Globin-coupled sensors (GCSs) are multiple-domain transducers, consisting of a regulatory globin-like heme-binding domain and a linked transducer domain(s). GCSs have been described in both Archaea and bacteria. They are generally assumed to bind O2 (and perhaps other gaseous ligands) and to transmit a conformational change signal through the transducer domain in response to fluctuating O2 levels. In this study, the heme-binding domain, AvGReg178, and the full protein, AvGReg of the Azotobacter vinelandii GCS, were cloned, expressed, and purified. After purification, the heme iron of AvGReg178 was found to bind O2. This form was stable over many hours. In contrast, the predominant presence of a bis-histidine coordinate heme in ferric AvGReg was revealed. Differences in the heme pocket structure were also observed for the deoxygenated ferrous state of these proteins. The spectra showed that the deoxygenated ferrous derivatives of AvGReg178 and AvGReg are characterized by a penta-coordinate and hexa-coordinate heme iron, respectively. O2 binding isotherms indicate that AvGReg178 and AvGReg show a high affinity for O2 with P50 values at 20 °C of 0.04 and 0.15 torr, respectively. Kinetics of CO binding indicate that AvGReg178 carbonylation conforms to a monophasic process, comparable with that of myoglobin, whereas AvGReg carbonylation conforms to a three-phasic reaction, as observed for several proteins with bis-histidine heme iron coordination. Besides sensing ligands, in vitro data suggest that AvGReg(178) may have a role in O2-mediated NO-detoxification, yielding metAvGReg(178) and nitrate. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc.

Thijs, L., Vinck, E., Bolli, A., Trandafir, F., Wan, X., Hoogewijs, D., et al. (2007). Characterization of a globin-coupled oxygen sensor with a gene-regulating function. THE JOURNAL OF BIOLOGICAL CHEMISTRY, 282(52), 37325-37340 [10.1074/jbc.M705541200].

Characterization of a globin-coupled oxygen sensor with a gene-regulating function

COLETTA, MASSIMILIANO;
2007-01-01

Abstract

Globin-coupled sensors (GCSs) are multiple-domain transducers, consisting of a regulatory globin-like heme-binding domain and a linked transducer domain(s). GCSs have been described in both Archaea and bacteria. They are generally assumed to bind O2 (and perhaps other gaseous ligands) and to transmit a conformational change signal through the transducer domain in response to fluctuating O2 levels. In this study, the heme-binding domain, AvGReg178, and the full protein, AvGReg of the Azotobacter vinelandii GCS, were cloned, expressed, and purified. After purification, the heme iron of AvGReg178 was found to bind O2. This form was stable over many hours. In contrast, the predominant presence of a bis-histidine coordinate heme in ferric AvGReg was revealed. Differences in the heme pocket structure were also observed for the deoxygenated ferrous state of these proteins. The spectra showed that the deoxygenated ferrous derivatives of AvGReg178 and AvGReg are characterized by a penta-coordinate and hexa-coordinate heme iron, respectively. O2 binding isotherms indicate that AvGReg178 and AvGReg show a high affinity for O2 with P50 values at 20 °C of 0.04 and 0.15 torr, respectively. Kinetics of CO binding indicate that AvGReg178 carbonylation conforms to a monophasic process, comparable with that of myoglobin, whereas AvGReg carbonylation conforms to a three-phasic reaction, as observed for several proteins with bis-histidine heme iron coordination. Besides sensing ligands, in vitro data suggest that AvGReg(178) may have a role in O2-mediated NO-detoxification, yielding metAvGReg(178) and nitrate. © 2007 by The American Society for Biochemistry and Molecular Biology, Inc.
Pubblicato
Rilevanza internazionale
Articolo
Sì, ma tipo non specificato
Settore BIO/10
English
Bacteria; Carbonylation; Genes; Isotherms; Ligands; Proteins; Archaea; Globin coupled sensors (GCS); Oxygen sensors; bacterial protein; carbon monoxide; ferrous ion; globin; heme; iron; myoglobin; protein AvGReg; protein AvGReg178; Archaebacterium; article; Azotobacter vinelandii; bacterium; binding affinity; carbonylation; conformational transition; detoxification; gene control; molecular cloning; nonhuman; nucleotide sequence; oxygen dissociation curve; oxygen sensing; priority journal; protein domain; protein expression; protein function; protein purification; signal transduction; Amino Acid Sequence; Azotobacter vinelandii; Bacterial Proteins; Escherichia coli; Gene Expression Regulation, Bacterial; Heme; Hemeproteins; Histidine; Iron; Kinetics; Ligands; Models, Biological; Molecular Sequence Data; Nitric Oxide; Oxygen; Protein Structure, Tertiary; Sequence Homology, Amino Acid; Spectrum Analysis, Raman; Time Factors; Azotobacter vinelandii
Thijs, L., Vinck, E., Bolli, A., Trandafir, F., Wan, X., Hoogewijs, D., et al. (2007). Characterization of a globin-coupled oxygen sensor with a gene-regulating function. THE JOURNAL OF BIOLOGICAL CHEMISTRY, 282(52), 37325-37340 [10.1074/jbc.M705541200].
Thijs, L; Vinck, E; Bolli, A; Trandafir, F; Wan, X; Hoogewijs, D; Coletta, M; Fago, A; Weber, R; Van Doorslaer, S; Ascenzi, P; Alam, M; Moens, L; Dewilde, S
Articolo su rivista
File in questo prodotto:
File Dimensione Formato  
JBC2007Moens.pdf

accesso aperto

Dimensione 990.97 kB
Formato Adobe PDF
990.97 kB Adobe PDF Visualizza/Apri

I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.

Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/2108/41165
Citazioni
  • ???jsp.display-item.citation.pmc??? 8
  • Scopus ND
  • ???jsp.display-item.citation.isi??? 32
social impact