Thermodynamic mechanism of free heme action on sickle cell hemoglobin polymerization

For insights into the mechanisms of heme action on the rate of sickle cell hemoglobin polymerization, we determine the erythrocytic concentration of free heme using a novel method based on enzymatic catalysis and luminescence. We find in sickle cell patients 44 ± 10 µM, in sickle trait individuals, 33 ± 4 µM, and in healthy adults, 21 ± 2 µM. We test the applicability of two mechanisms of heme action: a kinetic one, whereby heme aggregates serve as heterogeneous nucleation centers, and a thermodynamic pathway, in which free heme enhances the attraction between sickle hemoglobin (HbS) molecules in solution. We show that the latter mechanism exclusively operates. The enhanced attraction leads to increase of the total volume of a population of dense liquid clusters by about two orders of magnitude. As the dense liquid clusters serve as locations and precursors to the formation of the HbS polymer nuclei, their increased volume directly leads to faster polymer nucleation. © 2015 American Institute of Chemical Engineers AIChE J, 61: 2861–2870, 2015