were used to characterize the interaction between the proteins, and a model structure of the complex was obtained by molecular docking. The electrochemical observations point to the modulation of the NorC reduction potential by the presence of NorB, tuning its ability to receive electrons from cytochrome c552.

KW - Cytochrome c heterologous production

KW - Denitrification

KW - Electron transfer complex

KW - Marinobacter

KW - Nitric oxide redutase

KW - Protein interaction

UR - http://www.scopus.com/inward/record.url?scp=85029632658&partnerID=8YFLogxK

U2 - 10.1016/j.jinorgbio.2017.09.001

DO - 10.1016/j.jinorgbio.2017.09.001

M3 - Article

VL - 177

SP - 402

EP - 411

JO - Journal of Inorganic Biochemistry

JF - Journal of Inorganic Biochemistry

SN - 0162-0134

ER -