The reconstitution of the human erythrocyte sugar transporter in planar bilayer membranes.
The degradation of human erythrocyte membrane proteins in relation to the identification of the monosaccharide transporter has been investigated in whole membrane preparations and membrane protein extracts by polyacrylamide gel electrophoresis in sodium n-dodecyl sulphate and iodine-125 labelling. Evidence is presented for the degradation of band 3 polypeptide to lower molecular weight material some of which appears in region 4.5 of the polyacrylamide gel electrophoresis profile. It is found that the degradation process is inhibited by phenylmethylsulphonyl fluoride and is only significant in membrane extracts in the absence of detergent (Triton X-100) and on prolonged incubation at 37 degrees C, conditions which do not prevail during the isolation of membrane protein extracts for reconstitution studies. Extracts of band 3 and band 4.5 have been prepared and reconstituted in bilayer lipid membranes. The permeabilities of the reconstituted systems to D-glucose have been investigated and it is found that only bilayers incorporating band 4.5 exhibited enhanced monosaccharide transport. A linear relationship between D-glucose transport and the concentration of protein in the aqueous phase bathing the bilayers suggests a partitioning of the protein into the bilayer. Reconstitution is stereospecific and inhibited by cytochalasin B.
D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins / Q000378:metabolism; D002352:Carrier Proteins / Q000378:metabolism*; D004910:Erythrocyte Membrane / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism*; D006801:Humans; D007457:Iodine Radioisotopes; D008051:Lipid Bilayers; D008565:Membrane Proteins / Q000097:blood / Q000302:isolation & purification; D009004:Monosaccharide Transport Proteins; D011865:Radioisotope Dilution Technique
D001457:Anion Exchange Protein 1, Erythrocyte; D001798:Blood Proteins; D002352:Carrier Proteins; D007457:Iodine Radioisotopes; D008051:Lipid Bilayers; D008565:Membrane Proteins; D009004:Monosaccharide Transport Proteins