Structure-activity relationships of a series of analogues of the octadecaneuropeptide ODN on calcium mobilization in rat astrocytes.
The octadecaneuropeptide ODN (QATVGDVNTDRPGLLDLK), originally characterized as an endogenous ligand for central-type benzodiazepine receptors, increases intracellular calcium concentration ([Ca2+]i) in rat astroglial cells. A series of ODN analogues was synthesized, and each compound was studied for its ability to induce Ca2+ mobilization in cultured rat astrocytes. Replacement of each amino acid by an L-alanine residue (AlaScan) showed that the N-terminal region of the molecule was relatively tolerant to alanine substitution (2-8, 10), except for the Ala9-substituted analogue (9) which was totally devoid of activity. Pyroglutamization (21) and acetylation (22) of the Gln1 residue reduced the Ca2+ response suggesting that a free N-terminal amine function is required for full activity of ODN. Alanine substitution of the residues in the C-terminal region of the molecule (11-14, 16-18) significantly reduced the biological activity of ODN. In particular, modifications of the Leu15 residue (15, 20) abolished the Ca2+-mobilizing activity. The analogues [Ala9]ODN (9), [Ala15]ODN (15), [D-Thr9]ODN (19), and [D-Leu15]ODN (20) partially antagonized the Ca2+ response evoked by ODN. Most importantly, the octapeptide ODN11-18 (OP, 24) produced a dose-response curve that was superimposable to that obtained with ODN, indicating that the C-terminal region of the molecule possesses full biological activity. Finally, the AlaScan of OP revealed that replacement of the Leu5 residue by Ala (29) or D-Leu (33) totally suppressed the calcium response, confirming the crucial contribution of the Leu15 residue of ODN to the biological activity of the neuropeptide.
Leprince
J
J
European Institute for Peptide Research (IFRMP n degrees 23), Laboratory of Cellular and Molecular Neuroendocrinology, INSERM U413, France.
Gandolfo
P
P
Thoumas
J L
JL
Patte
C
C
Fauchère
J L
JL
Vaudry
H
H
Tonon
M C
MC
D000818:Animals; D001253:Astrocytes / Q000187:drug effects* / Q000378:metabolism; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D026261:Diazepam Binding Inhibitor; D058785:GABA-A Receptor Agonists; D058787:GABA-A Receptor Antagonists; D008024:Ligands; D009479:Neuropeptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D009842:Oligopeptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D010446:Peptide Fragments; D051381:Rats; D017208:Rats, Wistar; D013329:Structure-Activity Relationship
D026261:Diazepam Binding Inhibitor; D058785:GABA-A Receptor Agonists; D058787:GABA-A Receptor Antagonists; D008024:Ligands; D009479:Neuropeptides; D009842:Oligopeptides; D010446:Peptide Fragments; C067529:diazepam binding inhibitor (33-50); D002118:Calcium