Ion channels control both the resting membrane potential and the action potential in cells. As a result, improper physiological function of ion channels leads to a wide variety of disease states, including cardiac and neurological dysfunction, epilepsy, asthma, hearing loss, and others. Proper ion channel function requires appropriate ion selectivity and channel gating to control ion flux across biological membranes. However, our understanding of these fundamental properties of ion channels is still incomplete because they arise from a complex interplay of structure and dynamics. We will use solution NMR to experimentally test the role of protein dynamics in ion selectivity and gating of the NaK channel from Bacillus cereus solubilized in isotropic bicelles. NaK is an ideal model system because small mutations can shift its selectivity; its structure is similar to eukaryotic potassium channel pore domains, particularly biomedically important cyclic nucleotide gated and HERG potassium channels; and it is highly stable and amenable to structural and biophysical characterization. NMR provides a unique method to simultaneously and quantitatively probe ion channel structure and dynamics with site-specific resolution. The ability of NMR to measure dynamics on diverse timescales and detect transient and lowly populated states allows us to experimentally evaluate the role of protein dynamics on processes ranging from ion selectivity to channel gating. In this proposal we will use NMR studies of NaK to test three aims: Does selectivity arise from cooperative effects of ion binding on backbone dynamics? How does the N-terminal M0 helix regulate NaK gating? What structural and dynamic features allow allosteric communication between the selectivity filter and inner gate in potassium channels?

Public Health Relevance

To properly control ion flux across membranes, channels must maintain appropriate selectivity and gating behavior. Our understanding of these fundamental ion channel properties is still incomplete because they arise from a complex interplay of structure and dynamics. We will use solution NMR to experimentally test the role of protein dynamics in ion selectivity and gating in NaK, a non-selective cation channel.

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
5R01GM116047-02
Application #
9133430
Study Section
Biophysics of Neural Systems Study Section (BPNS)
Program Officer
Chin, Jean
Project Start
2015-09-01
Project End
2019-06-30
Budget Start
2016-07-01
Budget End
2017-06-30
Support Year
2
Fiscal Year
2016
Total Cost
Indirect Cost
Name
University of Wisconsin Madison
Department
Biochemistry
Type
Earth Sciences/Resources
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Brettmann, Joshua B; Urusova, Darya; Tonelli, Marco et al. (2015) Role of protein dynamics in ion selectivity and allosteric coupling in the NaK channel. Proc Natl Acad Sci U S A 112:15366-71