Takai Tomoyo, Takaya Takao, Nakano Mutsuko, Akutsu Hideo, Nakagawa Atsushi, Aimoto Saburo, Nagai Katsuya, Ikegami Takahisa
Institute for Protein Research, Osaka University, Yamadaoka 3-2, Osaka, 565-0871, Japan.
J Pept Sci. 2006 Jul;12(7):443-54. doi: 10.1002/psc.747.
Orexins-A and B, also called hypocretins-1 and 2, respectively, are neuropeptides that regulate feeding and sleep-wakefulness by binding to two orphan G protein-coupled receptors named orexin-1 (OX(1)R) and orexin-2 (OX(2)R). The sequences and functions of orexins-A and B are similar to each other, but the high sequence homology (68%) is limited in their C-terminal half regions (residues 15-33). The sequence of the N-terminal half region of orexin-A (residues 1-14), containing two disulfide bonds, is very different from that of orexin-B. The structure of orexin-A was determined using two-dimensional homonuclear and (15)N and (13)C natural abundance heteronuclear NMR experiments. Orexin-A had a compact conformation in the N-terminal half region, which contained a short helix (III:Cys6-Gln9) and was fixed by the two disulfide bonds, and a helix-turn-helix conformation (I:Leu16-Ala23 and II:Asn25-Thr32) in the remaining C-terminal half region. The C-terminal half region had both hydrophobic and hydrophilic residues, which existed on separate surfaces to provide an amphipathic character in helices I and II. The nine residues on the hydrophobic surface are also well conserved in orexin-B, and it was reported that the substitution of each of them with alanine resulted in a significant drop in the functional potency at the receptors. Therefore, we suggest that they form the surface responsible for the main hydrophobic interaction with the receptors. On the other hand, the residues on the hydrophilic surface, together with the hydrophilic residues in the N-terminal half region that form a cluster, are known to make only small contributions to the binding to the receptors through similar alanine-scan experiments. However, since our structure of orexin-A showed that large conformational and electrostatical differences between orexins-A and B were rather concentrated in the N-terminal half regions, we suggest that the region of orexin-A is important for the preference for orexin-A of OX(1)R.
食欲素-A和食欲素-B，也分别称为下丘脑泌素-1和下丘脑泌素-2，是通过与两种名为食欲素-1（OX(1)R）和食欲素-2（OX(2)R）的孤儿G蛋白偶联受体结合来调节进食和睡眠-觉醒的神经肽。食欲素-A和食欲素-B的序列和功能彼此相似，但它们C端半区（第15 - 33位残基）的高序列同源性（68%）是有限的。食欲素-A的N端半区（第1 - 14位残基）序列包含两个二硫键，与食欲素-B的序列非常不同。食欲素-A的结构通过二维同核以及（15）N和（13）C天然丰度异核NMR实验确定。食欲素-A在N端半区具有紧密构象，其中包含一个短螺旋（III：Cys6 - Gln9）并由两个二硫键固定，在其余的C端半区具有螺旋-转角-螺旋构象（I：Leu16 - Ala23和II：Asn25 - Thr32）。C端半区既有疏水残基又有亲水残基，它们存在于不同的表面，在螺旋I和II中提供两亲性特征。疏水表面上的九个残基在食欲素-B中也高度保守，据报道，将它们中的每一个用丙氨酸替代会导致受体功能效力显著下降。因此，我们认为它们形成了与受体主要疏水相互作用的表面。另一方面，通过类似的丙氨酸扫描实验已知，亲水表面上的残基以及在N端半区形成簇的亲水残基对与受体的结合仅起很小的作用。然而，由于我们的食欲素-A结构表明食欲素-A和食欲素-B之间的大的构象和静电差异相当集中在N端半区，我们认为食欲素-A的该区域对于OX(1)R对食欲素-A的偏好很重要。
