The mechanism by which muscle generates force has been the subject of considerable study. Key to our understanding of this mechanism is the conformational changes occurring in the myosin "head" as it interacts with the thin filament. Each myosin head individually binds to the thin filament, hydrolyzes ATP and generates force. This study has used electron paramagnetic resonance (EPR) spectroscopy to study the structure and function of the regulatory domain located on the distal portion of the myosin head. Skeletal myosin S1 was labeled with a variety of lysine targeting spin labels, under conditions optimized to label the reactive lysine residue Lys-83, to determine if spin label suitable for EPR studies could be found. Two spin labels (HO-226 and HO-2095T) had ordered populations. Competitions, of the two labels with selective blocking agents, determined that they were labeling both lysine and cysteine residues. The anisotropy for both labels was found to derive from labeled cysteine residues not Lys-83, rendering both unsuitable for EPR studies of myosin S1. The mobility of the essential light chain (ELC) and the regulatory light chain (RLC) subdomains of the regulatory domain was individually measured utilizing saturation transfer electron paramagnetic resonance (ST-EPR) spectroscopy. Their mobilities were found to be similar, and that similarity persisted under conditions, which increase the overall mobility of the myosin head: upon RLC phosphorylation, an increase of pH or the presence of divalent cations. Modeling of this mobility enabled calculation of the persistence length of the regulatory domain, which at 1.5 µm, is adequate for it to serve as a lever arm. This is consistent with theories of force generation where the regulatory domain serves as a lever to amplify movement of the catalytic domain during the power stroke of striated muscle. / A Dissertation Submitted to the Program in Molecular Biophysics in Partial
Fulfillment of the Requirements for the Degree of Doctor of Philosophy. / Spring Semester, 2003. / December 16, 2002. / muscle, force, mechanism / Includes bibliographical references. / Tim Logan, Professor Directing Dissertation; David Van Winkle, Outside Committee Member; Peter Fajer, Committee Member; Nancy L. Greenbaum, Committee Member; Kenneth A. Taylor, Committee Member.
| Identifer | oai:union.ndltd.org:fsu.edu/oai:fsu.digital.flvc.org:fsu_175702 |
| Contributors | Baumann, Bruce A. J. (authoraut), Logan, Tim (professor directing dissertation), Van Winkle, David (outside committee member), Fajer, Peter (committee member), Greenbaum, Nancy L. (committee member), Taylor, Kenneth A. (committee member), Department of Physics (degree granting department), Florida State University (degree granting institution) |
| Publisher | Florida State University, Florida State University |
| Source Sets | Florida State University |
| Language | English, English |
| Detected Language | English |
| Type | Text, text |
| Format | 1 online resource, computer, application/pdf |
| Rights | This Item is protected by copyright and/or related rights. You are free to use this Item in any way that is permitted by the copyright and related rights legislation that applies to your use. For other uses you need to obtain permission from the rights-holder(s). The copyright in theses and dissertations completed at Florida State University is held by the students who author them. |
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