| Popis: |
Troponin C (TnC) facilitates muscle contraction through calcium-binding within its N-terminal region (NTnC). Upon calcium binding, this regulatory NTnC domain opens to expose a hydrophobic patch region in order to bind the TnI-switch peptide which ultimately results in sliding of tropomyosin on the actin filament for myosin binding and muscle contraction to occur. This work has focused on understanding how the ability of cNTnC to open relates to its apparent calcium sensitivity and modifying this ability to open in order to modulate its native calcium sensitivity. To this end, we have developed an umbrella sampling protocol to assess known cardiomyopathy mutations’ effects on the cNTnC. This work was then extended to understand the mechanism of small molecule manipulation of calcium sensitivity. And then this scheme was applied in a design process to create potential candidates to offset cardiomyopathy-related muscle contraction problems. The umbrella sampling protocol was effective in showing that certain cardiomyopathy mutations and calcium-sensitizing mutations, as well as many of the small molecules tested, in fact, do alter this ability to expose the hydrophobic patch region, likely by destabilizing the closed conformation. From this confirmation of experimental work, our extension into predicting new variants was given solid grounding and testing these experimentally is the last step in validation. |
