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Role of arginine 220 in the oxygen sensor FixL from Bradyrhizobium japonicum

Abstract : In the heme-based oxygen sensor protein FixL, conformational changes induced by oxygen binding to the heme sensor domain regulate the activity of a neighboring histidine kinase, eventually restricting expression of specific genes to hypoxic conditions. The conserved arginine 220 residue is suggested to play a key role in the signal transduction mechanism. To obtain detailed insights into the role of this residue, we replaced Arg220 by histidine (R220H), glutamine (R220Q), glutamate (R220E), and isoleucine (R220I) in the heme domain FixLH from Bradyrhizobium japonicum. These mutations resulted in dramatic changes in the O2 affinity with Kd values in the order R220I < R220Q < wild type < R220H. For the R220H and R220Q mutants, residue 220 interacts with the bound O2 or CO ligands, as seen by resonance Raman spectroscopy. For the oxy-adducts, this H-bond modifies the π acidity of the O2 ligand, and its strength is correlated with the back-bonding-sensitive ν4 frequency, the koff value for O2 dissociation, and heme core-size conformational changes. This effect is especially strong for the wild-type protein where Arg220 is, in addition, positively charged. These observations strongly suggest that neither strong ligand fixation nor the displacement of residue 220 into the heme distal pocket are solely responsible for the reported heme conformational changes associated with kinase activity regulation, but that a significant decrease of the heme π* electron density because of strong back-bonding toward the oxygen ligand also plays a key role.
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Contributor : Denis Roura Connect in order to contact the contributor
Submitted on : Friday, June 7, 2013 - 4:44:07 PM
Last modification on : Tuesday, October 19, 2021 - 5:12:18 PM

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Véronique Balland, Latifa Bouzhir-Sima, Laurent Kiger, Michael C. Marden, Marten H. Vos, et al.. Role of arginine 220 in the oxygen sensor FixL from Bradyrhizobium japonicum. Journal of Biological Chemistry, 2005, 280, pp.15279-15288. ⟨10.1074/jbc.M413928200⟩. ⟨hal-00831800⟩



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