Investigation of cardiac myosin binding protein C (cMyBPC) domains and their interactions

Myosin binding protein C (MyBPC) is a multidomain protein present in the thick filament of striated muscles involved in both sarcomere formation and contraction. It appears that the C-terminus of the protein is involved in sarcomere formation, through interaction with Light Meromyosin (LMM), while the N-terminus seems to play a role in muscle contraction, being localized close to the motor domain of myosin that interacts with the thin filament protein actin, causing muscles to contract. It exists in three different isoforms, one for each type of muscle, the cardiac isoform being linked to hypertrophic cardiomyopathy (HCM), a genetic disorder associated with cardiac dysfunction that can manifest itself through arrhythmias, heart failure and sudden cardiac death, especially in the young. The present work has focused on two aspects of the cardiac isoform (cMyBPC) investigating both the function and the role in sarcomere assembly played by cMyBPC. The first part of the research presented in this thesis consisted in the determination of the three dimensional structure of the cardiac specific N-terminal domain cC0 using NMR spectroscopy, and the investigation of the interaction with its probable binding partner, the regulatory light chain of myosin, through ^1H/^{15}N HSQC NMR spectroscopy titrations. The knowledge of the three dimensional structure has proven a vital tool to map the interacting surface on cC0, giving the possibility to make hypothesis on the way cMyBPC might interact with the S1 domain of myosin, thus influencing muscle contraction. The second part of this research is based on the hypothesis that the central domain cC5 could interact with domain cC8 of another molecule of the same protein, suggesting that the way cMyBPC could incorporate in the sarcomere would be through a trimeric collar wrapped around the myosin filament. This aspect has being studied using ^1H/^{15}N HSQC NMR spectroscopy titrations but no conclusive results were obtained, suggesting that this interaction might not take place and indicating an axial arrangement, with cMyBPC running parallel to the thick filament, as the most likely.