Mechanism of Calmodulin Recognition of the Binding Domain of Isoform 1b of the Plasma Membrane Ca2+-ATPase: Kinetic Pathway and Effects of Methionine Oxidation
- 8 March 2007
- journal article
- research article
- Published by American Chemical Society (ACS) in Biochemistry
- Vol. 46 (13) , 4045-4054
- https://doi.org/10.1021/bi602481u
Abstract
Calmodulin (CaM) binds to a domain near the C-terminus of the plasma membrane Ca2+-ATPase (PMCA), causing the release of this domain and relief of its autoinhibitory function. We investigated the kinetics of dissociation and binding of Ca2+-CaM with a 28-residue peptide [C28W(1b)] corresponding to the CaM-binding domain of isoform 1b of PMCA. CaM was labeled with a fluorescent probe on either the N-terminal domain at residue 34 or the C-terminal domain at residue 110. Formation of complexes of CaM with C28W(1b) results in a decrease in the fluorescence yield of the fluorophore, allowing the kinetics of dissociation or binding to be detected. Using a maximum entropy method, we determined the minimum number and magnitudes of rate constants required to fit the data. Comparison of the fluorescence changes for CaM labeled on the C-terminal or N-terminal domain suggests sequential and ordered binding of the C-terminal and N-terminal domains of CaM with C28W(1b). For dissociation of C28W(1b) from CaM labeled on the N-terminal domain, we observed three time constants, indicating the presence of two intermediate states in the dissociation pathway. However, for CaM labeled on the C-terminal domain, we observed only two time constants, suggesting that the fluorescence label on the C-terminal domain was not sensitive to one of the kinetic steps. The results were modeled by a kinetic mechanism in which an initial complex forms upon binding of the C-terminal domain of CaM to C28W(1b), followed by binding of the N-terminal domain, and then formation of a tight binding complex. Oxidation of methionine residues in CaM resulted in significant perturbations to the binding kinetics. The rate of formation of a tight binding complex was reduced, consistent with the poorer effectiveness of oxidized CaM in activating the Ca2+ pump.Keywords
This publication has 39 references indexed in Scilit:
- Kinetic Analysis of the Calmodulin-Binding Region of the Plasma Membrane Calcium Pump Isoform 4bBiochemistry, 2005
- The kinetics of calcium binding to calmodulin: Quin 2 and ANS stopped-flow fluorescence studiesPublished by Elsevier ,2004
- Dissection of the Pathway of Molecular Recognition by CalmodulinBiochemistry, 2002
- Progressive Decline in the Ability of Calmodulin Isolated from Aged Brain To Activate the Plasma Membrane Ca-ATPaseBiochemistry, 1998
- Kinetic Control of the Dissociation Pathway of Calmodulin-Peptide ComplexesPublished by Elsevier ,1997
- Consequences of Functional Expression of the Plasma Membrane Ca2+ Pump Isoform 1aJournal of Biological Chemistry, 1996
- Oxidative Modification of a Carboxyl-Terminal Vicinal Methionine in Calmodulin by Hydrogen Peroxide Inhibits Calmodulin-Dependent Activation of the Plasma Membrane Ca-ATPaseBiochemistry, 1996
- Energetics and dynamics of molecular recognition by calmodulinBiochemistry, 1995
- Fluorescence energy transfer analysis of calmodulin.cntdot.peptide complexesBiochemistry, 1992
- A simple method for displaying the hydropathic character of a proteinJournal of Molecular Biology, 1982