The long-term objective of this project is to understand the mechanisms of voltage-dependent ion channels. This study will focus on three voltage-dependent ion channels, including a eukaryotic voltage-dependent K+ channel (Aim 1), a prokaryotic voltage-dependent K+ channel (Aim 2), and a eukaryotic voltage-dependent H+ channel (Aim 3). X-ray crystallography will be used to determine atomic structures of these membrane proteins and electrophysiological recordings and ion flux assays will be used to analyze the function. In addition, NMR spectroscopy will be used to study protein structure, dynamics, and lipid interactions. Work funded by this research proposal has already led to an understanding of the open conformation structure of a voltage- dependent K+ channel. Closed conformations are still unknown. The goal of the first two aims is to determine structures of partially and fully closed conformations of an entire voltage-dependent channel (Aim 1) and an isolated voltage sensor (Aim 2) and to correlate these conformations with function so that we can understand how membrane voltage controls the conformation of voltage sensors and voltage-dependent ion channels. The closed conformations will be induced through specific mutations that are identified and evaluated in electrophysiological assays. In the third aim we seek to determine a first atomic structure of a voltage- dependent H+ channel through x-ray crystallography. Because the H+ is invisible to x-rays we will use NMR spectroscopy to study the interaction of the H+ channel with its substrate. Voltage-dependent ion channels produce electrical signals in living cells. They are essential to nervous system as well as skeletal, cardiac and smooth muscle function. Certain voltage-dependent ion channels are targets of action for important pharmacological agents. An in-depth understanding of the atomic structure and mechanisms of voltage- dependent ion channels holds promise for new approaches in the future to treat diseases such as epilepsy and cardiac arrhythmia.

Public Health Relevance

Life has evolved a marvelous molecular system for producing electrical signals that underlie our thoughts and movements. The molecules in this system are known as voltage-dependent ion channels. This project seeks to understand the atomic basis of voltage-dependent ion channel function with the hope that this knowledge will lead to new therapies for diseases such as epilepsy and cardiac arrhythmia.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM043949-22
Application #
8106944
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Hagan, Ann A
Project Start
1990-04-01
Project End
2015-07-31
Budget Start
2011-09-15
Budget End
2012-07-31
Support Year
22
Fiscal Year
2011
Total Cost
$567,825
Indirect Cost
Name
Rockefeller University
Department
Biology
Type
Other Domestic Higher Education
DUNS #
071037113
City
New York
State
NY
Country
United States
Zip Code
10065
Hite, Richard K; MacKinnon, Roderick (2017) Structural Titration of Slo2.2, a Na+-Dependent K+ Channel. Cell 168:390-399.e11
Lee, Chia-Hsueh; MacKinnon, Roderick (2017) Structures of the Human HCN1 Hyperpolarization-Activated Channel. Cell 168:111-120.e11
Tao, Xiao; Hite, Richard K; MacKinnon, Roderick (2017) Cryo-EM structure of the open high-conductance Ca(2+)-activated K(+) channel. Nature 541:46-51
Wang, Weiwei; MacKinnon, Roderick (2017) Cryo-EM Structure of the Open Human Ether-à-go-go-Related K+ Channel hERG. Cell 169:422-430.e10
Park, Eunyong; Campbell, Ernest B; MacKinnon, Roderick (2017) Structure of a CLC chloride ion channel by cryo-electron microscopy. Nature 541:500-505
Hite, Richard K; Tao, Xiao; MacKinnon, Roderick (2017) Structural basis for gating the high-conductance Ca(2+)-activated K(+) channel. Nature 541:52-57
Whicher, Jonathan R; MacKinnon, Roderick (2016) Structure of the voltage-gated K? channel Eag1 reveals an alternative voltage sensing mechanism. Science 353:664-9
Su, Zhenwei; Brown, Emily C; Wang, Weiwei et al. (2016) Novel cell-free high-throughput screening method for pharmacological tools targeting K+ channels. Proc Natl Acad Sci U S A 113:5748-53
Touhara, Kouki K; Wang, Weiwei; MacKinnon, Roderick (2016) The GIRK1 subunit potentiates G protein activation of cardiac GIRK1/4 hetero-tetramers. Elife 5:
Wang, Weiwei; Touhara, Kouki K; Weir, Keiko et al. (2016) Cooperative regulation by G proteins and Na(+) of neuronal GIRK2 K(+) channels. Elife 5:

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