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Biochemistry. 2012 Apr 17;51(15):3264-72. doi: 10.1021/bi201623v. Epub 2012 Apr 6.

Interaction of deoxyhemoglobin with the cytoplasmic domain of murine erythrocyte band 3.

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Department of Chemistry, Purdue University, West Lafayette, Indiana 47907, United States.


The partial pressure of oxygen constitutes an important factor in the regulation of human erythrocyte physiology, including control of cell volume, membrane structure, and glucose metabolism. Because band 3 is thought to be involved in all three processes and because binding of hemoglobin (Hb) to the cytoplasmic domain of band 3 (cdb3) is strongly oxygen-dependent, the possibility that the reversible association of deoxyhemoglobin (deoxyHb) with cdb3 might constitute an O(2)-dependent sensor that mediates O(2)-regulated changes in erythrocyte properties arises. While several lines of evidence support this hypothesis, a major opposing argument lies in the fact that the deoxyHb binding sequence on human cdb3 is not conserved. Moreover, no effect of O(2) pressure on Hb-band 3 interactions has ever been demonstrated in another species. To explore whether band 3-Hb interactions might be widely involved in O(2)-dependent regulation of erythrocyte physiology, we undertook characterization of the effect of O(2) on band 3-Hb interactions in the mouse. We report here that murine band 3 binds deoxyHb with significantly greater affinity than oxyHb, despite the lack of significant homology within the deoxyHb binding sequence. We further map the deoxyHb binding site on murine band 3 and show that deletion of the site eliminates deoxyHb binding. Finally, we identify mutations in murine cdb3 that either enhance or eliminate its affinity for murine deoxyHb. These data demonstrate that despite a lack of homology in the sequences of both murine band 3 and murine Hb, a strong oxygen-dependent association of the two proteins has been conserved.

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