Single-Step Purification of Calpain-1, Calpain-2, and Calpastatin Using Anion-Exchange Chromatography.
Purification and separation of calpains and calpastatin are used to determine the individual activities of calpain-1 and calpain-2 and their inhibitor calpastatin. We discuss here a method to purify these enzymes using dialysis followed by separation using anion-exchange chromatography coupled with gradient elution. Swollen DEAE Sephacel is used as the column matrix in this method. Calpastatin and both domains of calpain are weakly basic molecules that effectively bind with the DEAE Sephacel and separate well using a stepwise, increasing gradient of NaCl to elute the proteins. Calpastatin binds most weakly with the column matrix, so it elutes first, followed by calpain-1 and, finally, calpain-2.
['Biswas|Ashim Kumar|AK|', 'Tandon|Samarth|S|']
D000818:Animals D000838:Anions D002135:Calcium-Binding Proteins D002154:Calpain D002645:Chickens D002852:Chromatography, Ion Exchange D003637:DEAE-Dextran D008967:Molecular Biology
Purification Calpastatin Anion-exchange chromatography Calpain DEAE Sephacel Chicken Muscle tissues