κ-Carrageenan as a new smart macroaffinity ligand for the purification of pullulanase

κ-Carrageenan is a polysaccharide from red seaweed which gets precipitated by K+ ions and dissolves again in water. This smart, K+-responsive polymer was found to selectively bind pullulanase activity from Bacillus acidopullulyticus. Gel filtration on Sephadex G-200 showed the formation of the polymer–pullulanase complex at the pre-precipitation stage. On the other hand, phospholipase D, an enzyme which did not co-precipitate with κ-carrageenan, did not form any complex with the polymer. Thus, K+ ions could be used to selectively precipitate the pullulanase activity. Then, 92% enzyme activity could be eluted with 1 M maltose solution. The single step protocol resulted in 50-fold purification, with a single band on sodium dodecylsulfate–polyacrylamide gel electrophoresis.