GPI-ANCHORED AND TRANSMEMBRANE-ANCHORED INFLUENZA HEMAGGLUTININ DIFFER IN STRUCTURE AND RECEPTOR-BINDING ACTIVITY

We investigated the influence of a glycosylphosphatidylinositol (GPI) anchor on the ectodomain of the influenza hemagglutinin (HA) by replac ing the wild type (wt) transmembrane and cytoplasmic domains with a GP I lipid anchor. GPI-anchored HA (GPI-HA) was transported to the cell s urface with equal efficiency and at the same rate as wt-HA. Like wt-HA , cell surface GPI-HA, and its ectodomain released with the enzyme PI- phospholipase C (PI-PLC), were 9S trimers. Compared to wt-HA, the GPI- HA ectodomain underwent additional terminal oligosaccharide modificati ons; some of these occurred near the receptor binding pocket and compl etely inhibited the ability of GPI-HA to bind erythrocytes. Growth of GPI-HA-expressing cells in the presence of the mannosidase I inhibitor deoxymannojirimycin (dMM) abrogated the differences in carbohydrate m odification and restored the ability of GPI-HA to bind erythrocytes. T he ectodomain of GPI-HA produced from cells grown in the presence or a bsence of dMM underwent characteristic low pH-induced conformational c hanges (it released its fusion peptides and became hydrophobic and pro teinase sensitive) but at 0.2 and 0.4 pH units higher than wt-HA, resp ectively. These results demonstrate that although GPI-HA forms a stabl e trimer with characteristics of the wt, its structure is altered such that its receptor binding activity is abolished. Our results show tha t transmembrane and GPI-anchored forms of the same ectodomain can exhi bit functionally important differences in structure at a great distanc e from the bilayer.