In response to membrane potential depolarization, voltage-dependent channels undergo a series of conformational changes from a non-conducting state (closed) to an activated (conducting), finally stabilizing in a non-conducting inactivated state. K+ channel function has been associated with such basic cellular functions as the regulation of electrical activity, signal transduction and osmotic balance. In higher organisms, K+ channel dysfunction may lead to uncontrolled periods of electrical hyperexcitability, like epileptic episodes, myotonia and cardiac arrhythmia. Consequently, efforts to understand K+ channel structure function and dynamics relate directly to human health and disease. The continuing long-term goal of this project is to further understand the molecular mechanisms of gating in voltage-dependent channels, by focusing on the analysis of K+ channel gating in prokaryotic and eukaryotic systems. Specifically we will address the following key questions: What are the atomic structures of the key conformations that determine channel activity? This question will be answered for membrane embedded systems as well as those ordered in a lattice. What are the molecular bases of voltage-dependent gating? We will be testing the hypothesis that a specific sliding helix movement (the one click motion) can explain charge translocation in certain voltage sensors, but perhaps not others. The more charge a sensor translocates, the larger the number of clicks its sensor needs to move. And how different parts of the channel interact to define open channel activity? We plan to study these problems by combining spectroscopic techniques (EPR and NMR), X-ray crystallography electrophysiological and computational methods. We intend to continue these structure-function studies by investigating a wealth of biochemically-defined systems from KcsA and KvAP, to the Shaker voltage sensor and the hyperpolarization- activated channel from Methanococcus janschii (MVP). In addition, we will focus our attention on the voltage- sensing domain from the Ciona intestinalis-Voltage-Sensor-containing Phosphatase (Ci-VSP) aiming to improve the resolution of our recent crystals structures. Finally, we will examine the structure of the human voltage-dependent proton channel Hv1 in membranes though an extensive site-directed spin labeling analysis and computational modeling. This proposal should open new experimental avenues that will contribute to our understanding of biologically important events such as electrical signaling, signal transduction and ion channel gating.

Public Health Relevance

Potassium channels are membrane proteins that catalyze the transfer of K+ ions down an electrochemical gradient with high efficiency and selectivity. Understanding of voltage-dependent K+ channel structure and function relates directly to health and disease. Their function has been associated with such basic cellular functions as the regulation of electrical activity, signal transduction and osmotic balance. K+ channels are members of the voltage-dependent channel superfamily, which include explicitly voltage-activated channels (Na+, Ca2+, and a large number of K+ channels), as well as voltage-independent channels (i.e., the inward rectifiers and the cyclic nucleotide activated channels). In higher organisms, K+ channel dysfunction may lead to uncontrolled periods of electrical hyperexcitability, like epileptic episodes and cardiac arrhythmias. Not surprisingly, voltage-dependent channels are also the target of many therapeutic agents.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
2R01GM057846-18
Application #
8818925
Study Section
Biochemistry and Biophysics of Membranes Study Section (BBM)
Program Officer
Nie, Zhongzhen
Project Start
1998-08-01
Project End
2019-01-31
Budget Start
2015-05-01
Budget End
2016-01-31
Support Year
18
Fiscal Year
2015
Total Cost
Indirect Cost
Name
University of Chicago
Department
Biochemistry
Type
Schools of Medicine
DUNS #
005421136
City
Chicago
State
IL
Country
United States
Zip Code
60637
Li, Jing; Ostmeyer, Jared; Cuello, Luis G et al. (2018) Rapid constriction of the selectivity filter underlies C-type inactivation in the KcsA potassium channel. J Gen Physiol 150:1408-1420
Zhao, Ruiming; Kennedy, Kelleigh; De Blas, Gerardo A et al. (2018) Role of human Hv1 channels in sperm capacitation and white blood cell respiratory burst established by a designed peptide inhibitor. Proc Natl Acad Sci U S A 115:E11847-E11856
Kratochvil, Huong T; Maj, Micha?; Matulef, Kimberly et al. (2017) Probing the Effects of Gating on the Ion Occupancy of the K+ Channel Selectivity Filter Using Two-Dimensional Infrared Spectroscopy. J Am Chem Soc 139:8837-8845
Vandenberg, Jamie I; Perozo, Eduardo; Allen, Toby W (2017) Towards a Structural View of Drug Binding to hERG K+ Channels. Trends Pharmacol Sci 38:899-907
Li, Jing; Ostmeyer, Jared; Boulanger, Eliot et al. (2017) Chemical substitutions in the selectivity filter of potassium channels do not rule out constricted-like conformations for C-type inactivation. Proc Natl Acad Sci U S A 114:11145-11150
Kratochvil, Huong T; Carr, Joshua K; Matulef, Kimberly et al. (2016) Instantaneous ion configurations in the K+ ion channel selectivity filter revealed by 2D IR spectroscopy. Science 353:1040-1044
Li, Qufei; Shen, Rong; Treger, Jeremy S et al. (2015) Resting state of the human proton channel dimer in a lipid bilayer. Proc Natl Acad Sci U S A 112:E5926-35
Li, Qufei; Wanderling, Sherry; Paduch, Marcin et al. (2014) Structural mechanism of voltage-dependent gating in an isolated voltage-sensing domain. Nat Struct Mol Biol 21:244-52
Li, Qufei; Wanderling, Sherry; Sompornpisut, Pornthep et al. (2014) Structural basis of lipid-driven conformational transitions in the KvAP voltage-sensing domain. Nat Struct Mol Biol 21:160-6
Raghuraman, H; Islam, Shahidul M; Mukherjee, Soumi et al. (2014) Dynamics transitions at the outer vestibule of the KcsA potassium channel during gating. Proc Natl Acad Sci U S A 111:1831-6

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