Development of an orexin-2 receptor selective agonist, [Ala(11), D-Leu(15)]orexin-B.
Investigation of L-alanine and D-amino acid replacement of orexin-B revealed that three L-leucine residues at the positions of 11, 14, and 15 in orexin-B were important to show selectivity for the orexin-2 receptor (OX(2)) over the orexin-1 receptor (OX(1)). L-Alanine substitution at position 11 and D-leucine substitution at positions 14 and 15 maintained the potency of orexin-B to mobilize [Ca(2+)](i) in CHO cells expressing the OX(2), while their potency for the OX(1) was significantly reduced. In combined substitutions, we identified that [Ala(11), D-Leu(15)]orexin-B showed a 400-fold selectivity for the OX(2) (EC(50)=0.13nM) over OX(1) (EC(50)=52nM). [Ala(11), D-Leu(15)]orexin-B is a beneficial tool for addressing the functional roles of the OX(2).
Asahi
Shuichi
S
Banyu Tsukuba Research Institute, Banyu Pharmaceutical Co., Ltd., Okubo-3, Tsukuba 300-2611, Ibaraki, Japan.
Egashira
Shin-Ichiro
S
Matsuda
Masao
M
Iwaasa
Hisashi
H
Kanatani
Akio
A
Ohkubo
Mitsuru
M
Ihara
Masaki
M
Morishima
Hajime
H
D019943:Amino Acid Substitution; D000818:Animals; D016466:CHO Cells; D020013:Calcium Signaling / Q000187:drug effects; D006224:Cricetinae; D015195:Drug Design; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides / Q000138:chemical synthesis* / Q000737:chemistry / Q000494:pharmacology; D064446:Orexin Receptors; D000068797:Orexins; D017433:Protein Structure, Secondary; D051381:Rats; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide / Q000819:agonists* / Q000378:metabolism
C000596229:HCRT protein, human; D047908:Intracellular Signaling Peptides and Proteins; D009479:Neuropeptides; D064446:Orexin Receptors; D000068797:Orexins; D043562:Receptors, G-Protein-Coupled; D018013:Receptors, Neuropeptide