Ghrelin-like peptide with fatty acid modification and O-glycosylation in the red stingray, Dasyatis akajei

BACKGROUND: Ghrelin (GRLN) is now known to be an appetite-stimulating and growth hormone (GH)-releasing peptide that is predominantly synthesized and secreted from the stomachs of various vertebrate species from fish to mammals. Here, we report a GRLN-like peptide (GRLN-LP) in a cartilaginous fish, the red stingray, Dasyatis akajei. RESULTS: The purified peptide contains 16 amino acids (GVSFHPQPRS(10)TSKPSA), and the serine residue at position 3 is modified by n-octanoic acid. The modification is the characteristic of GRLN. The six N-terminal amino acid residues (GVSFHP) were identical to another elasmobranch shark GRLN-LP that was recently identified although it had low identity with other GRLN peptides. Therefore, we designated this peptide stingray GRLN-LP. Uniquely, stingray GRLN-LP was O-glycosylated with mucin-type glycan chains [N-acetyl hexosamine (HexNAc)(3 )hexose(Hex)(2)] at threonine at position 11 (Thr-11) or both serine at position 10 (Ser-10) and Thr-11. Removal of the glycan structure by O-glycanase made the in vitro activity of stingray GRLN-LP decreased when it was evaluated by the increase in intracellular Ca(2+ )concentrations using a rat GHS-R1a-expressing cell line, suggesting that the glycan structure plays an important role for maintaining the activity of stingray GRLN-LP. CONCLUSIONS: This study reveals the structural diversity of GRLN and GRLN-LP in vertebrates.