Fully convergent chemical synthesis of ester insulin: determination of the high resolution X-ray structure by racemic protein crystallography.
Fully convergent chemical synthesis of ester insulin: determination of the high resolution X-ray structure by racemic protein crystallography.
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DOI：
10.1021/ja311408y
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发表时间：
2013-02-27
影响因子：
15
通讯作者：
Kent, Stephen B. H.
中科院分区：
文献类型：
--
作者：
Avital-Shmilovici, Michal;Mandal, Kalyaneswar;Gates, Zachary P.;Phillips, Nelson B.;Weiss, Michael A.;Kent, Stephen B. H.
胰岛素的高效全合成对于将药物化学应用于优化这一重要蛋白质分子的性质非常重要。最近，我们描述了一种新的形式的胰岛素，即胰岛素原的35个残基C肽的功能被一个共价键取代的一种新型胰岛素，它是有效合成胰岛素的关键中间体。在这里，我们描述了一种完全收敛的合成路线，从三个大小大致相同的未受保护的多肽片段合成胰岛素酯分子。在生理pH条件下，合成酯胰岛素多肽链的折叠速度远快于胰岛素原。单体DKP酯胰岛素的D蛋白和L蛋白对映体均为D-蛋白和L-蛋白对映体。采用全化学合成法合成了[AspB10，LysB28，ProB29]酯胰岛素。用外消旋蛋白质X-射线结晶学测定了合成酯胰岛素分子的原子结构，其分辨率为1.6？[D-DKP酯胰岛素+L-DKP酯胰岛素]外消旋混合物很容易获得衍射性晶体，而单用L-酯胰岛素则不能得到衍射性晶体。对单体DKP酯胰岛素的D蛋白和L蛋白对映体的受体结合进行了测定，并在糖尿病大鼠中皂化转化为相应的DKP胰岛素对映体前后进行了测定。L-DKP酯胰岛素与胰岛素受体结合较弱，而由L-DKP酯胰岛素中间体合成的L-DKP胰岛素与胰岛素受体结合活性强。D-和L-DKP酯胰岛素和D-DKP胰岛素对糖尿病大鼠的降血糖作用不明显，而合成的L-DKP胰岛素在本生物实验中完全具有活性。讨论了酯胰岛素缺乏生物活性的结构基础。
Efficient total synthesis of insulin is important to enable the application of medicinal chemistry to the optimization of the properties of this important protein molecule. Recently we described ‘ester insulin’ – a novel form of insulin in which the function of the 35 residue C-peptide of proinsulin is replaced by a single covalent bond – as a key intermediate for the efficient total synthesis of insulin. Here we describe a fully convergent synthetic route to the ester insulin molecule from three unprotected peptide segments of approximately equal size. The synthetic ester insulin polypeptide chain folded much more rapidly than proinsulin, and at physiological pH. Both the D-protein and L-protein enantiomers of monomeric DKP ester insulin (i.e. [AspB10, LysB28, ProB29]ester insulin) were prepared by total chemical synthesis. The atomic structure of the synthetic ester insulin molecule was determined by racemic protein X-ray crystallography to a resolution of 1.6 Å. Diffraction quality crystals were readily obtained from the racemic mixture of {D-DKP ester insulin + L-DKP ester insulin}, whereas crystals were not obtained from the L-ester insulin alone even after extensive trials. Both the D-protein and L-protein enantiomers of monomeric DKP ester insulin were assayed for receptor binding and in diabetic rats, before and after conversion by saponification to the corresponding DKP insulin enantiomers. L-DKP ester insulin bound weakly to the insulin receptor, while synthetic L-DKP insulin derived from the L-DKP ester insulin intermediate was fully active in binding to the insulin receptor. The D- and L-DKP ester insulins and D-DKP insulin were inactive in lowering blood glucose in diabetic rats, while synthetic L-DKP insulin was fully active in this biological assay. The structural basis of the lack of biological activity of ester insulin is discussed.
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影响因子：
56.9
作者：
DAWSON, PE;MUIR, TW;KENT, SBH
通讯作者：
KENT, SBH
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
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通讯作者：
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影响因子：
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通讯作者：
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影响因子：
5.6
作者：
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通讯作者：
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影响因子：
4.8
作者：
Hua, Qing-xin;Nakagawa, Satoe;Weiss, Michael A.
通讯作者：
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