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.
Department of Physics, University of Antwerp, Universiteitsplein 1, Antwerp B-2610, Belgium.
Florin
F
Trandafir
Sabine
S
Van Doorslaer
Sylvia
S
Dewilde
Luc
L
Moens
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