Abstract
Cholesterol is a major regulator of multiple types of ion channels. Although there is increasing information about cholesterol binding sites, the molecular mechanisms through which cholesterol binding alters channel function are virtually unknown. In this study, we used a combination of Martini coarse-grained simulations, a network theory-based analysis, and electrophysiology to determine the effect of cholesterol on the dynamic structure of the Kir2.2 channel. We found that increasing membrane cholesterol reduced the likelihood of contact between specific regions of the cytoplasmic and transmembrane domains of the channel, most prominently at the subunit-subunit interfaces of the cytosolic domains. This decrease in contact was mediated by pairwise interactions of specific residues and correlated to the stoichiometry of cholesterol binding events. The predictions of the model were tested by site-directed mutagenesis of two identified residues—V265 and H222—and high throughput electrophysiology.
| Original language | English (US) |
|---|---|
| Article number | 104329 |
| Journal | iScience |
| Volume | 25 |
| Issue number | 5 |
| DOIs | |
| State | Published - May 20 2022 |
Funding
Notice: This manuscript has been authored by UT-Battelle, LLC, under contract DE-AC05-00OR22725 with the US Department of Energy (DOE) . The US government retains and the publisher, by accepting the article for publication, acknowledges that the US government retains a nonexclusive, paid-up, irrevocable worldwide license to publish or reproduce the published form of this manuscript, or allow others to do so, for US government purposes. DOE will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan ( http://energy.gov/downloads/does-public-access-plan ). We thank Ms. Dana Lazarko for her help with performing imaging experiments. We also thank the Imaging Core of Research Resource Center of the UIC for supporting these experiments. This research is supported by NIH NHLBI grants R01HL073965, R01HL141120 R01HL083298, and R01HL122010. Notice: This manuscript has been authored by UT-Battelle, LLC, under contract DE-AC05-00OR22725 with the US Department of Energy (DOE). The US government retains and the publisher, by accepting the article for publication, acknowledges that the US government retains a nonexclusive, paid-up, irrevocable worldwide license to publish or reproduce the published form of this manuscript, or allow others to do so, for US government purposes. DOE will provide public access to these results of federally sponsored research in accordance with the DOE Public Access Plan (http://energy.gov/downloads/does-public-access-plan). Funding: This research used resources at the Joint Institute for Biological Sciences (JIBS), a DOE Office of Science User Facility operated by the Oak Ridge National Laboratory. Conceptualization, N.B. S.T.G. B.S.A. and I.L.; Methodology, N.B. S.T.G. S.J.A, A.L.G. B.S.A. and I.L.; Software, N.B. D.K. and B.S.A.; Formal Analysis, N.B. and S.T.G.; Investigation, N.B. S.T.G, C.G.V. and T.V.A.; Resources, A.L.G. and I.L.; Writing – Original Draft, N.B. S.T.G. B.S.A. and I.L.; Writing – Review & Editing, N.B. S.T.G. B.S.A. and I.L.; Visualization, N.B. and S.T.G.; Supervision, A.L.G. B.S.A. and I.L.; Funding Acquisition, A.L.G. B.S.A. and I.L. The authors declare no competing interests. We thank Ms. Dana Lazarko for her help with performing imaging experiments. We also thank the Imaging Core of Research Resource Center of the UIC for supporting these experiments. This research is supported by NIH NHLBI grants R01HL073965 , R01HL141120 R01HL083298 , and R01HL122010 .
Keywords
- Biophysics
- Cellular physiology
- Molecular biology
ASJC Scopus subject areas
- General