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Blood, 1 November 2008, Vol. 112, No. 9, pp. 3900-3906. Prepublished online as a Blood First Edition Paper on August 12, 2008; DOI 10.1182/blood-2008-03-146159.
RED CELLS Characterization of glycolytic enzyme interactions with murine erythrocyte membranes in wild-type and membrane protein knockout mice1 Department of Chemistry, Purdue University, West Lafayette, IN; 2 Department of Pediatrics and Children's Research Institute, Medical College of Wisconsin, and Blood Research Institute, Blood Center of Wisconsin, Milwaukee; 3 The Jackson Laboratory, Bar Harbor, ME; 4 New York Blood Center, New York, NY; and 5 Puget Sound Blood Center, University of Washington School of Medicine, Seattle
Previous research has shown that glycolytic enzymes (GEs) exist as multienzyme complexes on the inner surface of human erythrocyte membranes. Because GE binding sites have been mapped to sequences on the membrane protein, band 3, that are not conserved in other mammalian homologs, the question arose whether GEs can organize into complexes on other mammalian erythrocyte membranes. To address this, murine erythrocytes were stained with antibodies to glyceraldehyde-3-phosphate dehydrogenase, aldolase, phosphofructokinase, lactate dehydrogenase, and pyruvate kinase and analyzed by confocal microscopy. GEs were found to localize to the membrane in oxygenated erythrocytes but redistributed to the cytoplasm upon deoxygenation, as seen in human erythrocytes. To identify membrane proteins involved in GE assembly, erythrocytes from mice lacking each of the major erythrocyte membrane proteins were examined for GE localization. GEs from band 3 knockout mice were not membrane associated but distributed throughout the cytoplasm, regardless of erythrocyte oxygenation state. In contrast, erythrocytes from mice lacking
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| Copyright © 2008 by American Society of Hematology Online ISSN: 1528-0020 | |||||||||
-spectrin, ankyrin, protein 4.2, protein 4.1, β-adducin, or dematin headpiece exhibited GEs bound to the membrane. These data suggest that oxygenation-dependent assembly of GEs on the membrane could be a general phenomenon of mammalian erythrocytes and that stability of these interactions depends primarily on band 3. 
