A Novel Conformation in a Highly Potent, Constrained Gonadotropin-Releasing Hormone Antagonist

Through design, synthesis, and biological testing of constrained
gonadotropin releasing hormone (GnRH)
antagonists, we are studying the structural requirements for biological
activity. Here we describe the conformational
analysis in solution of a highly potent, dicyclic GnRH antagonist,
dicyclo(4-10/5,5‘-8)[Ac-d-2Nal1,d-pClPhe2,d-3Pal3,Asp4,Glu5(Gly),d-Arg6,Dbu8,Dpr10]GnRH
(1), using NMR spectroscopy. The dicyclic part of this
molecule
adopts a preferred conformation containing a type II β turn around
residues 5−6, nested with a type I‘ β turn around
residues 6−7, and a type II β-turn-like structure involving residue
9 and the side chain of residue 10, which is
stabilized by hydrogen bonds between Leu7
NH/Asp4 CO, Dbu8
NHδ/Glu5 CO, and Dpr10
NHγ/Dbu8 CO. This is
a novel conformation that had not been observed previously in any
constrained GnRH antagonist and is remarkably
different from that found for another dicyclic (4-10/5-8) GnRH
antagonist with very similar sequence, dicyclo(4-10/5-8)[Ac-d-2Nal1,d-pClPhe2,d-Trp3,Asp4,Glu5,d-Arg6,Lys8,Dpr10]GnRH
(2) (Bienstock et al. J. Med. Chem.
1993,
36, 3265−3273). The conformation of 2
contains a type II‘ β turn around residues 6−7, which had been
proposed
to be essential for GnRH activity. These results are important for
our general understanding of polypeptide
conformation, since they show that the dicyclo(4-10/5-8) backbone
can adopt more than one family of conformations
despite its dicyclic nature, and from the point of view of the design
of GnRH antagonists, since they suggest that the
presence of a turn around residues 6−7, rather than the type of β
turn, may be necessary for biological activity.

本研究通过对受限促性腺激素释放激素（gonadotropin releasing hormone, GnRH）拮抗剂开展设计、合成与生物学测试，旨在阐明其生物学活性的结构必需条件。本研究采用核磁共振波谱法（nuclear magnetic resonance spectroscopy, NMR），对一种高效双环GnRH拮抗剂dicyclo(4-10/5,5'-8)[Ac-d-2Nal¹,d-pClPhe²,d-3Pal³,Asp⁴,Glu⁵(Gly),d-Arg⁶,Dbu⁸,Dpr¹⁰]GnRH（1）在溶液中的构象进行分析。该分子的双环骨架呈现优势构象：在第5-6位残基处形成II型β转角（type II β turn），嵌套有第6-7位残基处的I’型β转角（type I’ β turn），以及涉及第9位残基与第10位残基侧链的类II型β转角结构（type II β-turn-like structure）；该构象可通过下述氢键网络得以稳定：Leu⁷的NH与Asp⁴的CO、Dbu⁸的NHδ与Glu⁵的CO、Dpr¹⁰的NHγ与Dbu⁸的CO之间形成氢键。该构象为全新构象，此前在任何受限GnRH拮抗剂中均未被观测到，且与另一序列高度相似的双环(4-10/5-8)GnRH拮抗剂dicyclo(4-10/5-8)[Ac-d-2Nal¹,d-pClPhe²,d-Trp³,Asp⁴,Glu⁵,d-Arg⁶,Lys⁸,Dpr¹⁰]GnRH（2）的构象存在显著差异（Bienstock等人，《药物化学杂志》，1993年，第36卷，3265-3273页）。拮抗剂2的构象在第6-7位残基处包含II’型β转角（type II’ β turn），该转角曾被认为是GnRH活性所必需的结构。这些研究结果对于我们全面理解多肽构象（polypeptide conformation）具有重要意义：研究表明，尽管双环(4-10/5-8)骨架具有双环结构特性，却可容纳不止一类构象；同时从GnRH拮抗剂的设计角度而言，本研究提示，第6-7位残基处存在转角，而非特定类型的β转角，或许才是其具备生物学活性的必要条件。