Expression and reconstitution of calcineurin A and B subunits.
Calcineurin consists of two subunits, a catalytic A subunit of 60 kDa and a regulatory B subunit of 19 kDa. Both the A and B subunits of rat brain calcineurin were expressed in E. coli. The B-subunit was readily overexpressed in the pET-21a vector with yields of > 70 mg of purified B subunit per 1 culture, representing > 17% of the soluble E. coli protein. About 8 mg of purified A subunit was obtained. The enzyme activities of the A-subunit and the reconstituted AB complex were found to be comparable to that of the bovine brain enzyme. The reconstitution of the AB complex was studied and shown to be rapid.
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Department of Biochemistry and Molecular Biology, University of Miami School of Medicine, FL 33101, USA.
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D001483:Base Sequence; D001921:Brain; D019703:Calcineurin; D002135:Calcium-Binding Proteins / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D003001:Cloning, Molecular; D004926:Escherichia coli; D007527:Isoenzymes / Q000302:isolation & purification; D017510:Protein Folding; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification
D002135:Calcium-Binding Proteins; D007527:Isoenzymes; D011994:Recombinant Proteins; D019703:Calcineurin