Circular dichroism spectra of human hemoglobin reveal a reversible structural transition at body temperature.

Previously we have shown that human red blood cells (RBCs) undergo a sudden change from blocking to passing through a 1.3+/-0.2-microm micropipette when applying an aspiration pressure of 2.3 kPa at a critical transition temperature (Tc = 36.4+/-0.3 degrees C). Low-shear viscosity measurements suggested that changes in the molecular properties of hemoglobin might be responsible for this effect. To evaluate structural changes in hemoglobin at the critical temperature, we have used circular dichroism (CD) spectroscopy. The thermal denaturation curves of human hemoglobin A (HbA) and hemoglobin S (HbS) upon heating between 25 and 60 degrees C were non-linear and showed accelerated denaturation between 35 and 39 degrees C with a midpoint at 37.2+/-0.6 degrees C. The transition was reversible below 39 degrees C and independent of solution pH (pH 6.8-7.8). It was also independent of the oxygenation state of hemoglobin, since a sample that was extensively deoxygenated with N2 showed a similar transition by CD. These findings suggest that a structural change in hemoglobin may enable the cellular passage phenomenon as well as the temperature-dependent decrease in viscosity of RBC solutions.