Solution 1H NMR characterization of the distal H-bond network and the effective axial field in the resting-state, high-spin ferric, substrate-bound complex of heme oxygenase from N. meningitidis.
The solution (1)H 1D and 2D NMR spectra of the high-spin ferric, resting-state, substrate-bound complex of heme oxygenase, HO, from the pathological bacterium N. meningitidis have been investigated to assess the prospects for definitive assignment of hyperfine shifted and relaxed residue protons and the interpretation of those shifts in terms of the anisotropy and orientation of the paramagnetic susceptibility tensor, chi. Appropriately tailored 1D/2D NMR data, together with analyses of paramagnetic relaxation and a preliminary estimate of the magnetic anisotropy, reveal a chi that is axially anisotropic and oriented along the Fe-His vector. Together with T(-)(2) dependence of the shifts, Deltachi(ax) yields a zero-field splitting constant, D = 9.1 cm(-)(1), which is expected to serve as a very sensitive probe of H-bond interactions between the iron-ligated water and a series of distal ordered water molecules implicated in the mechanism of HO action. The side chains, Gln49 and His53, involved in the stabilization of catalytically relevant water molecules, were found to exhibit orientations rotated by 180 degrees about the beta-gamma bonds in solution relative to those in the crystal. The implication of these reorientations on the details of the distal H-bond network is discussed. The H-bond donor strengths of Gln 49 and His53 were found to respond appropriately to H-bond donor (water) versus H-bond acceptor (cyanide) iron ligands. Very slow NH exchange for the N-terminal portion of the distal helix suggest that an intrinsically "unstable" distal helix may be valid only for the C-terminal portion.
Liu
Yangzhong
Y
Department of Chemistry, University of California, Davis, California 95616, USA.
Zhang
Xuhong
X
Yoshida
Tadashi
T
La Mar
Gerd N
GN
D000596:Amino Acids / Q000737:chemistry / Q000378:metabolism; D005290:Ferric Compounds / Q000737:chemistry* / Q000378:metabolism; D006419:Heme Oxygenase (Decyclizing) / Q000737:chemistry* / Q000378:metabolism; D006860:Hydrogen Bonding; D008958:Models, Molecular; D009345:Neisseria meningitidis / Q000201:enzymology*; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D011487:Protein Conformation; D012996:Solutions; D014867:Water / Q000737:chemistry
D000596:Amino Acids; D005290:Ferric Compounds; D012996:Solutions; D014867:Water; D006419:Heme Oxygenase (Decyclizing)