Acceleration of hemoglobin C crystallization by hemoglobin S

Abstract

We previously reported that circulating hemoglobin (Hb) CC erythrocytes contain oxygenated HbC crystals with little or no HbF and that HbF inhibits in vitro crystallization of HbC. We now report that HbS accelerates in vitro crystallization of HbC. Crystals were formed in 1.8 mol/L potassium phosphate buffer, pH 7.4, at 30 degrees C and were counted in several time intervals with a hematocytometer. The hemoglobin composition of Millipore-isolated crystals and supernatant were also analyzed. Under the conditions selected, 100% HbS formed needle-shaped crystals only after two hours. Pure HbC does not form crystals within 15 minutes, whereas a ratio of 10% HbS:90% HbC forms 1,100 crystals/mm3, 20% HbS:80% HbC forms 370 crystals/mm3, and 30% HbS:70% HbC forms 5 crystals/mm3. Crystals formed in the presence of HbS are tetragonal, as are pure HbC crystals. As compared with 100% HbC, HbA or albumin mixed with HbC showed a decreased number of crystals as a result of dilution. Analysis of the Hb content of isolated crystals by citrate agar gel electrophoresis showed that HbS was rapidly incorporated into the crystal in the same ratio over time. These results demonstrate that HbS accelerates crystallization of HbC with respect to the rates of crystallization of any of these two Hbs separately, through a mechanism that involves cocrystallization. These results may be significant in understanding SC disease.