TROPONIN COMPLEX BIOPHYSICAL CHANGES INDUCED BY CARDIAC TROPONIN T MUTATIONS

picture of Ofer Fass presenting his/her poster: TROPONIN COMPLEX BIOPHYSICAL CHANGES INDUCED BY CARDIAC TROPONIN T MUTATIONS

Ofer Fass , James Jayant, Mark McConnel, Sara Fliehman, Jil Tardiff

TROPONIN COMPLEX BIOPHYSICAL CHANGES INDUCED BY CARDIAC TROPONIN T MUTATIONS

Cardiomyopathies are diseases of the heart muscle that impair cardiovascular function, often leading to heart disease or sudden cardiac death. Various genetic mutations have been implicated in the initiation and progression of cardiomyopathies. My research focuses on mutations to one of the contractile regulatory proteins found in heart muscle cells, cardiac troponin T (cTnT). cTnT is one of three regulatory proteins that together form the troponin complex, the other two being cardiac troponin I, (cTnI) and cardiac troponin C, (cTnC). During cardiac contraction, a conformational change occurs causing a pocket to form between cTnC and cTnI, exposing the cTnI phosphorylation site at serine 23/24. Phosphorylation of cTnI by protein kinase A (PKA) is a crucial step in increasing the speed of cardiac relaxation during beta-adrenergic stimulation. Therefore, the inability of PKA to properly phosphorylate cTnI can lead to impaired relaxation of the heart and subsequent cardiovascular complications.

The purpose of this study is to determine whether mutations to cTnT induce aberrant allosteric changes that decrease the size of the cTnI-cTnC pocket, preventing PKA phosphorylation of cTnI. The main tool utilized is fluorescence resonance energy transfer (FRET). FRET is used to measure very small distances between two fluorophores. By tagging cTnI with a donor fluorophore and cTnT with an acceptor fluorophore, size measurements of the cTnI-cTnC pocket can be made. Measurements are being conducted for troponin complexes containing wild type cTnT, cTnT mutants and various non-cTnT related mutants. In this way, it can be determined whether cTnT mutations are altering the troponin pocket size. Due to the ongoing nature of this research, no results have yet been obtained. Currently, cTnC, cTnI and cTnT have been isolated and various mutant variants for use with FRET. In the near future I will reconstitute the thin filament and make the fluoroscopic measurements. By understanding the biophysical changes that cTnT mutations induce in the troponin complex, it may be possible to develop novel pharmaceuticals that will prevent these changes from occurring. Research is supported in part by funding from the UA Provost's Office. 

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