Off-campus UMass Amherst users: To download campus access dissertations, please use the following link to log into our proxy server with your UMass Amherst user name and password.
Non-UMass Amherst users: Please talk to your librarian about requesting this dissertation through interlibrary loan.
Dissertations that have an embargo placed on them will not be available to anyone until the embargo expires.
Author ORCID Identifier
Open Access Dissertation
Doctor of Philosophy (PhD)
Year Degree Awarded
Month Degree Awarded
Myosins are a family of motor proteins responsible for various forms of cellular motility, including muscle contraction and vesicular transport. The most fundamental aspect of myosin is its ability to transduce the chemical energy from the hydrolysis of ATP into mechanical work, in the form of force and motion. A key unanswered question of the transduction process is, ”what is the relative timing between the pow- erstroke and release of phosphate Pi from the active site?”. We examined the ability of single-headed myosin Va to generate a powerstroke in a single molecule laser trap assay while maintaining Pi in its active site in of of two ways: 1) by elevating Pi in solution or 2) by introducing a mutation in myosin’s active site (S217A) which slow Pi-release from the active site. An autaomted analysis program for the detection of single molecule binding events was developed and showed that upon binding to the actin filament, WT myosin generated a powerstoke rapidly (500/s) and without a detectable delay, both in the absence and presence of 30 mM Pi . The elevated levels of Pi did, however, affect event lifetime, eliminating the longest 25 percent of binding events, confirming that Pi rebound to myosin’s active site and accelerated detachment. The S217A construct also generated a powerstroke similar in size and rate upon binding to actin despite the slower Pi release rate. These findings provide direct evidence that myosin Va generates a powerstroke with Pi still in the active site. Therefore, the findings are most consistent with a model in which the powerstroke occurs prior to the release of Pi from the active site.
Scott, Brent, "What is the Relative Timing Between Myosin’s Powerstroke and Pi-release?" (2022). Doctoral Dissertations. 2644.
Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 License.