Journal
PROTEIN EXPRESSION AND PURIFICATION
Volume 28, Issue 1, Pages 190-195Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/S1046-5928(02)00673-3
Keywords
alginates; smart polymers; glucoamylase; pullulanase; three-phase partitioning (TPP); macroaffinity ligand-facilitated three-phase; partitioning (MLFTPP)
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Starch-degrading enzymes glucoamylase (from Aspergillus niger), and pullulanase (from Bacillus acidopullulyticus) were purified using alginates (polysaccharides consisting of mannuronic acids and guluronic acids) by a recently developed technique called macroaffinity ligand-facilitated three-phase partitioning (MLFTPP). In this process, a crude preparation of the enzyme was mixed with alginate. On addition of appropriate amounts of ammonium sulfate and t-butanol, the alginate bound enzyme appeared as an interfacial precipitate between the lower aqueous and the upper t-butanol phase. Enzyme activity from this interfacial precipitate was recovered using I M maltose. Glucoamylase and pullulanase were purified 20- and 38-fold with 83% and 89% activity recovery, respectively. Both the purified preparations showed a single band on SDS-PAGE. (C) 2002 Elsevier Science (USA). All rights reserved.
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