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Article

NMR Analysis of Caenorhabditis elegans FLP-18 Neuropeptides: Implications for NPR-1 Activation^†

Supporting Info

Aaron T. Dossey,^‡^§ Vincenzina Reale,^§ Heather Chatwin, Cherian Zachariah,^‡ Mario deBono, Peter D. Evans, and Arthur S. Edison*^‡

McKnight Brain Institute, University of Florida, 100 South Newell Drive, Building 59, Room LG-150, Gainesville, Florida 32611, The Inositide Laboratory, The Babraham Institute, Cambridge, CB2 4AT, United Kingdom, and The MRC Laboratory of Molecular Biology, Hills Road, Cambridge, CB2 2QH, United Kingdom

Biochemistry, 2006, 45 (24), pp 7586–7597

DOI: 10.1021/bi0603928

Publication Date (Web): May 23, 2006

†

This work was supported by a NSF CAREER grant to A.S.E., NIH 5P41RR016105, and the Human Frontier Science Program.

‡

University of Florida.

These authors contributed equally to this work.

The Babraham Institute.

The MRC Laboratory of Molecular Biology.

To whom correspondence should be addressed: Box 100245, Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, FL 32610-0245. Telephone: 352-392-4535. Fax: 352-392-3422. E-mail: art@mbi.ufl.edu.

Abstract

Phe-Met-Arg-Phe-NH₂ (FMRFamide)-like peptides (FLPs) are the largest neuropeptide family in animals, particularly invertebrates. FLPs are characterized by a C−N-terminal gradient of decreasing amino acid conservation. Neuropeptide receptor 1 (NPR-1) is a G-protein coupled receptor (GPCR), which has been shown to be a strong regulator of foraging behavior and aggregation responses in Caenorhabditis elegans. Recently, ligands for NPR-1 were identified as neuropeptides coded by the precursor genes flp-18 and flp-21 in C. elegans. The flp-18 gene encodes eight FLPs including DFDGAMPGVLRF-NH₂ and EMPGVLRF-NH₂. These peptides exhibit considerably different activities on NPR-1, with the longer one showing a lower potency. We have used nuclear magnetic resonance and biological activity to investigate structural features that may explain these activity differences. Our data demonstrate that long-range electrostatic interactions exist between N-terminal aspartates and the C-terminal penultimate arginine as well as N-terminal hydrogen-bonding interactions that form transient loops within DFDGAMPGVLRF-NH₂. We hypothesize that these loops, along with peptide charge, diminish the activity of this peptide on NPR-1 relative to that of EMPGVLRF-NH₂. These results provide some insight into the large amino acid diversity in FLPs.