Conformational Control of Integrin-Subtype Selectivity in isoDGR Peptide Motifs: A Biological Switch
Conformational Control of Integrin-Subtype Selectivity in isoDGR Peptide Motifs: A Biological Switch
复制标题
DOI：
10.1002/anie.201004363
复制
发表时间：
2010-01-01
影响因子：
16.6
通讯作者：
Kessler, Horst
中科院分区：
文献类型：
--
作者：
Frank, Andreas O.;Otto, Elke;Kessler, Horst
天冬酰胺重排为异天冬氨酸是多肽合成中众所周知的(不想要的)副反应，通常会导致结构失去生物活性(方案1)。[2]异天冬氨酸的形成在体内也会发生，可能导致蛋白质功能的丧失。因此，这个过程被认为是一个限制蛋白质寿命的生化钟。最近发现，细胞外基质(ECM)蛋白纤维连接蛋白(FN)中的ASN-Gly-Arg(NGR)基序脱酰胺化为等DGR，通过为整合素创建新的黏附结合位点而导致蛋白质功能的增强。基于IsoDGR基序，我们提出了高度活性的环化五肽，选择性结合密切相关的αvβ3和α5β1整合素。整合素是参与基本生物学过程的细胞黏附受体。[8]肽序列精氨酸-甘氨酸-天冬氨酸(RgD)是促进整合素介导的细胞与细胞外基质蛋白(如FN、Vitronectin和Fb)黏附的最主要基序。[9]FN中的RGD三肽至少由四种不同的整合素(α5β1、αvβx、α8β1和αIIbβ3)识别。[10]在小鼠中，第10型III型重复FN模块(见支持信息中的图SI_1)中的RGD序列突变为RGE会破坏整合素与突变基序的结合。[6]有趣的是，尽管存在这种结合缺陷，含有RGE突变体的FN仍然可以通过αvβ3整合素组装成FN纤维[6]。因此，有人认为，从第5类重复I FN模块(见支持信息中的图SI_1)中的NGR序列产生的等DGR基序是新的整合素结合位点。[6]然而，最近一份使用NgR基序突变的重组FN的报告对这一概念提出了质疑。[11]尽管两个关键的FN受体αvβ3和α5β1具有相同的配体识别基序，但它们的功能并不多余。当FN结合时，它们会诱导不同的细胞信号和行为，这对伤口愈合、血管生成和癌症转移等许多生理和病理生理条件都是重要的。[12]为了证明NgR脱酰胺为等DGR产生αvβ3整合素从头结合表位的假设，我们研究了限制性等DGR多肽与整合素亚型的亲和力。此外，受rgd序列的构象控制αvβ3和血小板整合素αIIbβ3之间的选择性的发现的启发，[13]我们测试了这些多肽与紧密相关的整合素αvβ3和α5β1的选择性结合。
The rearrangement of asparagine into isoaspartate is a wellknown (unwanted) side reaction in peptide synthesis,[1] which usually results in structures that lose biological activity (Scheme 1).[2] Isoaspartate formation occurs also invivo, potentially leading to a loss of protein function. Therefore, this process has been proposed to be a biochemical clock that limits protein lifetimes.[4] In contrast, Curnis etal. have recently shown that deamidation of the Asn-Gly-Arg (NGR) motif in the extracellular matrix (ECM) protein fibronectin (FN) into isoDGR results in a gain of protein function by creating a new adhesion binding site for integrins.[5, 6] Here, based on the isoDGR motif we present highly active head-to-tail-cyclized pentapeptides selective for the closely related αvβ3 and α5β1 integrins which have been identified by the study of spatial screening libraries [7] in vitro and in cellular assays.Integrins are cell adhesion receptors that are involved in fundamental biological processes.[8] The peptide sequence Arg-Gly-Asp (RGD) is the most prominent motif to promote integrin-mediated cell adhesion to ECM proteins such as FN, vitronectin, and fibrinogen.[9] The RGD tripeptide in FN is recognized by at least four different integrins (α5β1, αvβx, α8β1, and αIIbβ3), leading to the assembly of an FN matrix around cells.[10] A mutation of the RGD sequence in the 10th type III repeat FN module (see Figure SI_1 in the Supporting Information) to RGE in mice abrogates integrin binding to the mutant motif.[6] Interestingly, despite this binding defect, FN containing the RGE mutant can still be assembled into FN fibrils [6] via αvβ3 integrin. Hence, it was suggested that the isoDGR motif, generated from the NGR sequences in the 5th type repeat I FN module (see Figure SI_1 in the Supporting Information), serves as novel integrin binding site.[6] However, this notion has recently been questioned in a report using recombinant FN with mutations in the NGR motif.[11] Even though the two key FN receptors αvβ3 and α5β1 share the same ligand-recognition motif, their function is not redundant. Upon FN binding they induce different cellular signals and behaviors, which is important for many physiological and pathophysiological conditions such as wound healing, angiogenesis, and cancer metastasis.[12] To prove the hypothesis that deamidation of NGR into isoDGR generates de novo binding epitopes for the αvβ3 integrin, we studied constrained isoDGR peptides for their affinities for integrin subtypes. In addition, inspired by our previous findings that the conformation of the RGD sequence controls the selectivity between αvβ3 and the platelet integrin αIIbβ3,[13] we tested these peptides for selective binding to the closely related integrins αvβ3 and α5β1. For this purpose, we created different libraries of head-to-tail-cyclized pentapep-