Ke, Ying and Hunter, Mark J and Ng, Chai Ann and Perry, Matthew D and Vandenberg, Jamie I (2014) Role of the cytoplasmic N-terminal Cap and Per-Arnt-Sim (PAS) domain in trafficking and stabilization of Kv11.1 channels. The Journal of Biological Chemistry, 289 (20). pp.13782-91. ISSN 1083-351X (OA)
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Abstract
The N-terminal cytoplasmic region of the Kv11.1a potassium channel contains a Per-Arnt-Sim (PAS) domain that is essential for the unique slow deactivation gating kinetics of the channel. The PAS domain has also been implicated in the assembly and stabilization of the assembled tetrameric channel, with many clinical mutants in the PAS domain resulting in reduced stability of the domain and reduced trafficking. Here, we use quantitative Western blotting to show that the PAS domain is not required for normal channel trafficking nor for subunit-subunit interactions, and it is not necessary for stabilizing assembled channels. However, when the PAS domain is present, the N-Cap amphipathic helix must also be present for channels to traffic to the cell membrane. Serine scan mutagenesis of the N-Cap amphipathic helix identified Leu-15, Ile-18, and Ile-19 as residues critical for the stabilization of full-length proteins when the PAS domain is present. Furthermore, mutant cycle analysis experiments support recent crystallography studies, indicating that the hydrophobic face of the N-Cap amphipathic helix interacts with a surface-exposed hydrophobic patch on the core of the PAS domain to stabilize the structure of this critical gating domain. Our data demonstrate that the N-Cap amphipathic helix is critical for channel stability and trafficking.
(Grant-in-aid G11S 5829 from the Health Foundation of Australia and NHMRC Research Fellowship 1019693).
| Item Type: | Article |
|---|---|
| Subjects: | R Medicine > R Medicine (General) |
| Depositing User: | Repository Administrator |
| Date Deposited: | 21 Jan 2016 05:21 |
| Last Modified: | 27 May 2016 06:49 |
| URI: | https://eprints.victorchang.edu.au/id/eprint/204 |
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