Rapid purification of calcium-activated protease by calcium-dependent hydrophobic-interaction chromatography.
Both low Ca2+- and high Ca2+-requiring forms of Ca2+-activated protease (calpains I and II) were found to bind to phenyl-Sepharose in a calcium-dependent manner, suggesting that both enzymes expose a hydrophobic surface region in the presence of Ca2+. Inclusion of leupeptin in column buffers prevented the loss of activity during hydrophobic-interaction and substrate-affinity chromatography. Under these conditions calpain II (high calcium-requiring form) was rapidly purified from bovine brain and rabbit skeletal muscle using successive phenyl-Sepharose and casein-Sepharose columns.
Gopalakrishna
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Head
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JF
D000818:Animals; D001921:Brain / Q000201:enzymology; D002118:Calcium / Q000494:pharmacology; D002154:Calpain; D002417:Cattle; D002846:Chromatography, Affinity / Q000379:methods; D002848:Chromatography, DEAE-Cellulose / Q000379:methods; D010450:Endopeptidases / Q000302:isolation & purification*; D046911:Macromolecular Substances; D008970:Molecular Weight; D009132:Muscles / Q000201:enzymology; D011817:Rabbits
D046911:Macromolecular Substances; D010450:Endopeptidases; D002154:Calpain; D002118:Calcium