Affinity purification, physicochemical and immunological characterization of a galactose-specific lectin from the seeds of Dolichos lablab (Indian lablab beans)

Affinity Chromatography of Lectins

“…However, the link formed is unstable in alkaline solutions (Lis & Sharon, 1981;Pepper, 1992), so acidic elution of bound lectins is advised. The method has recently been used for coupling sugars to Sepharose in order to isolate lectins from various sources (Almanza et al, 2004;Lavanya Latha et al, 2006;Pohleven et al, 2011).…”

“…Moreover, different lectins can sometimes be co-isolated from the extract, especially when the ligand is immobilized to polysaccharide matrices, which also bind lectins. In that case, serial carbohydrate affinity chromatography can be applied using two or more adsorbents in series (Chen et al, 1999;Kato et al, 2011;Lavanya Latha et al, 2006;Moreira et al, 1998;Ooi et al, 2002;Pohleven et al, 2011;Trindade et al, 2006). For example, the extract is applied to the affinity column that binds, besides the lectin of interest, non-desired lectins.…”

“…Recently, isolation of lectins from various sources has been described using numerous sugars, such as lactose (Almanza et al, 2004;Braga et al, 2006;Fujii et al, 2011;Hamako et al, 2007;Kawsar et al, 2009;Mendonça-Franqueiro et al, 2011;Naeem et al, 2007b;Pohleven et al, 2009;Rocha et al, 2009;Silva et al, 2007), sucrose and glucose (Pohleven et al, 2011), galactose (Konami et al, 1991;Lavanya Latha et al, 2006;Mo et al, 2000;Moreira et al, 1998;Naeem et al, 2007b;Nagata, 2005), mannose (Andon et al, 2003;Ooi et al, 2010;Suseelan et al, 2002), fucose (Cammarata et al, 2007;Mansour & Abdul-Salam, 2009), L-rhamnose (Jimbo et al, 2007;Watanabe et al, 2008), and sugars with 2'-acetamido groups N-acetylgalactosamine (Gerlach et al, 2005;Perçin et al, 2009;Qureshi et al, 2006), N-acetylglucosamine (Kaur et al, 2005;Kim et al, 2006;Maheswari et al, 2002;Wang & Ng, 2003), and N,N'-diacetylchitobiose (Konami et al, 1991;Koyama et al, 2002).…”

“…The non-bound lectin of interest is washed off the column and thus purified. Several authors have purified lectins from a variety of sources using serial carbohydrate affinity chromatography (Chen et al, 1999;Kato et al, 2011;Lavanya Latha et al, 2006;Moreira et al, 1998;Ooi et al, 2002;Pohleven et al, 2011;Trindade et al, 2006) or Ultrogel-A4 agarose beads to remove non-specific agarose-binding proteins from the extract prior to subjecting it to carbohydrate affinity chromatography (Kvennefors et al, 2008). In contrast, nonpolysaccharide matrices, such as silica carriers (Synsorb) (Kaur et al, 2006;Mo et al, 2000;Naeem et al, 2007b;Shangary et al, 1995), poly(2-hydroxyethyl methacrylate) matrix of Spheron 300 (Nahálková et al, 2001), fractogel affinity matrix (Guzmán-Partida et al, 2004) or polyacrylamide beads (Dresch et al, 2008;Hořejší & Kocourek, 1978;Veau et al, 1999), can be used to avoid non-specific binding of lectins to the polysaccharide matrix.…”

“…The most widely used have been differently activated Sepharoses -divinyl sulphone (Almanza et al, 2004;Lavanya Latha et al, 2006;Pohleven et al, 2011) and epoxy (epichlorohydrin) activated Sepharose have been used to immobilize carbohydrates (Chen et al, 1999;Kaur et al, 2005;Maheswari et al, 2002;Mora et al, 2008;Nagata, 2005), and cyanogen bromide activated Sepharose for glycoproteins (Bhat et al, 2010;Bhowal et al, 2005;Kajiya et al, 2003;Mo et al, 1999;Naganuma et al, 2006;Nagre et al, 2010;Vega & Pérez, 2006;Yang et al, 2007). Nhydroxysuccinimide Sepharose was used for coupling with sugar derivatives containing amino groups (Takeuchi et al, 2011), and thiopropyl Sepharose for immobilizing iodoacetamidyl glycan derivatives (Watanabe et al, 2007).…”