Purification and characterization of anti-N lectin from Vicia unijuga leaves.
1. An anti-N lectin was extracted from Vicia unijuga leaves with phosphate-buffered saline (PBS). Purification of the lectin was achieved, after pretreatment of the PBS extract by ammonium sulfate fractionation and absorption with human M erythrocytes, by using a combination of conventional chromatographic techniques with asialoglycophorin AN-Sepharose CL-4B affinity chromatography. Purification steps were followed by increase of specific activity. 2. Homogeneity of the purified lectin was demonstrated by HPLC and SDS-PAGE. The purified lectin was a glycoprotein with 11.4% carbohydrate and relatively high percentages of serine, threonine and aspartic acid residues and had a Mw of 120,000 Da. 3. This lectin agglutinated human N and MN erythrocytes, but did not agglutinate M erythrocytes. Hemagglutination of the lectin was inhibited by glycophorin AN and N-active sialoglycopeptide released from human N erythrocytes by treatment with Pronase or trypsin. However, it was not inhibited by any of mono- and di-saccharides, ABH-active glycoproteins, glycophorin AM and M-active sialoglycopeptide liberated from human M erythrocytes by treatment with Pronase or trypsin.
Yanagi
K
K
Department of Biochemistry, Tokyo College of Pharmacy, Japan.
Ohyama
K
K
Yamakawa
T
T
Hashimoto
K
K
Ohkuma
S
S
D000596:Amino Acids / Q000032:analysis; D001212:Asialoglycoproteins; D002241:Carbohydrates / Q000032:analysis; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D006384:Hemagglutination; D006801:Humans; D037102:Lectins / Q000302:isolation & purification*; D008951:MNSs Blood-Group System / Q000276:immunology*; D008970:Molecular Weight; D037121:Plant Lectins
D000596:Amino Acids; D001212:Asialoglycoproteins; D002241:Carbohydrates; D037102:Lectins; D008951:MNSs Blood-Group System; D037121:Plant Lectins; C042242:Vicia lectins; C026503:asialoglycophorin