Stoichiometry of wheat germ agglutinin as a morphology controlling agent and as a morphology controlling agent and as a morphology protective agent for the human erythrocyte

The lectin wheat germ agglutinin (WGA) is an unusually effective agent in controlling both the forward and reverse reactions of the reversible morphology conversion discocyte in equilibrium with echinocyte for the human erythrocyte. Under conditions severe enough to drive the reactions to completion in either direction without the lectin, WGA is able to stabilize both these morphologies and to fully prevent conversion of either morphology. The lectin can quantitatively block both reactions. The ability of WGA to carry out these functions has no obvious rate limitation. Its effectiveness depends mainly on its binding stoichiometry, particularly toward the transmembrane glycoprotein, glycophorin. The critical binding stoichiometries for both the lectin and the echinocytic agent were determined in relation to the binding isotherms using 125I-labeled WGA and 35S-labeled dodecyl sulfate. There appear to be two principal stoichiometries for WGA binding that are important in its control of erythrocyte morphology. The first stoichiometry marks the threshold of obvious protection of the discocyte against strong echinocytic agents such as detergents and, likely, is simply a 1:1 stoichiometry of WGA: glycophorin, assuming currently recognized values of 3--5 x 10(5) copies of glycophorin per cell. The second important stoichiometry, whereby the cell's morphology is protected against extremely severe stress, involves binding of approximately 4--5 WGA molecules per glycophorin. The controls that WGA exerts can be instantly abolished by added N-acetylglucosamine. However, N-acetylglucosamine ligands on the erythrocyte are of less importance than membrane neuraminic acid residues in enabling WGA to control the cell's morphology, as is shown by comparing intact cells with completely desialated cells. WGA can also be used to produce elliptocytes in vitro, but it does this at levels approaching monolayer coverage of the cell with WGA.