Molecular transporter between polymer platforms: Highly efficient chemoenzymatic glycopeptide synthesis by the combined use of solid-phase and water-soluble polymer supports
Molecular transporter between polymer platforms: Highly efficient chemoenzymatic glycopeptide synthesis by the combined use of solid-phase and water-soluble polymer supports
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DOI：
10.1002/anie.200463065
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发表时间：
2005-01-01
影响因子：
16.6
通讯作者：
Nishimura, SI
中科院分区：
文献类型：
--
作者：
Fumoto, M;Hinou, H;Nishimura, SI
聚合物负载合成是一种实用方便的合成方法，因为它简化了最终产物的纯化，使组合工艺变得可行。固相聚合物的化学合成使核苷酸（DNA/RNA）和肽（蛋白质）的自动合成成为可能[1,2]，它们现在是研究基因组和蛋白质的功能作用以及开发各种治疗试剂不可或缺的设备。然而，糖缀合物的化学合成比核酸或多肽的合成要困难得多，因为糖缀合物的复杂结构需要极其耗时和繁琐的区域选择性保护和立体选择性糖基化反应过程。尽管固相化学合成的进步使得构建多种低聚糖成为可能，但这些方法对于一般生物化学家或医学科学家来说，仍然存在目标结构的限制和技术难度。由于糖基化反应具有立体化学和区域选择性的特异性，酶促合成是替代化学合成复合寡糖的一种潜在方法然而，固定在固体载体上的糖基受体底物在实际合成中的效率和通用性方面不适合这些酶促反应。在我们酶促合成的研究过程中，基于糖附着的水溶性聚合物作为多价受体底物的簇效应，我们的兴趣一直集中在糖肽作为细胞识别的重要信号分子的有效合成上为了实现基于化学和酶结合策略的协调和高效的糖肽合成，我们认为一种结合固相肽合成和液相碳水化合物合成的合适方法的出现是非常可取的在此，我们报告了一种快速高效合成糖肽的新策略，该策略通过使用一种方便的“分子转运体”来连接两种不同的聚合物载体。我们的综合策略如下图1所示：a)以分子转运蛋白1为末端的光敏o - glcnac肽的固相合成，b)从树脂中脱保护并释放转运蛋白，c)使用含烷氧基氨基官能团的水溶性聚合物对携带糖肽引物的分子转运蛋白进行化学选择性印迹，b[8] d)糖基转移酶的一锅糖延伸，e)通过光选择性切割反应从聚合物平台上的转运体释放全长糖肽。分子转运体1是由l -脯氨酸作为目标肽的n端残基，通过具有光敏连接基团4-[4-（1-羟乙基）-2-甲氧基-5-硝基苯氧基]丁酸[9]修饰的活性酮基合成的（见支持信息）这种异双功能连接体作为两种不同聚合物平台之间的转运体，并允许糖肽的化学选择性印迹和光选择性切割（捕获和释放）（图1，步骤c和e）。我们通过构建一个以sialyl LewisX四糖残基(7)为模型化合物的十二肽来证明我们方法的可行性。方案1采用fmoc保护氨基酸（Fmoc-AA）和Fmoc-Ser (Ac3GlcNAcβ)-OH -[11]，在Fmoc-Arg (Pbf)-NovaSynTGA树脂上固相合成了携带中间糖肽(3)的转运体分子接下来，转运体上的中间体(3…
Polymer-supported synthesis is a practical and convenient method because it simplifies purification of the final products and makes combinatorial processes feasible. Chemical synthesis on solid-phase polymers made the automated synthesis of nucleotides (DNA/RNA) and peptides (proteins) possible,[1, 2] and they are now indispensable devices for the investigation of the functional roles of genomes and proteins as well as the development of a variety of therapeutic reagents. Chemical synthesis of glycoconjugates, however, is a much more difficult task than the synthesis of nucleic acids or polypeptides because complex structures of glycoconjugates require extremely time-consuming and tedious procedures of regioselective protection and stereoselective glycosylation reactions. Although progress in solid-phase chemical synthesis meanwhile allowed the construction of a variety of oligosaccharides,[3] these methods still entail limitation of the target structures and technical difficulty for general biochemists or medical scientists. Enzymatic synthesis is a potential alternative to the chemical synthesis of complex oligosaccharides because of the specificities of both the stereochemistry and regioselectivity in the glycosylation reactions.[4] However, glycosyl acceptor substrates immobilized on solid supports are not suited for these enzymatic reactions in terms of efficiency and versatility in practical synthesis. In the course of our studies of enzymatic synthesis, based on the cluster effect [5] of sugar-attached water-soluble polymers as multivalent acceptor substrates,[6] our interest has been focused on the efficient synthesis of glycopeptides as important signal molecules in cellular recognition.[7] To achieve a concerted and efficient glycopeptide synthesis based on a combined chemical and enzymatic strategy, we thought that the advent of an appropriate methodology to combine solid-phase peptide synthesis and liquid-phase carbohydrate synthesis was highly desirable.[4] Herein, we report a novel strategy of rapid and efficient synthesis of glycopeptides by using a convenient “molecular transporter” that interfaces two different polymer supports. Our synthetic strategy is summarized in Figure1 as follows: a) solid-phase synthesis of the photosensitive O-GlcNAc-peptides terminated by the molecular transporter 1, b) deprotection and release of the transporter from the resin, c) chemoselective blotting of the molecular transporter that carries glycopeptide primers using a water-soluble polymer with alkoxyamino functional groups,[8] d) one-pot sugar elongation with glycosyltransferases, and e) release of fulllength glycopeptides from the transporter on the polymer platform with a photoselective cleavage reaction. The molecular transporter 1 was synthesized from L-proline, which was to be used as the N-terminal residue of the target peptide, by modification with a reactive ketone group with a photolabile linker moiety, 4-[4-(1-hydroxyethyl)-2-methoxy-5-nitrophenoxy] butyric acid [9](see the Supporting Information).[10] This heterobifunctional linker acts as a transporter between the two different polymer platforms and allows both chemoselective blotting and photoselective cleavage (catch and release) of glycopeptides (Figure 1, steps c and e). We demonstrated the feasibility of our method by constructing a dodecapeptide with a sialyl LewisX tetrasaccharide residue (7) as a model compound. Scheme 1 shows the solid-phase synthesis of a transporter molecule carrying an intermediate glycopeptide (3) on a Fmoc-Arg (Pbf)-NovaSynTGA resin using Fmoc-protected amino acids (Fmoc-AA) and Fmoc-Ser (Ac3GlcNAcβ)-OH.[11] Next, the intermediate on the transporter (3 …