Characterization and optimization of two-chain folding pathways of insulin via native chain assembly
Characterization and optimization of two-chain folding pathways of insulin via native chain assembly
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DOI：
10.1038/s42004-018-0024-0
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发表时间：
2018-05-03
影响因子：
5.9
通讯作者：
Iwaoka, Michio
中科院分区：
文献类型：
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作者：
Arai, Kenta;Takei, Toshiki;Iwaoka, Michio
直到最近，胰岛素的全合成仍然是一个未解决的挑战，它的特征是由两个链间和一个链内二硫键(SS)桥连接的异二聚体结构。通过优化这两条链的合成和定向二硫键交联，并应用仿生单组分胰岛素原方法，实现了胰岛素的高效合成。在这里，我们报告了另一种策略，氧化天然链组装的优化和特征。在该方法中，无保护的A-链和B-链在热力学控制下氧化组装，以39%的产率合成了牛胰腺胰岛素。折叠主要通过结构1SS(STAR)和2SS(STAR)中间体进行，具有共同的链间半胱氨酸(A20)-半胱氨酸(B19)二硫键。这些结果表明，长期以来被认为效率低下的天然链组装可能是获得胰岛素变异体的合理策略。人胰岛素和人II型松弛素产率分别高达49%和47%的合成支持了这一点，尽管应用于人胰岛素Val(A16)变异体并不成功。
Until recently the total synthesis of insulin, with its characteristic heterodimeric structure crosslinked by two interchain and one intrachain disulfide (SS) bridge, remained largely an unsolved challenge. By optimizing the synthesis and directed disulfide crosslinking of the two chains, and by applying biomimetic monocomponent proinsulin approaches, efficient insulin syntheses have been realized. Here we report the optimization and characterisation of an alternative strategy, oxidative native chain assembly. In this method unprotected A- and B-chains assemble oxidatively under thermodynamic control to afford bovine pancreatic insulin in 39% yield. Folding is found to proceed predominantly via structured 1SS(star) and 2SS(star) intermediates with a common interchain Cys(A20)-Cys(B19) disulfide. These results suggest that native chain assembly, long considered inefficient, may represent a reasonable strategy to access insulin variants. This is supported by the synthesis of human insulin and human type-II relaxin in yields of up to 49 and 47%, respectively, although the application to human insulin Val(A16) variant is unsuccessful.