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Journal Articles Journal of Biological Chemistry Year : 2011

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 Trp(66), which can modulate this proximal H-bond. We investigated the effects of these mutations on different NOS complexes (Fe-III, (FeCO)-C-II, and (FeNO)-N-II), 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 sigma-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.

Domains

Biophysics

Dates and versions

hal-00720044 , version 1 (23-07-2012)

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Cite

A. Brunel, A. Wilson, L. Henry, P. Dorlet, J. Santolini. The Proximal Hydrogen Bond Network Modulates Bacillus subtilis Nitric-oxide synthase Electronic and structural Properties. Journal of Biological Chemistry, 2011, 286 (14), pp.9. ⟨10.1074/jbc.M110.195446⟩. ⟨hal-00720044⟩

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