Rational optimization of a short human P-selectin-binding peptide leads to nanomolar affinity antagonists
Rational optimization of a short human P-selectin-binding peptide leads to nanomolar affinity antagonists
复制标题
DOI：
10.1074/jbc.m209267200
复制
发表时间：
2003-03-21
影响因子：
4.8
通讯作者：
Biessen, EAL
中科院分区：
文献类型：
--
作者：
Appeldoorn, CCM;Molenaar, TJM;Biessen, EAL
P-选择素在动脉粥样硬化和血栓形成等多种疾病的发生发展中起着重要作用。在我们的实验室中，我们最近发现了一些含有Glu-Trp-Val-Asp-Val共识基序的人P-选择素结合肽，显示出对P-选择素的低微摩尔亲和力(IC50=2 gm)。为了寻找更有效的P-选择素拮抗剂，我们通过两步组合化学方法优化了EWVDV五肽核心基序。通过在基序的N端和C端引入7个取代基，构建了一个专门的肽衍生物文库。特别是，N端带有没食子酸或1，3，5-苯三羧酸取代基的五肽对P-选择素结合的抑制作用明显强于亲本多肽。在从核心序列中去除N-末端谷氨酸后，合成了第二个文库，以绘制导致P-选择素结合增强的没食子酸或1，3，5-苯三羧酸基中的化学部分。此外，通过改变连接基团的长度和刚性，进一步优化了N-端取代基的空间取向。结合使用噬菌体展示和随后的组合。化学导致了一些在静态和动态条件下(IC50=15.4 nM)对P-选择素具有低纳摩尔亲和力的含没食子酸的多肽的设计。这些合成的小分子拮抗剂与天然配体P-选择素糖蛋白配体-1同样有效，有望成为抗动脉粥样硬化血栓治疗的先导。
P-selectin plays an important role in the development of various diseases, including atherosclerosis and thrombosis. In our laboratory we recently identified a number of specific human P-selectin-binding peptides containing a Glu-Trp-Val-Asp-Val consensus motif, displaying a low micromolar affinity for P-selectin (IC50 = 2 gm). In search of more potent antagonists for P-selectin, we have optimized the EWVDV pentapeptide core motif via a two-step combinatorial chemistry approach. A dedicated library of peptide derivatives was generated by introducing seven substituents at the N and C termini of the motif. In particular, pentapeptides with gallic acid or 1,3,5-benzenetricarboxylic acid substituents at the N terminus proved to be considerably more potent inhibitors of P-selectin binding than the parental peptide. After removal of the N-terminal glutamic acid from the core sequence, which appeared to be replaceable by a carboxamide function without loss of affinity, a second library was synthesized to map the chemical moieties within the gallic acid or 1,3,5-benzenetricarboxyl acid groups responsible for the enhanced P-selectin binding. Moreover, by varying the length and rigidity of the connective spacer, we have further optimized the spatial orientation of the N-terminal substituent. The combined use of phage display and subsequent combinatorial. chemistry led to the design of a number of gallic acid-containing peptides with low nanomolar affinity for P-selectin both under static and dynamic conditions (IC50 = 15.4 nM). These small synthetic antagonists, which are equally as potent as the natural ligand P-selectin glycoprotein ligand-1, are promising leads in anti-atherothrombotic therapy.