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Calpain pp 3-11 | Cite as

Single-Step Purification of Calpain-1, Calpain-2, and Calpastatin Using Anion-Exchange Chromatography

  • Ashim Kumar Biswas
  • Samarth Tandon
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1915)

Abstract

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.

Key words

Purification Chicken Muscle tissues Calpain Calpastatin DEAE Sephacel Anion-exchange chromatography 

Notes

Acknowledgment

This work is supported by DST (Govt. of India) grant SB/FT/LS-283/2012 and the Indian Council of Agricultural Research, New Delhi (Ministry of Agriculture).

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Ashim Kumar Biswas
    • 1
  • Samarth Tandon
    • 1
  1. 1.Division of Post-Harvest TechnologyICAR-Central Avian Research InstituteBareillyIndia

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