Characterization of the calcium-binding sites of calcineurin B.
Calcineurin (CaN) is a calcium- and calmodulin-dependent serine/threonine phosphatase whose inhibition by the immunosuppressant-immunophilin complexes (cyclosporin-cyclophilin and FK506-FKBP) is considered key to the mechanism of immunosuppression. CaN is a heterodimer, consisting of a 59 kDa catalytic subunit (A) and a 19 kDa calcium-binding regulatory subunit (B). The latter is postulated to harbor four calcium binding domains of the EF hand type. The titration of the CaN B apoprotein with the isomorphic Cd2+ was followed by 113Cd NMR and these data support one high-affinity metal binding site and three lower-affinity ones. Flow dialysis data with Ca2+ indicate one high affinity calcium binding site with Kd approximately 2.4 x 10(-8) M and three other sites with Kd approximately 1.5 x 10(-5) M. The chemical shifts of all four 113Cd resonances (-75, -93, -106 and -119 ppm) are in the same range as found in other 113Cd substituted calcium-binding proteins, and are indicative of all-oxygen coordination of pentagonal bipyramidal geometry.
Kakalis
L T
LT
Department of Pharmacology, Yale University School of Medicine, New Haven, CT 06520, USA.
Kennedy
M
M
Sikkink
R
R
Rusnak
F
F
Armitage
I M
IM
D000595:Amino Acid Sequence; D001665:Binding Sites; D002104:Cadmium; D019703:Calcineurin; D002118:Calcium / Q000378:metabolism*; D002135:Calcium-Binding Proteins / Q000737:chemistry* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data
D002135:Calcium-Binding Proteins; D002104:Cadmium; D019703:Calcineurin; D002118:Calcium