The CaMKII enzyme is involved in diverse biological processes including control of heart rate and synaptic plasticity. CaMKII is activated by binding of calmodulin (CaM) and ATP. Its intricate structure and dynamics are thought to underlie its mode of self-regulation. The enzyme consists of catalytic, regulatory (auto-inhibitory), and association domains. Unregulated CaMKII activity has been implicated in cardiac pathologies. As such, a better understanding of the activation mechanism of the enzyme could contribute to the development of CaMKII-specific inhibitors. The hypothesis tested was that binding of ATP to CaMKII alters the flexibility of the regulatory domain and induces conformational rearrangements in the catalytic domain of the kinase. Toward this end, studies were made on monomeric constructs lacking the association domain. Spin labels were placed at specific positions in the regulatory and catalytic domains for distance measurements by Double Electron-Electron Resonance (DEER) in basal, +ATP, and +CaM conditions. Additionally, the effect of ATP on the CaM-binding region was studied on a per-residue basis by NMR. The findings demonstrate 1) a re-structuring of the regulatory domain upon CaM binding, 2) a change in regulatory domain dynamics due to ATP binding, consistent with increased exposure of the CaM-binding segment to the solvent, and 3) shorter distances within the catalytic domain after addition of ATP, in concordance with a closed catalytic domain conformation.
| Identifer | oai:union.ndltd.org:VANDERBILT/oai:VANDERBILTETD:etd-07192016-152059 |
| Date | 22 July 2016 |
| Creators | Rodriguez Mena, Francisco Guillermo |
| Contributors | Hassane Mchaourab, Ph.D, Roger Colbran, Ph.D, Michael Stone, Ph.D, Jens Meiler, Ph.D |
| Publisher | VANDERBILT |
| Source Sets | Vanderbilt University Theses |
| Language | English |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | http://etd.library.vanderbilt.edu/available/etd-07192016-152059/ |
| Rights | restricted, I hereby certify that, if appropriate, I have obtained and attached hereto a written permission statement from the owner(s) of each third party copyrighted matter to be included in my thesis, dissertation, or project report, allowing distribution as specified below. I certify that the version I submitted is the same as that approved by my advisory committee. I hereby grant to Vanderbilt University or its agents the non-exclusive license to archive and make accessible, under the conditions specified below, my thesis, dissertation, or project report in whole or in part in all forms of media, now or hereafter known. I retain all other ownership rights to the copyright of the thesis, dissertation or project report. I also retain the right to use in future works (such as articles or books) all or part of this thesis, dissertation, or project report. |
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