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The Catalytically Active Domain in the A Subunit of Calcineurin

2003; De Gruyter; Volume: 384; Issue: 10-11 Linguagem: Inglês

10.1515/bc.2003.158

1437-4315

Bosong Xiang, Ping Liu, Guangde Jiang, Kun Zou, Fei Yi, Shujie Yang, Qun Wei,

Biochemical and Structural Characterization

Calcineurin (CaN) is a heterodimer composed of a catalytic subunit A (CaNA) and a regulatory subunit B (CaNB). We report here an active truncated mutation of the rat CaNAδ that contains only the catalytic domain (residues 1-347, also known as a/CaNA). The pnitrophenyl phosphatase activity and protein phosphatase activity of a/CaNA were higher than that of CaNA. Both pnitrophenyl phosphatase activity and protein phosphatase activity of a/CaNA were unaffected by CaM and the B-subunit; the B-subunit and CaM have relatively little effect on pnitrophenyl phosphatase activity and a crucial effect on protein phosphatase activity of CaNA. Mn[2+] and Ni[2+] ions effeciently activated CaNA. The Km of a/CaNA was about 16 mM, and the kcat of a/CaNA was 10.03 1/s using pNPP as substrate. With RII peptide as a substrate, the Km of a/CaNA was about 21 uM and the kcat of a/CaNA was 0.51 1/s. The optimum reaction temperature was about 45C, and the optimum reaction pH was about 7.2. Our results indicate that a/CaNA is the catalytic core of CaNA, and CaN and the Bsubunit binding domain itself might play roles in the negative regulation of the phosphatase activity of CaN. The results provide the basis for future studies on the catalytic domain of CaN.