Kinetics of calmodulin binding to calcineurin.
Calcineurin (CaN) binds Ca(2+)-saturated calmodulin (CaM) with relatively high affinity; however, an accurate steady-state K(d) value has not been determined. In this report, we describe, using steady-state and stopped-flow fluorescence techniques, the rates of association and dissociation of Ca(2+)-saturated CaM from CaN heterodimer (CaNA/CaNB) and CaNA only. The rate of Ca(2+)/CaM association was determined to be 4.6 x 10(7) M(-1)s(-1). The rate of Ca(2+)/CaM dissociation from CaN was slower than previously reported and was approximately 0.0012 s(-1). In preparations of CaNA alone (no regulatory CaNB subunit), the dissociation rate was slowed further to 0.00026 s(-1). From these data we calculate a K(d) for binding of Ca(2+)-saturated CaM to CaN of 28 pM. This K(d) is significantly lower than previously reported estimates of approximately 1 nM and indicates that CaN is one of the highest affinity CaM-binding proteins identified to date.
Quintana
Andrea R
AR
Department of Neurobiology and Anatomy, The University of Texas Medical School, Houston, TX 77030, USA.
Wang
Dan
D
Forbes
Joanna E
JE
Waxham
M Neal
MN
D019703:Calcineurin / Q000032:analysis* / Q000737:chemistry*; D002147:Calmodulin / Q000032:analysis* / Q000737:chemistry*; D004789:Enzyme Activation; D007700:Kinetics; D011485:Protein Binding
D002147:Calmodulin; D019703:Calcineurin