A conducting state with properties of a slow inactivated state in a shaker K(+) channel mutant.
In Shaker K(+) channel, the amino terminus deletion Delta6-46 removes fast inactivation (N-type) unmasking a slow inactivation process. In Shaker Delta6-46 (Sh-IR) background, two additional mutations (T449V-I470C) remove slow inactivation, producing a noninactivating channel. However, despite the fact that Sh-IR-T449V-I470C mutant channels remain conductive, prolonged depolarizations (1 min, 0 mV) produce a shift of the QV curve by about -30 mV, suggesting that the channels still undergo the conformational changes typical of slow inactivation. For depolarizations longer than 50 ms, the tail currents measured during repolarization to -90 mV display a slow component that increases in amplitude as the duration of the depolarizing pulse increases. We found that the slow development of the QV shift had a counterpart in the amplitude of the slow component of the ionic tail current that is not present in Sh-IR. During long depolarizations, the time course of both the increase in the slow component of the tail current and the change in voltage dependence of the charge movement could be well fitted by exponential functions with identical time constant of 459 ms. Single channel recordings revealed that after prolonged depolarizations, the channels remain conductive for long periods after membrane repolarization. Nonstationary autocovariance analysis performed on macroscopic current in the T449V-I470C mutant confirmed that a novel open state appears with increasing prepulse depolarization time. These observations suggest that in the mutant studied, a new open state becomes progressively populated during long depolarizations (>50 ms). An appealing interpretation of these results is that the new open state of the mutant channel corresponds to a slow inactivated state of Sh-IR that became conductive.
Olcese
R
R
Department of Anesthesiology, School of Medicine, University of California Los Angeles, Los Angeles, CA 90095-7115, USA.
Sigg
D
D
Latorre
R
R
Bezanilla
F
F
Stefani
E
E
D000818:Animals; D016477:Artifacts; D004553:Electric Conductivity; D015640:Ion Channel Gating / Q000502:physiology*; D007700:Kinetics; D008564:Membrane Potentials / Q000502:physiology; D016296:Mutagenesis / Q000502:physiology; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000235:genetics* / Q000378:metabolism*; D011336:Probability; D051637:Shaker Superfamily of Potassium Channels; D014982:Xenopus laevis
D015221:Potassium Channels; D051637:Shaker Superfamily of Potassium Channels