title: Probing the architecture of an L-type calcium channel with a charged phenylalkylamine: evidence for a widely open pore and drug trapping.
abstract: Voltage-gated calcium channels are in a closed conformation at rest and open temporarily when the membrane is depolarized. To gain insight into the molecular architecture of Ca(v)1.2, we probed the closed and open conformations with the charged phenylalkylamine (-)devapamil ((-)qD888). To elucidate the access pathway of (-)D888 to its binding pocket from the intracellular side, we used mutations replacing a highly conserved Ile-781 by threonine/proline in the pore-lining segment IIS6 of Ca(v)1.2 (1). The shifted channel gating of these mutants (by 30-40 mV in the hyperpolarizing direction) enabled us to evoke currents with identical kinetics at different potentials and thus investigate the effect of the membrane potentials on the drug access per se. We show here that under these conditions the development of channel block by (-)qD888 is not affected by the transmembrane voltage. Recovery from block at rest was, however, accelerated at more hyperpolarized voltages. These findings support the conclusion that Ca(v)1.2 must be opening widely to enable free access of the charged (-)D888 molecule to its binding site, whereas drug dissociation from the closed channel conformation is restricted by bulky channel gates. The functional data indicating a location of a trapped (-)D888 molecule close to the central pore region are supported by a homology model illustrating that the closed Ca(v)1.2 is able to accommodate a large cation such as (-)D888.
authors.affiliation: Institute for Pharmacology and Toxicology, University of Vienna, Althanstrasse 14, A-1090 Vienna, Austria.
authors.forename: Stanislav
authors.initials: S
authors.lastname: Beyl
authors.forename: Eugen N
authors.initials: EN
authors.lastname: Timin
authors.forename: Annette
authors.initials: A
authors.lastname: Hohaus
authors.forename: Anna
authors.initials: A
authors.lastname: Stary
authors.forename: Michaela
authors.initials: M
authors.lastname: Kudrnac
authors.forename: Robert H
authors.initials: RH
authors.lastname: Guy
authors.forename: Steffen
authors.initials: S
authors.lastname: Hering
mesh_terms: D001665:Binding Sites; D002121:Calcium Channel Blockers / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D020746:Calcium Channels, L-Type / Q000737:chemistry* / Q000378:metabolism* / Q000502:physiology; D002460:Cell Line; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology; D011485:Protein Binding; D011487:Protein Conformation; D055672:Static Electricity; D014700:Verapamil / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology
chemical_list: D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; C043184:4-desmethoxyverapamil; D014700:Verapamil