Ingrid Hayden

Heart disease is a leading cause of death for Americans (CDC, 2023), propelling the urgent need for drugs that restore the heart's function after a heart attack as well as preventing the onset and progression of heart diseases. The focus of my research was on the sarcoplasmic reticulum (SR) Ca-ATPase pump (SERCA). SERCA is the enzyme responsible for active transport of Ca2+ in order to relax the muscle, which allows the ventricle to fill with blood after the Ca-induced muscle contraction. Insufficient activity of SERCA2a is a leading cause of heart failure, since it prevents complete filling of the ventricle, in a “vicious cycle” that leads to toxic levels of cytoplasmic Ca2+. By examining the impact of the membrane protein, Dwarf open reading frame (DWORF), which is a direct protein activator of SERCA as well as the membrane protein PLB which inhibits SERCA, the background activity of SERCA, through fluorescence resonance energy transfer (FRET), information on how to improve the regulation of SERCA can be determined. To achieve this, DNA had to be cloned with the appropriate sequencing to encode for the expression of SERCA and the membrane proteins. The results of my research provided me with successful cloning of seven different combinations of DNA sequencing for combinations of SERCA with PLB or DWORF as well as fluorescent tags that will allow for FRET measurements. The tag mCyRFP goes with DWORF which is the donor, and the tag mMaroon goes with SERCA2a which is the acceptor. These results can either be co-expressed or expressed all at once, which provides baseline readings for future testing using small molecule drugs.