Derivation of Inhibitory Peptides to Target the Cardiac Troponin C–I Interaction as Potential Therapeutics for Heart Failure

Abstract

The Ca²⁺-sensitive cardiac troponin (cTn) is a hetero-trimer complex consisting of three subunits cTnC, cTnI, and cTnT, which has been recognized as an important biomarker and a potential target of cardiovascular diseases. Previously, several small-molecule agents such as levosimendan and pimobendan have been successfully developed to target this protein for the treatment of heart failure. Here, instead of small-molecule chemical drugs, we purposed rational derivation of self-inhibitory peptides as potential biologic disruptors of cTnC–cTnI interaction from the interaction complex interface. In the procedure, the crystal structure of cTn trimer was examined in detail using bioinformatics approach, from which a peptide-mediated interaction between the N-terminal domain of cTnC and the switch region of cTnI was identified. The switch is a 19-mer peptide segment Swt that contains a structured helical core capped by a short N-terminal tripeptide and a disordered C-terminal tail. Structural and energetic analysis revealed that the Swt peptide binds independently to cTnC N-terminal domain, which can be stripped from the intact cTnI subunit to interact effectively with cTnC. Further investigations found that truncation of two N-terminal residues and five C-terminal residues of the full-length Swt peptide, resulting in a shortened version namely Swt-ΔN2ΔC5 peptide, would not cause substantial loss in its binding potency to cTnC. The computational finding was then confirmed by using fluorescence-based affinity assays; the Swt and Swt-ΔN2ΔC5 peptides was experimentally measured to have a moderately high affinity to the recombinant protein of human cTnC N-terminal domain with K _d values at micromolar level. The Swt and Swt-ΔN2ΔC5 are considered as inhibitory peptides that can be further optimized and modified to obtain high-affinity disruptors of cTnI–cTnC interaction.