Nitrogen binding to deoxyhemoglobin at high pressures and its relation to changes in hemoglobin-oxygen affinity.
The stoichiometric and thermodynamic properties of nitrogen (N2) binding to human deoxyhemoglobin (Hb) at N2 saturation pressures up to 400 atm were derived from measured N2 solubilities in protein-free buffers (pH 7.1) and in corresponding buffer + Hb (6.5% w/w) solutions at 20.0, 25.0, and 37.0 degrees C. At each temperature, approximately 3 N2 molecules bind per Hb tetramer at N2 pressures of 100 atm, while about 7 N2 molecules bind per tetramer at 400 atm N2 pressure, where available binding sites are still not fully saturated. Calculated N2-Hb binding isotherms are well described by a simple binding model with 12 independent and equivalent binding sites/Hb tetramer. N2 binding at each of the sites is hydrophobic, exhibiting the typical increase of the dissociation enthalpy with temperature. Enthalpies of dissociation are slightly more negative, while corresponding unitary entropies are somewhat less negative than those for the transfer of N2 from olive oil to water. Calculated partial molar volumes of N2 in Hb are positive but less than the corresponding partial molar volumes of N2 in buffer. Results indicate that N2 binding to Hb is accompanied by only small protein conformational changes which entail slight structural destabilization and loss of free volume in the protein that partially accommodates the volume of the N2 ligand. Results are related to previously reported effects of high pressure and high-pressure N2 on HbO2 affinity, illuminating essential features of the molecular mechanisms for these effects.
F.G. Hall Laboratory for Environmental Research, Duke University Medical Center, Durham, North Carolina 27710.
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D006454:Hemoglobins / Q000378:metabolism*; D006801:Humans; D046911:Macromolecular Substances; D009584:Nitrogen / Q000378:metabolism*; D010100:Oxygen / Q000097:blood*; D011312:Pressure; D012995:Solubility; D012996:Solutions; D013696:Temperature; D013816:Thermodynamics
D006454:Hemoglobins; D046911:Macromolecular Substances; D012996:Solutions; C100003:deoxyhemoglobin; D009584:Nitrogen; D010100:Oxygen