Theses and Dissertations
ORCID
https://orcid.org/0000-0002-6179-7653
Advisor
Johnson, Christopher
Committee Member
Fitzkee, Nicholas
Committee Member
Webster, Charles Edwin
Committee Member
Kroncke, Brett
Committee Member
Emerson, Joseph
Date of Degree
5-15-2026
Original embargo terms
Embargo 2 years
Document Type
Dissertation - Open Access
Major
Chemistry
Degree Name
Doctor of Philosophy (Ph.D.)
College
College of Arts and Sciences
Department
Department of Chemistry
Abstract
Voltage-gated sodium channels (NaV) initiate the initial upstroke of the action potential in excitable cells. Dysfunctional channels often cause disease such as arrhythmia, epilepsy, and neuropathy. An accessory protein of NaV is calmodulin (CaM), a Ca2+-sensing protein involvedin many signaling pathways. The interaction between CaM and the cardiac NaV (NaV1.5) is of interest as it renders the channel sensitive to Ca2+. Notably, CaM binds to two cytosolic NaV1.5 components: the IQ motif on the C-terminal domain and the DII-DIV linker known as the inactivation gate. In cardiomyocytes, intracellular pH (pHi) is decreased in ischemic conditions, altering many cellular processes. Here, pHi modification of the NaV1.5 and CaM was investigated through H1915, a residue on the NaV1.5 IQ motif. Point mutations (H1915A, H1915Y, H1915K) were used to test intermolecular forces between CaM and NaV1.5 H1915 at pH 7.4 and 6.6 in the presence and absence of Ca2+. Fluorescence spectroscopy data demonstrated that CaM-IQ complex Ca2+-binding (affinity and cooperativity) was sensitive to pH. Mutations to H1915 were able to minimize or remove this pH effect. ITC was used to investigate the effect of pH on CaM-IQ binding affinity (protein-protein interaction). Thermodynamic properties of H1915A and H1915K (enthalpy and entropy) were pH sensitive. Conversely, CaM-IQ binding affinity was not pH sensitive irrespective of point mutation. NMR spectroscopy data demonstrated that pH altered all CaM-IQ motif interactions. In the absence of Ca2+, many Ca2+-binding residues displayed pH-driven chemical shift perturbations. There was less pH sensitivity in the presence of Ca2+. Whole-cell patch clamp characterized the influence of pHi on NaV function. WT NaV1.5 function was not sensitive to changes in pHi. Intriguingly, H1915A and H1915K NaV1.5 displayed alterations in function from pHi 7.4 to 6.6. Together, these results highlight the importance of H1915 for appropriate function of this CaM-IQ complex, as well as the pH effect on this interaction and CaM Ca2+-binding.
Sponsorship (Optional)
National Institute of Health, American Heart Association
Recommended Citation
Agee, Taylor Ann, "A multimodal biophysical investigation into the pH sensitivity of the voltage-gated cardiac sodium channel IQ motif – calmodulin complex" (2026). Theses and Dissertations. 6856.
https://scholarsjunction.msstate.edu/td/6856
