Unexpected affinity of polysaccharides and its application in separation of enzymes on fluidized beds

Abstract

Fluidized bed chromatography is emerging as a powerful tool in bioseparation of proteins/enzymes. In this work, alginate and chitosan beads have been used as affinity media in fluidized mode to selectively capture α-amylases and Aspergillus niger cellulase, respectively. Fairly high yields (>80%), with homogeneity at the level of single band on sodium dodecyl sulfate–polyacrylamide gel electrophoresis, were achieved in all these cases. The main novel features are unexpected affinity of the polysaccharides towards the target enzymes; such macroaffinity ligands avoid any costly chemical steps for affinity ligands; and possibility of easy scale up in fluidized bed mode and single step purification protocols.

Keywords:

• Alginate
• α-Amylase
• Cellulase
• Chitosan
• Expanded bed affinity chromatography
• Macroaffinity ligand

Acknowledgments

The travel support provided by Council for Scientific and Industrial Research (Govt. of India) and support for the Conference registration fees provided by Birkhauser Verlag AG, Basel, for presenting this work in SPICA 2000 are gratefully acknowledged. Protocol material related to this work will be available in Methods for Affinity Based Separations of Enzymes and Proteins (Methods in Biosciences and Medicine), ISBN 3-7643-6306-1, Birkhauser Verlag, Basel, Switzerland, due in 2002.