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Studies of the Structure of Potassium Channel KcsA in the Open Conformation and the Effect of Anionic Lipids on Channel Inactivation
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| 자료유형 | 학위논문 |
|---|---|
| 서명/저자사항 | Studies of the Structure of Potassium Channel KcsA in the Open Conformation and the Effect of Anionic Lipids on Channel Inactivation. |
| 개인저자 | Zhang, Dongyu (Allison). |
| 단체저자명 | Columbia University. Chemistry. |
| 발행사항 | [S.l.]: Columbia University., 2019. |
| 발행사항 | Ann Arbor: ProQuest Dissertations & Theses, 2019. |
| 형태사항 | 103 p. |
| 기본자료 저록 | Dissertations Abstracts International 81-04B. Dissertation Abstract International |
| ISBN | 9781392448816 |
| 학위논문주기 | Thesis (Ph.D.)--Columbia University, 2019. |
| 일반주기 |
Source: Dissertations Abstracts International, Volume: 81-04, Section: B.
Advisor: McDermott, Ann. |
| 이용제한사항 | This item must not be sold to any third party vendors. |
| 요약 | Membrane proteins play a vital role in cellular processes. In this thesis, we use KcsA, a prokaryotic potassium channel, as a model to investigate the gating mechanism of ion channels and the effect of anionic lipids on the channel activity using solid-state NMR spectroscopy.KcsA activity is known to be highly dependent on the presence of negatively charged lipids. Multiple crystal structures combined with biochemistry assays suggest that KcsA is co-purified with anionic lipids with phosphatidylglycerol headgroup. Here, we identified this specifically bound, isotopically labeled lipid in the protein 13C-13C correlation spectra. Our results reveal that the lipid cross peaks show stronger intensity when the channel is in the inactivated state compared to the activated state, which indicates a stronger protein-lipid interaction when KcsA is inactivated. In addition, our data shows that including anionic lipids into proteoliposomes leads to a weaker potassium ion affinity at the selectivity filter. Considering ion loss as a model of inactivation, our results suggest anionic lipids promote channel inactivation. However, the surface charge is not the only physical parameter that regulates channel gating or conformational preference. We found that the channel adapted to different conformations when reconstituted into liposome either made of DOPC or DOPE, two zwitterionic lipids.Also, we were able to stabilize the open-conformation of KcsA in 3:1 DOPE/DOPG liposome at pH 4.0 and acquired several multi-dimensional solid-state NMR experiments for site-specific resonance assignments. This is the first time that we obtain wild-type full-length KcsA structural information on the transient state. The structure is not only important for understanding channel gating, but can also serve as a homology model for investigating drug binding with more complicated potassium channels such as human voltage gated channel (hERG). |
| 일반주제명 | Chemistry. Biophysics. |
| 언어 | 영어 |
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