Structure-Activity Relationships of FMRF-NH2 Peptides Demonstrate A Role for the Conserved C Terminus and Unique N-Terminal Extension in Modulating Cardiac Contractility

Abstract

FMRF-NH2 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-NH2 peptides, PDNFMRF-NH2, SDNFMRF-NH2, DPKQDFMRF-NH2, SPKQDFMRF-NH2, and TPAEDFMRF-NH2, which bind FMRFa-R, were investigated. The hypothesis tested was the C-terminal tetrapeptide FMRF-NH2, 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-NH2 peptides were compared. Results for FMRF-NH2 and analogs were consistent with the hypothesis; F1 made extensive, strong ligand-receptor contacts with FMRFa-R; Y→F (YMRF-NH2) retained binding, yet A→F (AMRF-NH2) 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-NH2 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-NH2 which docked with two equally favorable poses, consistent with the N terminus influencing docking to define specific ligand-receptor contacts. Furthermore, SDNAMRF-NH2 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.

Related neuropeptides use different balances of unitary mechanisms to modulate the cardiac neuromuscular system in the American lobster,Homarus americanus

“…In the locust, for example, it appears that different receptors may respond to the FLRFamide peptides (particularly myosuppressins) in abdominal muscles than in muscles of the oviduct (Lange and Cheung 1999). In both Caenorhabditis elegans and Drosophila melanogaster, single neuropeptides, including those in the FMRFamide family, have been shown to activate two or more receptors; conversely, the same receptors are often activated by more than one related peptide, making for complex signaling possibilities (Johnson et al 2003;Klose et al 2010;Li 2005;Maynard et al 2013). Among the most intriguing of these studies is that by Klose et al (2010), demonstrating that two receptors are required for enhancement at the NMJ in a larval Drosophila muscle.…”

Modulation of neuromuscular synapses and contraction inDrosophila3rd instar larvae

“…Greenberg & Price, 1992;Lange, 2001;Mercier, Friedrich, & Boldt, 2003). In Drosophila, they are implicated in cardiac function, locomotion, flight, ecdysis and stress-induced sleep (Agrawal, Sadaf, & Hasan, 2013;Kim et al, 2006;Klose, Dason, Atwood, Boulianne, & Mercier, 2010;Lenz, Xiong, Nelson, Raizen, & Williams, 2015;Maynard et al, 2013;Taghert, 1999). The dFMRFa gene (Nambu et al, 1988;Schneider & Taghert, 1988) encodes eight distinct peptides, five of which contain the amidated C-terminal sequence FMRFamide and three with sequences similar to FMRFamide (SAPQDFVRSamide, MDSNFIRFamide and SVQDNFMHFamide).…”

“…Thus, many people have favoured the view that there is only one receptor for the peptides encoded in dFMRFa, at least at physiologically relevant peptide concentrations. Receptor and ligand modeling indicates that the five Drosophila peptides containing the sequence 'FMRFamide' exhibit subtle differences in docking and linking with the FMRFamide receptor, but they all elicit very similar responses in cardiac bioassays (Maynard et al, 2013). Hewes et al (1998) also found that DPKQDFMRFamide increases synaptic current in muscle fiber 6, indicating increased transmitter release from presynaptic axons.…”

Cardiac Contractility Structure-Activity Relationship and Ligand-Receptor Interactions; the Discovery Of Unique and Novel Molecular Switches in Myosuppressin Signaling

“…Myosuppressins are members of a family of peptides with an identical C-terminal RF-NH 2 , however, the N-terminal extension is unique and differs in length and sequence [ 4 ]. The identical C terminus and variant N terminus are both important in binding and activating signaling pathways [ 5 , 6 ]. FMRF-NH 2 , the first RF-NH 2 -containing peptide isolated, was identified from a neural extract applied to a clam heart preparation [ 7 ].…”