title: Nitrosylhemoglobin Wood: effects of inositol hexaphosphate on thiol reactivity and electron paramagnetic resonance spectrum.
abstract: Properties of Hb Wood (beta-97(FG4)His leads to Leu), a high oxygen affinity hemoglobin with reduced hemeheme interaction, were examined in its nitric oxide liganded form. The reactivity of the beta-93 thiol groups and the electron paramagnetic resonance (EPR) spectrum were examined to determine what effect the amino acid substitution, which occurs at the alpha1beta2 interface, would have on inositol hexaphosphate induced transition of this form of the tetramer. Binding of inositol hexaphosphate (IHP) in a 1:1 stoichiometry was demonstrated. In spite of apparently normal interaction with IHP, there was little or no change in the reactivity of the beta-93 thiol groups and in the electron paramagnetic resonance (EPR) spectrum as contrasted with the marked changes characteristic of normal hemoglobin (HbA). In contrast with NO-HbA, there was also no development of the EPR hyperfine structure in NO-Hb Wood with increased protonation of the protein at pH below 7.0. Taken together with the observations of Henry and Banerjee ((1973), J. Mol. Biol. 73, 469) on the development of NO-Hb EPR hyperfine structure and of Perutz et al. (1974a), Biochemistry 13, 2174) on changes in thiol reactivity with the R leads to T transition, the results suggest that IHP or H+ cannot switch NO-Hb Wood to the T conformation. Since the atomic structures of met- and deoxyhemoglobin offer no indication that His-97 plays any special part in the allosteric mechanism (M. E. Perutz, personal communication), it appears that the replacement of His-97 by Leu reduces the stability of the T structure relative to that of R.
mesh_terms: D001665:Binding Sites; D002248:Carbon Monoxide / Q000097:blood; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004578:Electron Spin Resonance Spectroscopy; D006455:Hemoglobins, Abnormal / Q000302:isolation & purification; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007294:Inositol / Q000031:analogs & derivatives*; D008396:Maryland; D009569:Nitric Oxide / Q000097:blood*; D010100:Oxygen / Q000097:blood; D010833:Phytic Acid / Q000097:blood*; D011485:Protein Binding; D011487:Protein Conformation; D013438:Sulfhydryl Compounds / Q000097:blood*; D014922:Wisconsin
chemical_list: D006455:Hemoglobins, Abnormal; D013438:Sulfhydryl Compounds; D009569:Nitric Oxide; D007294:Inositol; D010833:Phytic Acid; D002248:Carbon Monoxide; D010100:Oxygen