Molecular Mechanism of Use-Dependent Calcium Channel Block by Phenylalkylamines: Role of Inactivation

The role of channel inactivation in the molecular mechanism of calcium (Ca2+) channel block by phenylalkylamines (PAA) was analyzed by designing mutant Ca2+ channels that carry the high affinity determinants of the PAA receptor site [Hockerman, G. H., Johnson, B. D., Scheuer, T., and Catterall, W. A...

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Veröffentlicht in:Proceedings of the National Academy of Sciences - PNAS 1997-11, Vol.94 (24), p.13323-13328
Hauptverfasser: Hering, Steffen, Aczél, Stefan, Kraus, Richard L., Berjukow, Stanislav, Striessnig, Jörg, Timin, Eugen N.
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Sprache:eng
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Zusammenfassung:The role of channel inactivation in the molecular mechanism of calcium (Ca2+) channel block by phenylalkylamines (PAA) was analyzed by designing mutant Ca2+ channels that carry the high affinity determinants of the PAA receptor site [Hockerman, G. H., Johnson, B. D., Scheuer, T., and Catterall, W. A. (1995) J. Biol. Chem. 270,22119-22122] but inactivate at different rates. Use-dependent block by PAAs was studied after expressing the mutant Ca2+ channels in Xenopus oocytes. Substitution of single putative poreorientated amino acids in segment IIIS6 by alanine (F-1499-A, F-1500-A, F-1510-A, I-1514-A, and F-1515-A) gradually slowed channel inactivation and simultaneously reduced inhibition of barium currents (IBa) by (-)D600 upon depolarization by 100 ms steps at 0.1 Hz. This apparent reduction in drug sensitivity was only evident if test pulses were applied at a low frequency of 0.1 Hz and almost disappeared at the frequency of 1 Hz. (-)D600 slowed IBa recovery after maintained membrane depolarization (1-3 sec) to a comparable extent in all channel constructs. A drug-induced delay in the onset of IBa recovery from inactivation suggests that PAAs promote the transition to a deep inactivated channel conformation. These findings indicate that apparent PAA sensitivity of Ca2+ channels is not only defined by drug interaction with its receptor site but also crucially dependent on intrinsic gating properties of the channel molecule. A molecular model for PAA-Ca2+ channel interaction that accounts for the relationship between drug induced inactivation and channel block by PAA is proposed.
ISSN:0027-8424
1091-6490
DOI:10.1073/pnas.94.24.13323