Unexpected different binding of mistletoe lectins from plant extracts to immobilized lactose and N-acetylgalactosamine.
Mistletoe Extracts (ME) are of growing interest to pharmacological research because of their apoptosis-inducing/cytostatic and immunomodulatory effects. The standardization of the three different groups of Mistletoe Isolectins (ML-I, II and III) is often rendered more difficult since the primary structures are nearly identical. Their classification is based on their Galactose- and N-acetyl-D-galactosamine (GalNAc)-specificity which was measured by various inhibitory assays. The aim of the present study was to improve the characterization of the direct binding activity of the isolectins from ME to immobilized lactose, GalNAc and to the oligosaccharide asialofetuin. After careful ultrafiltration of fresh ME, affinity chromatography was carried out using lactose- agarose, GalNAc--agarose and asialofetuin--affigel 15 columns. MLs were further purified by Sephadex G-100 or by cation exchange chromatography which was adapted to a Fast Protein Liquid Chromatography (FPLC) system. Proteins from both fresh plants and commercial ME were able to bind immobilized lactose to a considerable extent. The majority of this lectin has a B-chain with a Molecular Weight (MW) of 34 kD and an A-chain with a MW of 29 kD (ML-I). Only a minor part of the lactose-binding proteins has a lower MW, namely 32 kD and 27 kD (MLII). However, neither MLs which were eluted from lactose columns, nor the proteins from fresh plant or ME showed a direct binding to the immobilized GalNAc. In spite of this deficiency, GalNAc was able to induce a considerable (25% and 32%) inhibitory effect on their binding to immobilized asialofetuin indicating a discrepancy between the lectin binding and inhibiting effects of GalNAC. Consequently, for an improved standardization of ME more specific sugar molecules are necessary.
Hajtò
Tibor
T
Department of Immunology and Biotechnology, Faculty of Medicine, University of Pècs, Hungary. drhajtot@axelero.hu
Krisztina
Fodor
F
Ildikò
Aponyi
A
Zsolt
Pallai
P
Pèter
Balogh
B
Pèter
Németh
N
Pàl
Perjési
P