Screening of lectins from South American plants used as affinity ligands to purify rhEPO

Abstract

Two groups of isoforms of rhEPO, at a concentration of 300 µg/ml, were tested as putative inhibitors of the lectinic hemagglutination reaction in order to obtain affinity ligand(s) for hormone purification: groups I (pI: 3.80; 3.89; 3.95; 4.07, 4.15 and 4.26) and groups II (pI: 4.15, 4.26; 4.38; 4.51; 4.72 and 4.93) Crude extracts from the vegetable materials Abrus precatorious (Abrin), Artocarpus incisa (Frutalin), Artocarpus integrifolia (Jacalin), Canavalia ensiformes (ConA), Canavalia brasiliensis (Conbr), Cratylia floribunda, Dioclea altissima (DAL), Dioclea grandiflora (DGL), Erythrina vellutina (EVL), Erythrina cristagalli, Lutaelburgia auriculata (lectin not fully characterized yet), Lycopersicum esculentum (LEA), Phaseolus vulgaris (PHA), Ricinus communis (Ricin) and Triticum vulgaris (WGA) were used. Only some of the galactose-specific lectins and the GlcNAc-specific lectins showed rapid full inhibition of the hemagglutination reaction for the less acidic isoforms and the total isoforms of rhEPO, respectively. On this basis, the selected lectins were purified by affinity chromatoghraphy and covalently coupled to cyanogen bromide activated Sepharose® (Amersham-Pharmacia). CHO.K1 cell culture supernatant containing rhEPO was loaded onto the lectin resins and the recoveries were calculated by using specific elutions

Detection of isoforms of recombinant human erythropoietin by various plant lectins after isoelectric focusing

“…However, only a weak interaction between ConA and rHuEPO was observed. Comparisons with previous studies confirmed these observations [22][23][24]. Interestingly, the three isoforms detected by ConA did not correspond to the three most intense bands of the immunoblot.…”

Comparison of affinity membrane adsorbers for the purification of recombinant human erythropoietin (rhEPO)

“…Affinity ligands that have been used are Cibacron Blue (CB) 3 , anti-EPO monoclonal antibodies 4,5 and lectins, such as concanavalin A (ConA), 6 wheat germ agglutinin (WGA), 7,8 potato and tomato lectins. 8 However, chromatographic processes are traditionally carried out in packed beds, which may present limitations, especially at larger scales. Traditional preparative agarose-based column chromatography may suffer from resin deformation and bed consolidation over time, and this may lead to the formation of preferential channels and to an increase in pressure drop along the column.…”

Cashew Tree Bark Secretion - Persectives for its Use in Protein Isolation Strategies

“…Synthesis generally occurs in all organs of the plant, with different quantitative composition; appearing to be genetically controlled and little influenced by environmental conditions. The gum is similar to gum arabic and may be used as to a substitute for liquid glue for paper, in the pharmaceutical/cosmetic industry and as an agglutinant for capsules and pills [1,2,7]. Fifteen species of trees or perennial bushes compose the genus Anacardium, native of tropical parts of South America and Central and of western India [8][9][10].…”

“…The cashew tree is a hardy, fast-growing, evergreen with an umbrella-like canopy. The tree has a rather messy look with its gnarled stem and crooked branches [7]. They are easily cultivated, vigorous, and drought-resistant, requiring little care.…”

“…The presence of phenolic resine [13,14] is still a nuisance for harvesting and shelling. Although the fruit can be used for making many typical fruit products (jellies, jams, juice, wine and liquor), the apple is often discarded, in pursuit of the nut [7,9].…”