Proteomics of Stored Red Blood Cell Membrane and Storage-Induced Microvesicles Reveals the Association of Flotillin-2 With Band 3 Complexes
Abstract
The storage of erythrocyte concentrates (ECs) induces lesions that notably affect metabolism, protein activity, deformability of red blood cells (RBCs), as well as the release of oxygen. Band 3 is one of the proteins affected during the ex vivo aging of RBCs. This membrane protein is an anion transporter, an anchor site for the cytoskeleton and other membrane proteins as well as a binding site for glycolytic enzymes and bears blood group antigens. In the present study, band 3 complexes were isolated from RBCs stored for 7 and 42 days in average (n = 3), as well as from microvesicles (n = 3). After extraction of membrane proteins with a deoxycholate containing buffer, band 3 complexes were co-immunoprecipitated on magnetic beads coated with two anti-band 3 antibodies. Both total membrane protein extracts and eluates (containing band 3 complexes) were separated on SDS-PAGE and analyzed by bottom-up proteomics. It revealed that three proteins were present or absent in band 3 complexes stemming from long-stored or short-stored ECs, respectively, whereas the membrane protein contents remained equivalent. These potential markers for storage-induced RBC aging are adenylosuccinate lyase (ADSL), α-adducin and flotillin-2, and were furth...Continue Reading
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