title: The potassium channel interacting protein 3 (DREAM/KChIP3) heterodimerizes with and regulates calmodulin function.
abstract: Downstream regulatory element antagonistic modulator (DREAM/KChIP3), a neuronal EF-hand protein, modulates pain, potassium channel activity, and binds presenilin 1. Using affinity capture of neuronal proteins by immobilized DREAM/KChIP3 in the presence and absence of calcium (Ca(2+)) followed by mass spectroscopic identification of interacting proteins, we demonstrate that in the presence of Ca(2+), DREAM/KChIP3 interacts with the EF-hand protein, calmodulin (CaM). The interaction of DREAM/KChIP3 with CaM does not occur in the absence of Ca(2+). In the absence of Ca(2+), DREAM/KChIP3 binds the EF-hand protein, calcineurin subunit-B. Ca(2+)-bound DREAM/KChIP3 binds CaM with a dissociation constant of ∼3 μM as assessed by changes in DREAM/KChIP3 intrinsic protein fluorescence in the presence of CaM. Two-dimensional (1)H,(15)N heteronuclear single quantum coherence spectra reveal changes in chemical shifts and line broadening upon the addition of CaM to (15)N DREAM/KChIP3. The amino-terminal portion of DREAM/KChIP3 is required for its binding to CaM because a construct of DREAM/KChIP3 lacking the first 94 amino-terminal residues fails to bind CaM as assessed by fluorescence spectroscopy. The addition of Ca(2+)-bound DREAM/KChIP3 increases the activation of calcineurin (CN) by calcium CaM. A DREAM/KChIP3 mutant incapable of binding Ca(2+) also stimulates calmodulin-dependent CN activity. The shortened form of DREAM/KChIP3 lacking the NH(2)-terminal amino acids fails to activate CN in the presence of calcium CaM. Our data demonstrate the interaction of DREAM/KChIP3 with the important EF-hand protein, CaM, and show that the interaction alters CN activity.
authors.affiliation: Division of Nephrology and Hypertension, Mayo Clinic, Rochester, Minnesota 55905, USA.
authors.forename: Pradeep L
authors.initials: PL
authors.lastname: Ramachandran
authors.forename: Theodore A
authors.initials: TA
authors.lastname: Craig
authors.forename: Elena A
authors.initials: EA
authors.lastname: Atanasova
authors.forename: Gaofeng
authors.initials: G
authors.lastname: Cui
authors.forename: Barbara A
authors.initials: BA
authors.lastname: Owen
authors.forename: H Robert
authors.initials: HR
authors.lastname: Bergen
authors.forename: Georges
authors.initials: G
authors.lastname: Mer
authors.forename: Rajiv
authors.initials: R
authors.lastname: Kumar
mesh_terms: D019703:Calcineurin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D002118:Calcium / Q000737:chemistry / Q000378:metabolism*; D002147:Calmodulin / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D051605:Kv Channel-Interacting Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D011485:Protein Binding; D055503:Protein Multimerization / Q000502:physiology*; D017434:Protein Structure, Tertiary; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D002147:Calmodulin; C496749:KCNIP3 protein, human; D051605:Kv Channel-Interacting Proteins; D012097:Repressor Proteins; D019703:Calcineurin; D002118:Calcium