Nitrogen binding to deoxyhemoglobin at high pressures and its relation to changes in hemoglobin-oxygen affinity

Area of Science:

• Biophysics
• Physiological Chemistry

Background:

• Human hemoglobin (Hb) is crucial for oxygen transport.
• Understanding ligand interactions with Hb is vital for physiological studies.
• Nitrogen (N2) binding to Hb under high pressure remains incompletely understood.

Purpose of the Study:

• To determine the stoichiometric and thermodynamic properties of N2 binding to human deoxyhemoglobin (Hb).
• To investigate N2 binding at high N2 saturation pressures (up to 400 atm).
• To elucidate the molecular mechanisms of N2 effects on Hb.

Main Methods:

• Measured N2 solubilities in protein-free buffers and Hb solutions at varying temperatures (20.0, 25.0, 37.0 °C).
• Analyzed N2 binding isotherms using a simple binding model.
• Calculated partial molar volumes of N2 in Hb and buffer solutions.

Main Results:

• Approximately 3 N2 molecules bind per Hb tetramer at 100 atm, increasing to about 7 at 400 atm.
• Binding data fit a model with 12 independent, equivalent binding sites per Hb tetramer.
• N2 binding is hydrophobic, with dissociation enthalpies increasing with temperature and slight protein conformational changes.

Conclusions:

• N2 binding to Hb involves hydrophobic interactions and minimal protein conformational changes.
• These findings provide insights into the molecular mechanisms of high-pressure N2 effects on HbO2 affinity.
• The study quantifies N2 binding stoichiometry and thermodynamics to Hb under high pressure.