Characterization of erythrocyte membrane-associated enzymes (glyceraldehyde-3-phosphate dehydrogenase and phosphoglyceric kinase).
The purpose of our study was to determine why several of the glycolytic enzymes of the erythrocyte have the propensity for adhering to erythrocyte membranes while others do not. Two of these membrane-associated enzymes, glyceraldehyde phosphate dehydrogenase (GAPD) and phosphoglyceric kinase (PGK), have been shown to have controlling functions on intra-erythrocytic glycolysis and sodium-potassium transport. In order to test the hypothesis that the membrane-associated fraction of these enzymes consisted of isozymes with increased capacity to become membrane associated, the enzymes from cytosol or membrane sources were partially purified and characterized. Determination of molecular weight by gel filtration chromatography, measurement of certain kinetic parameters, and the curves relating to pH optimal activity indicated that there were no measurable differences between membrane and cytosol PGK and membrane and cytosol GAPD. It was possible to raise inhibitory antibodies to GAPD in certain species of mice, and these antibodies did not distinguish between GAPD isolated from either membrane or cytosol sources. GAPD was firmly bound to membranes and the membrane-associated fraction counted for approximately 60 per cent of total erythrocytic enzyme. Membrane-associated PGK was only loosely adherent, and accounted for only 1 per cent of the total erythrocytic enzyme. The reasons for membrane association have yet to be determined, but these inward facing membrane-associated enzymes appear to function by carrying the organization of the membrane into the cell interior.
D000818:Animals; D000906:Antibodies; D002462:Cell Membrane / Q000201:enzymology* / Q000276:immunology; D002850:Chromatography, Gel; D003600:Cytosol / Q000201:enzymology; D004912:Erythrocytes / Q000201:enzymology*; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000097:blood* / Q000276:immunology / Q000302:isolation & purification; D006461:Hemolysis; D006863:Hydrogen-Ion Concentration; D007527:Isoenzymes / Q000097:blood*; D007700:Kinetics; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D008815:Mice, Inbred Strains; D008970:Molecular Weight; D010735:Phosphoglycerate Kinase / Q000097:blood* / Q000302:isolation & purification
D000906:Antibodies; D007527:Isoenzymes; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010735:Phosphoglycerate Kinase