Purification and characterization of four isoforms of Himalayan mistletoe ribosome-inactivating protein from Viscum album having unique sugar affinity.
Ribosome-inactivating proteins having antitumor and immunomodulatory properties constitute the active principle of widely used mistletoe therapy in Europe. This is the first report of the four isoforms of Himalayan mistletoe ribosome-inactivating proteins (HmRips) from Viscum album parasitized on wild apple inhabiting NW Himalayas. HmRips were purified by affinity chromatography and four isoforms were separated by ion-exchange chromatography. HmRip 1, 2, 3, and 4 have isoelectric points of 6.6, 6.1, 5.2, and 4.7, respectively. Disulfide linked toxin and lectin subunits of HmRip 1 and 2 isoforms have molecular weights of 28 and 34kDa while those of HmRip 3 and 4 have 28 and 32kDa. The isoforms lacked blood group specificity and showed positive activity with seven mammalian erythrocyte types but did not show any activity with avian erythrocyte type. Lectin activity of HmRips remained unchanged for a wide range of temperatures (0-65 degrees C) and pH (3-9). Unlike other type II Rips, the HmRip 1, 2, and 4 showed unique affinity towards l-rhamnose, meso-inositol, and l-arabinose while HmRip 3 has specificity to gal/galNAc. Sugar binding studies with 22 sugars also suggested that the C-4 hydroxyl of galactose might be the critical site involved in sugar binding of HmRips. Type II Rips are known to be galactoside specific and do not have affinity for l-rhamnose and meso-inositol. However, HmRip 1, 2, and 4 having equal affinity for galactose and l-rhamnose do not strictly fit into any of the four structural classes of the lectins and represent a new class of type II Rips and plant lectins.
Department of Botay, University of Delhi, Delhi-110 007, India.
Vandana
V
Mishra
Radhey S
RS
Sharma
Savita
S
Yadav
Cherukuri R
CR
Babu
Tej P
TP
Singh
D000595:Amino Acid Sequence; D000818:Animals; D050260:Carbohydrate Metabolism; D002241:Carbohydrates / Q000737:chemistry; D002846:Chromatography, Affinity; D002852:Chromatography, Ion Exchange; D015180:Electrophoresis, Gel, Two-Dimensional; D004912:Erythrocytes / Q000276:immunology; D006384:Hemagglutination / Q000187:drug effects; D006386:Hemagglutination Tests / Q000379:methods; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D014784:Mistletoe / Q000737:chemistry*; D008969:Molecular Sequence Data; D037121:Plant Lectins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism / Q000494:pharmacology; D010940:Plant Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism* / Q000494:pharmacology; D020033:Protein Isoforms; D017386:Sequence Homology, Amino Acid; D013696:Temperature
D002241:Carbohydrates; D037121:Plant Lectins; D010940:Plant Proteins; D020033:Protein Isoforms