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. 2020 Jun 24;9:e57640. doi: 10.7554/eLife.57640

Figure 1. Comparison of the sequences of echinoderm NPY/NPF/PrRP-like peptides with related peptides in other taxa.

(A) Comparison with PrRP-type neuropeptides. Conserved residues are highlighted in black (identical) or grey (conservative substitutions) (B) Comparison with NPY/NPF-type neuropeptides. Conserved residues are highlighted in black (identical) or grey (conservative substitutions). The arrowheads indicate residues that have been shown to be important for the three-dimensional structure of the NPY/NPF-type peptides but which are not present in the echinoderm peptides. The colour coding of phyla is as follows: dark blue (Echinodermata), light blue (Hemichordata), purple (Chordata), orange (Platyhelminthes), red (Lophotrochozoa), yellow (Priapulida), green (Arthropoda), grey (Nematoda). The full names of the species and the accession numbers of the sequences are listed in Figure 1—source data 1.

Figure 1—source data 1. Accession numbers of the precursor sequences used for the peptide alignments in Figure 1.
Accession numbers of the precursor sequences used for the peptide alignments in Figure 1.

Figure 1.

Figure 1—figure supplement 1. Sequencing of the A. rubens NPY/NPF/PrRP-like precursor cDNA and determination of the structure of the neuropeptide derived from the precursor.

Figure 1—figure supplement 1.

(A) Sequence of a cDNA encoding the precursor of a NPY/NPF/PrRP-like peptide in A. rubens. The cDNA sequence (lowercase) comprises an open reading frame of 324 bases that encode a 108-residue protein (uppercase). The predicted signal peptide is shown in blue, the predicted cleavage site is shown in green and the predicted neuropeptide is shown in red, with an N-terminal glutamine that is a potential substrate for pyroglutamation shown in purple and with a C-terminal glycine that is a potential substrate for amidation shown in orange. The cDNA was amplified by PCR from A. rubens radial nerve cord cDNA using primers corresponding to the sequences highlighted in yellow. The cDNA was cloned in the vector pBluescript II SK (+) and the T3 and T7 primers were used for the sequencing. A single nucleotide that differs from a contig sequence (1060225) identified from A. rubens radial nerve cord transcriptome data is highlighted in black, but this is a synonymous substitution. This sequence has been deposited in GenBank under the accession number MK033631.1 (B) Annotated mass spectrum showing the structure of a NPY/NPF/PrRP-like peptide isolated from an A. rubens radial nerve cord extract. The peptide QDRSKAMQAERTGQLRRLNPRF, with Q1-Pyroglutamate (−17.02655 Da) and F22-amidated (−0.98402 Da), was observed at charge state +6, monoisotopic m/z 440.90631 Da with a precursor mass error of 0.12 ppm [MH+ 2640.40148 Da] and with a retention time (RT) of 66.3484 min. The b series of peptide fragment ions are shown in red, the y series in blue and additional identified peptide fragment ions in green. The peptide was identified with: Sequest HT (v1.17); XCorr: 4.27, Percolator q-Value: 0.0e0, Percolator PEP:1.6e-2. The fragment match tolerance used for the search was 0.8 Da. The fragmentation table for this mass spectrum can be found in Figure 1—figure supplement 1—source data 1.
Figure 1—figure supplement 1—source data 1. Fragmentation table of the mass spectrum for the A. rubens neuropeptide ArPrRP (QDRSKAMQAERTGQLRRLNPRF) shown in Figure 1—figure supplement 1B.
F22-amidated (−0.98402 Da), Q1-Pyroglutamate (−17.02655 Da), observed at charge state +6, monoisotopic m/z 440.90631 Da with a precursor mass error of 0.12 ppm).