Factors responsible for the Ca2+‐dependent inactivation of calcineurin in brain

• Published in: FEBS letters Vol. 374; no. 2; pp. 237 - 240
• Main Authors: Stemmer, Paul M., Wang, Xutong, Krinks, Marie H., Klee, Claude B.
• Format: Journal Article
• Language: English
• Published: 30-10-1995
• Subjects: Ca2; Calcineurin; Calmodulin; Protein phosphatase
• ISSN: 0014-5793; 1873-3468
• DOI: 10.1016/0014-5793(95)01095-V

The Ca2+‐dependent protein phosphatase activity of crude rat brain extracts measured in the presence of okadaic acid, exhibits the characteristic properties of the calmodulin‐stimulated protein phosphatase, calcineurin. It is stimulated more than 200‐fold by Ca2+ and inhibited by the calmodulin‐binding peptide, M13, and by the immunosuppressive drug, FK506. It is insensitive to rapamycin at concentrations up to 1 μM. Its specific activity, based on calcineurin concentration determined by quantitative analysis of Western blots exposed to anti‐bovine brain IgG, is ten to twenty times that of purified rat brain calcineurin assayed under similar conditions. Unlike the purified enzyme it is rapidly and irreversibly inactivated in a time‐, temperature‐, and Ca2+/calmodulin‐dependent fashion without evidence of extensive proteolytic degradation. The enzyme is converted to a state which does not lose activity by removal of low molecular weight material by gel filtration. Reconstitution of a labile enzyme is achieved by the addition of the low molecular weight‐containing fraction eluted from the gel filtration column. These observations indicate that calcineurin in crude brain extracts is under the control of Ca2+/calmodulin‐dependent positive and negative regulatory mechanisms which involve unidentified endogenous factor(s).