Temperature dependence of NO binding modes in human neuroglobin.
Both the ferrous and ferric forms of wild-type neuroglobin are found to be hexacoordinated with axial ligation of the F8-His and E7-His. Rapidly growing Escherichia coli cell cultures with low O2 concentration generate nitric oxide (NO). Combined electron paramagnetic resonance (EPR) and optical measurements show that wild-type human recombinant neuroglobin, overexpressed in such E. coli cells, still favors the F8His-Fe2+ -E7His conformation, whereby only a small fraction of the protein binds NO. Upon mutation of the E7-His to Leu and Gln, the competition with the distal histidine disappears and the nitrosyl ferrous form is readily observed. At low temperature, the EPR spectra of the NO-ligated Ngb proteins consist of contributions from two geometrically different NO-heme conformations. In combination with EPR data of vertebrate hemoglobins and myoglobins, the temperature dependence of the EPR spectra of the NO adducts of ferrous hNgb and its E7-mutants proves a strong stabilization of one isomer by the E7-histidine in wt hNgb. It is shown that this is not related to the polarity of histidine, but to its specific binding characteristics.
Trandafir
Florin
F
Department of Physics, University of Antwerp, Universiteitsplein 1, Antwerp B-2610, Belgium.
Van Doorslaer
Sabine
S
Dewilde
Sylvia
S
Moens
Luc
L
D004578:Electron Spin Resonance Spectroscopy; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D005914:Globins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D000078787:Neuroglobin; D009569:Nitric Oxide / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013696:Temperature
D009419:Nerve Tissue Proteins; D000078787:Neuroglobin; D011994:Recombinant Proteins; D009569:Nitric Oxide; D005914:Globins