Turn structures in CGRP C-terminal analogues promote stable arrangements of key residue side chains
Turn structures in CGRP C-terminal analogues promote stable arrangements of key residue side chains
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DOI：
10.1021/bi0102860
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发表时间：
2001-07-17
期刊：
影响因子：
2.9
通讯作者：
Walpole, C
中科院分区：
文献类型：
--
作者：
Carpenter, KA;Schmidt, R;Walpole, C
37 个氨基酸的降钙素基因相关肽 (CGRP) 是一种有效的内源性血管扩张剂，被认为与偏头痛发作的发生有关。因此，CGRP拮抗剂对于治疗偏头痛可能具有治疗价值。 CGRP C 末端衍生肽 [D-31,P-34,F-35]CGRP(27-37)-NH2 最近被鉴定为高亲和力 hCGRP(1) 受体选择性拮抗剂。还证明了多种相关类似物具有合理的 CGRP(1) 亲和力，包括 [D-31,A(34),F-35]CGRP(27-37)-NH2。在此介绍的研究中，探讨了对于 hCGRP(1) 受体结合很重要的 CGRP(27-37)-NH2 类似物的构象和结构特征。对 [D-31,P-34,F-35]CGRP(27-37)-NH2 进行的结构-活性研究得到了 [D-31,P-34,F-35]CGRP(30-37)-NH2，这是报道的最短的 CGRP C 末端肽类似物，表现出合理的 hCGRP(1) 受体亲和力 (K-i = 29.6 nM)。进一步从肽的 N 末端去除 T-30 大大降低了受体亲和力，从纳摩尔到微摩尔范围。通过 [A(34),F-35]CGRP(28-37)-NH2 的丙氨酸扫描，鉴定出被认为对 hCGRP(1) 受体结合至关重要的其他残基，包括 V-32 和 F-37。此外，用羧基取代同一肽中的 C 端酰胺，导致 hCGRP(1) 亲和力降低 50 倍以上，因此表明酰胺部分在受体结合中具有直接作用。通过 NMR 波谱和分子模型检查了两类 CGRP(27-37)-NH2 肽 [D-31,X-34,F-35]CGRP(27-37)-NH2（X 是 A 或 P）的构象特性。以 P-29 为中心的 β 转角是两个系列的活性肽中一致观察到的显着特征。这一转变导致关键的 T-30 残留物暴露于周围环境中。 A(34) 系列中的肽的另外一个特征是稳定的 C 端螺旋转角，导致三个重要残基（T-30、V-32 和 F-37）彼此采用一致的空间位置。尽管大部分 [D-31,P-34,F-35]CGRP(27-37)-NH2 计算的构象异构体包含以 P-34 为中心的 γ 转角，但 P-34 系列中的肽在 C 末端相对更灵活。这些结果共同表明，CGRP(27-37)-NH2 类似物 C 末端和 N 末端的转角结构可能有助于适当定向 hCGRP(1) 受体结合的关键残基（T-30、V-32 和 F-37）。
The 37-amino acid calcitonin gene-related peptide (CGRP) is a potent endogenous vasodilator thought to be implicated in the genesis of migraine attack. CGRP antagonists may thus have therapeutic value for the treatment of migraine. The CGRP C-terminally derived peptide [D-31,P-34,F-35]CGRP(27-37)-NH2 was recently identified as a high-affinity hCGRP(1) receptor selective antagonist. Reasonable CGRP(1) affinity has also been demonstrated for several related analogues, including [D-31,A(34),F-35]CGRP(27-37)-NH2. In the study presented here, conformational and structural features in CGRP(27-37)-NH2 analogues that are important for hCGRP(1) receptor binding were explored. Structure-activity studies carried out on [D-31,P-34,F-35]CGRP(27-37)-NH2 resulted in [D-31,P-34,F-35]CGRP(30-37)-NH2, the shortest reported CGRP C-terminal peptide analogue exhibiting reasonable hCGRP(1) receptor affinity (K-i = 29.6 nM). Further removal of T-30 from the peptide's N-terminus greatly reduced receptor affinity from the nanomolar to micromolar range. Additional residues deemed critical for hCGRP(1) receptor binding were identified from an alanine scan of [A(34),F-35]CGRP(28-37)-NH2 and included V-32 and F-37. Replacement of the C-terminal amide in this same peptide with a carboxyl, furthermore, resulted in a greater than 50-fold reduction in hCGRP(1) affinity, thus suggesting a direct role for the amide moiety in receptor binding. The conformational properties of two classes of CGRP(27-37)-NH2 peptides, [D-31,X-34,F-35]CGRP(27-37)-NH2 (X is A or P), were examined by NMR spectroscopy and molecular modeling. A beta -turn centered on P-29 was a notable feature consistently observed among active peptides in both series. This turn led to exposure of the critical T-30 residue to the surrounding environment. Peptides in the A(34) series were additionally characterized by a stable C-terminal helical turn that resulted in the three important residues (T-30, V-32, and F-37) adopting consistent interspatial positions with respect to one another. Peptides in the P-34 series were comparatively more flexible at the C-terminus, although a large proportion of the [D-31,P-34,F-35]CGRP(27-37)-NH2 calculated conformers contained a gamma -turn centered on P-34. These results collectively suggest that turn structures at both the C-terminus and N-terminus of CGRP(27-37)-NH2 analogues may help to appropriately orient critical residues (T-30, V-32, and F-37) for hCGRP(1) receptor binding.