Isolation and characterization of glycophorin from nucleated (chicken) erythrocytes.
A sialoglycoprotein fraction was isolated from chicken erythrocytes by two methods based on the phenol extraction or chloroform/2-propanol extraction of differently prepared erythrocyte membranes. Both preparations gave in SDS-PAGE two major PAS-stained bands (GP2 and GP3), which migrated as 60- and 33-kDa species, respectively, compared to reference proteins, or as 44- and 23-kDa molecules, compared to human glycophorins. Some less abundant slower migrating PAS-stained components, antigenically related to GP2 and GP3, also were detected. No evidence for the presence of antigenically distinct glycoproteins of leukosialin type was obtained. Interconversion in SDS-PAGE, similar carbohydrate composition, and similar antigenic properties of GP2 and GP3 indicated that they are a dimer and monomer, respectively, of the same glycoprotein which shows properties that allow it to be classified as a glycophorin. Lectin binding studies and methylation analysis of beta-elimination products of chicken glycophorin preparation showed the presence of O-glycans and N-glycans. The major O-glycans include sialylated Galbeta1-3GalNAc units and more complex GlcNAc-containing chains. Among the N-glycans, there are complex-type biantennary structures with a bisecting GlcNAc residue, accompanied by chains with additional antennas linked to alpha-mannose residues. A characteristic feature of the chicken glycophorin is a relatively high proportion of N-glycans to O-glycans, compared to the glycophorin A from human erythrocytes.
Duk
M
M
Department of Immunochemistry, Ludwik Hirszfeld Institute of Immunology and Experimental Therapy, Polish Academy of Sciences, Rudolf Weigl Street 12, Wroclaw, 53-114, Poland.
Krotkiewski
H
H
Stasyk
T V
TV
Lutsik-Kordovsky
M
M
Syper
D
D
Lisowska
E
E
D000818:Animals; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000032:analysis; D002462:Cell Membrane / Q000378:metabolism; D002645:Chickens; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000276:immunology; D004912:Erythrocytes / Q000737:chemistry*; D006021:Glycophorins / Q000737:chemistry* / Q000276:immunology / Q000302:isolation & purification*; D006031:Glycosylation; D015151:Immunoblotting; D037102:Lectins / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012795:Sialoglycoproteins / Q000737:chemistry / Q000302:isolation & purification; D013347:Subcellular Fractions / Q000737:chemistry
D002241:Carbohydrates; D000939:Epitopes; D006021:Glycophorins; D037102:Lectins; D012795:Sialoglycoproteins