title: Adenosine triphosphate regulates the activity of guinea pig Cav1.2 channel by direct binding to the channel in a dose-dependent manner.
abstract: The present study is to investigate the mechanism by which ATP regulates Cav1.2 channel activity. Ventricular tissue was obtained from adult guinea pig hearts using collagenase. Ca(2+) channel activity was monitored using the patch-clamp technique. Proteins were purified using wheat germ agglutinin-Sepharose, and the concentration was determined using the Coomassie brilliant blue technique. ATP binding to the Cav1.2 channel was examined using the photoaffinity method. EDA-ATP-biotin maintains Ca(2+) channel activity in inside-out membrane patches. ATP directly bound to the Cav1.2 channel in a dose-dependent manner, and at least two molecules of ATP bound to one molecule of the Cav1.2 channel. Low levels of calmodulin (CaM) increased ATP binding to the Cav1.2 channel, but higher levels of CaM decreased ATP binding to the Cav1.2 channel. In addition, Ca(2+) was another regulator for ATP binding to the Cav1.2 channel. Furthermore, ATP bound to GST-fusion peptides of NH2-terminal region (amino acids 6-140) and proximal COOH-terminal region (amino acids 1,509-1,789) of the main subunit (α1C) of the Cav1.2 channel. Our data suggest that ATP might regulate Cav1.2 channel activity by directly binding to the Cav1.2 channel in a dose-dependent manner. In addition, the ATP-binding effect to the Cav1.2 channel was both CaM- and Ca(2+) dependent.
authors.affiliation: Department of Pharmaceutical Toxicology, School of Pharmacy, China Medical University, Shenyang, China; and.
authors.forename: Rui
authors.initials: R
authors.lastname: Feng
authors.forename: Jianjun
authors.initials: J
authors.lastname: Xu
authors.forename: Etsuko
authors.initials: E
authors.lastname: Minobe
authors.forename: Asako
authors.initials: A
authors.lastname: Kameyama
authors.forename: Lei
authors.initials: L
authors.lastname: Yang
authors.forename: Lifeng
authors.initials: L
authors.lastname: Yu
authors.forename: Liying
authors.initials: L
authors.lastname: Hao
authors.forename: Masaki
authors.initials: M
authors.lastname: Kameyama
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D001665:Binding Sites; D002118:Calcium / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000187:drug effects* / Q000378:metabolism; D002147:Calmodulin / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D005260:Female; D006168:Guinea Pigs; D006352:Heart Ventricles / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D008564:Membrane Potentials; D011993:Recombinant Fusion Proteins / Q000378:metabolism
chemical_list: C079146:CACNA1C protein, human; D020746:Calcium Channels, L-Type; D002147:Calmodulin; C419385:L-type calcium channel alpha(1C); D011993:Recombinant Fusion Proteins; D000255:Adenosine Triphosphate; D002118:Calcium
keywords: L-type Ca2+ channel; adenosine-triphosphate; calmodulin; phosphorylation