Spin Label Studies of the Hemoglobin–Membrane Interaction During Sickle Hemoglobin Polymerization

Abstract

An enhanced hemoglobin–membrane association has been previously documented in sickle cell anemia. However, it is not known how this interaction is modified during the hemoglobin S polymerization process. In this work, we use a model of reconstituted erythrocytes from ghost membranes whose cytoskeleton proteins had been previously labeled with the 4-maleimido Tempo spin label, and that were subsequently resealed with hemoglobin S or A solutions. Using electron paramagnetic resonance spectroscopy, we studied the time dependence of the spectral W/S parameter, indicative of the conformational state of cytoskeleton proteins (mainly spectrin) under spontaneous deoxygenation, with the aim of detecting the eventual effects due to hemoglobin S polymerization. The differences observed in the temporal behavior of W/S in erythrocytes reconstituted with both hemoglobins were considered as experimental evidence of an increment in hemoglobin S–membrane interaction as a result of the polymerization process of hemoglobin S under spontaneous deoxygenation.