Event Title
Oxidation Effects the Myosin Motor Domain Function and Structure
Location
CSU Ballroom
Start Date
11-4-2017 10:00 AM
End Date
11-4-2017 11:30 AM
Student's Major
Chemistry and Geology
Student's College
Science, Engineering and Technology
Mentor's Name
Rebecca Moen
Mentor's Department
Chemistry and Geology
Mentor's College
Science, Engineering and Technology
Description
This research examines the functional and structural effects of oxidative modification in Dicty myosin II in the motor domain. Myosin has been shown to be a target of oxidative stress. Reactive oxygen species target specific amino acids within contractile proteins leading to site directed oxidative modifications. Oxidative stress of muscle proteins is associated with aging and chronic diseases that induce muscle wasting. The goal was to obtain molecular level information on the myosin-actin relationship to understand the effects of myosin oxidation and muscle dysfunction using site-directed mutagenesis (SDM), molecular modeling, enzyme kinetics, site-direct spin- labeling and electron paramagnetic resonance (EPR). The structural focus was on the force- generating domain of myosin, specifically the relay helix, which undergoes a large structural change with contraction. SDM was used to introduce new cysteine sites to monitor structural changes in the relay helix. Labels bind by disulfide linkage to amino acid residues at i and i+4 on an alpha helix at sites 492 and 496, and 639 and 643 in the myosin primary sequence. The function of this mutated myosin was tested using an actin-activated ATPase assay to confirm that neither mutagenesis nor spin-labeling affected myosin's function. The oxidized labeled-myosin actin- activated ATPase activity showed a 3-fold decrease in activity compared to the unoxidized labeled myosin. Results for changes in myosin structural dynamics based on EPR data is yet to be determined although simulations of the data suggest a change will be detectable. These results provide knowledge on protein oxidation effects with potential therapeutic significance.
Oxidation Effects the Myosin Motor Domain Function and Structure
CSU Ballroom
This research examines the functional and structural effects of oxidative modification in Dicty myosin II in the motor domain. Myosin has been shown to be a target of oxidative stress. Reactive oxygen species target specific amino acids within contractile proteins leading to site directed oxidative modifications. Oxidative stress of muscle proteins is associated with aging and chronic diseases that induce muscle wasting. The goal was to obtain molecular level information on the myosin-actin relationship to understand the effects of myosin oxidation and muscle dysfunction using site-directed mutagenesis (SDM), molecular modeling, enzyme kinetics, site-direct spin- labeling and electron paramagnetic resonance (EPR). The structural focus was on the force- generating domain of myosin, specifically the relay helix, which undergoes a large structural change with contraction. SDM was used to introduce new cysteine sites to monitor structural changes in the relay helix. Labels bind by disulfide linkage to amino acid residues at i and i+4 on an alpha helix at sites 492 and 496, and 639 and 643 in the myosin primary sequence. The function of this mutated myosin was tested using an actin-activated ATPase assay to confirm that neither mutagenesis nor spin-labeling affected myosin's function. The oxidized labeled-myosin actin- activated ATPase activity showed a 3-fold decrease in activity compared to the unoxidized labeled myosin. Results for changes in myosin structural dynamics based on EPR data is yet to be determined although simulations of the data suggest a change will be detectable. These results provide knowledge on protein oxidation effects with potential therapeutic significance.
Recommended Citation
Ott, Rachel. "Oxidation Effects the Myosin Motor Domain Function and Structure." Undergraduate Research Symposium, Mankato, MN, April 11, 2017.
https://cornerstone.lib.mnsu.edu/urs/2017/poster-session-A/32
