title: The proximal hydrogen bond network modulates Bacillus subtilis nitric-oxide synthase electronic and structural properties.
abstract: Bacterial nitric-oxide synthase (NOS)-like proteins are believed to be genuine NOSs. As for cytochromes P450 (CYPs), NOS-proximal ligand is a thiolate that exerts a push effect crucial for the process of dioxygen activation. Unlike CYPs, this catalytic electron donation seems controlled by a hydrogen bond (H-bond) interaction between the thiolate ligand and a vicinal tryptophan. Variations of the strength of this H-bond could provide a direct way to tune the stability along with the electronic and structural properties of NOS. We generated five different mutations of bsNOS Trp66, which can modulate this proximal H-bond. We investigated the effects of these mutations on different NOS complexes (FeIII, FeIICO, and FeIINO), using a combination of UV-visible absorption, EPR, FTIR, and resonance Raman spectroscopies. Our results indicate that (i) the proximal H-bond modulation can selectively decrease or increase the electron donating properties of the proximal thiolate, (ii) this modulation controls the σ-competition between distal and proximal ligands, (iii) this H-bond controls the stability of various NOS intermediates, and (iv) a fine tuning of the electron donation by the proximal ligand is required to allow at the same time oxygen activation and to prevent uncoupling reactions.
authors.affiliation: Commissariat à l'Energie Atomique, iBiTec-S, SBSM, F-91191 Gif-sur-Yvette, France.
authors.forename: Albane
authors.initials: A
authors.lastname: Brunel
authors.forename: Adjélé
authors.initials: A
authors.lastname: Wilson
authors.forename: Laura
authors.initials: L
authors.lastname: Henry
authors.forename: Pierre
authors.initials: P
authors.lastname: Dorlet
authors.forename: Jérôme
authors.initials: J
authors.lastname: Santolini
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology*; D004578:Electron Spin Resonance Spectroscopy; D006860:Hydrogen Bonding; D019001:Nitric Oxide Synthase / Q000737:chemistry* / Q000378:metabolism*; D018384:Oxidative Stress; D017433:Protein Structure, Secondary; D017550:Spectroscopy, Fourier Transform Infrared; D013059:Spectrum Analysis, Raman
chemical_list: D019001:Nitric Oxide Synthase