Purification of a bifunctional amylase/protease inhibitor from ragi (Eleusine coracana) by chromatography and its use as an affinity ligand.
An ammonium sulphate fraction (20-60%) of bifunctional amylase/protease inhibitor from ragi (Eleusine coracana) was purified by affinity chromatography to give 6.59-fold purity with 81.48% yield. The same ammonium sulphate fraction was also subjected to ion exchange chromatography and was purified 4.28-fold with 75.95% yield. The ion exchange fraction was subjected to gel filtration and the inhibitor was purified to 6.67-fold with 67.36% yield. Further sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) was performed to check the homogeneity of purified amylase/trypsin inhibitor obtained through affinity, ion exchange and gel chromatography. The molecular weight of the inhibitor was found to be 14 kDa. This purified inhibitor was used as affinity ligand for the purification of a commercial preparation of pancreatic amylase.
Saxena
Lalit
L
Food Engineering and Technology Department, Institute of Chemical Technology, University of Mumbai, Nathalal Parikh Marg, Matunga, Mumbai-400 019, India.
Iyer
Bharti K
BK
Ananthanarayan
Laxmi
L
D000681:Amylases / Q000037:antagonists & inhibitors*; D002846:Chromatography, Affinity / Q000379:methods*; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D031748:Eleusine / Q000737:chemistry*; D004791:Enzyme Inhibitors / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D055767:Immobilized Proteins / Q000737:chemistry / Q000378:metabolism*; D055292:Pancreatic alpha-Amylases / Q000302:isolation & purification / Q000378:metabolism; D011480:Protease Inhibitors / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism
D004791:Enzyme Inhibitors; D055767:Immobilized Proteins; D011480:Protease Inhibitors; D000681:Amylases; D055292:Pancreatic alpha-Amylases