Structure-Activity Relationships of FMRF-NH(2) Peptides Demonstrate A Role for the Conserved C Terminus and Unique N-Terminal Extension in Modulating Cardiac Contractility

FMRF-NH(2) peptides which contain a conserved, identical C-terminal tetrapeptide but unique N terminus modulate cardiac contractility; yet, little is known about the mechanisms involved in signaling. Here, the structure-activity relationships (SARs) of the Drosophila melanogaster FMRF-NH(2) peptides, PDNFMRF-NH(2), SDNFMRF-NH(2), DPKQDFMRF-NH(2), SPKQDFMRF-NH(2), and TPAEDFMRF-NH(2), which bind FMRFa-R, were investigated. The hypothesis tested was the C-terminal tetrapeptide FMRF-NH(2), particularly F1, makes extensive, strong ligand-receptor contacts, yet the unique N terminus influences docking and activity. To test this hypothesis, docking, binding, and bioactivity of the C-terminal tetrapeptide and analogs, and the FMRF-NH(2) peptides were compared. Results for FMRF-NH(2) and analogs were consistent with the hypothesis; F1 made extensive, strong ligand-receptor contacts with FMRFa-R; Y→F (YMRF-NH(2)) retained binding, yet A→F (AMRF-NH(2)) did not. These findings reflected amino acid physicochemical properties; the bulky, aromatic residues F and Y formed strong pi-stacking and hydrophobic contacts to anchor the ligand, interactions which could not be maintained in diversity or number by the small, aliphatic A. The FMRF-NH(2) peptides modulated heart rate in larva, pupa, and adult distinctly, representative of the contact sites influenced by their unique N-terminal structures. Based on physicochemical properties, the peptides each docked to FMRFa-R with one best pose, except FMRF-NH(2) which docked with two equally favorable poses, consistent with the N terminus influencing docking to define specific ligand-receptor contacts. Furthermore, SDNAMRF-NH(2) was designed and, despite lacking the aromatic properties of one F, it binds FMRFa-R and demonstrated a unique SAR, consistent with the N terminus influencing docking and conferring binding and activity; thus, supporting our hypothesis.