Iterative in situ click chemistry assembles a branched capture agent and allosteric inhibitor for Akt1.
Iterative in situ click chemistry assembles a branched capture agent and allosteric inhibitor for Akt1.
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DOI：
10.1021/ja2064389
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发表时间：
2011-11-16
影响因子：
15
通讯作者：
Heath, James R.
中科院分区：
文献类型：
--
作者：
Millward, Steven W.;Henning, Ryan K.;Kwong, Gabriel A.;Pitram, Suresh;Agnew, Heather D.;Deyle, Kaycie M.;Nag, Arundhati;Hein, Jason;Lee, Su Seong;Lim, Jaehong;Pfeilsticker, Jessica A.;Sharpless, K. Barry;Heath, James R.
我们描述了使用迭代原位点击化学来设计Akt特异性分支肽三配体，其在多种生化测定中是单克隆抗体的直接替代物。分支结构中的每个肽模块对亲和力和/或特异性做出独特贡献，产生200 nM亲和配体，其有效地从癌细胞裂解物中免疫沉淀Akt并标记固定细胞中的Akt。我们在筛选之前使用小分子预抑制Akt导致低微摩尔抑制效力和变构抑制模式，这通过一系列竞争性酶动力学测定来证明。为了证明蛋白质模板原位点击反应的效率和选择性，我们开发了一种新的基于QPCR的方法，能够定量评估其产率。这些结果指出了迭代原位点击化学产生具有新型抑制特性的有效的、合成可及的抗体替代物的潜力
We describe the use of iterative in situ click chemistry to design an Akt-specific branched peptide triligand that is a drop-in replacement for monoclonal antibodies in multiple biochemical assays. Each peptide module in the branched structure makes unique contributions to affinity and/or specificity resulting in a 200 nM affinity ligand that efficiently immunoprecipitates Akt from cancer cell lysates and labels Akt in fixed cells. Our use of a small molecule to pre-inhibit Akt prior to screening resulted in low micromolar inhibitory potency and an allosteric mode of inhibition, which is evidenced through a series of competitive enzyme kinetic assays. To demonstrate the efficiency and selectivity of the protein-templated in situ click reaction, we developed a novel QPCR-based methodology that enabled a quantitative assessment of its yield. These results point to the potential for iterative in situ click chemistry to generate potent, synthetically accessible antibody replacements with novel inhibitory properties
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影响因子：
3.7
作者：
Ostroff RM;Bigbee WL;Franklin W;Gold L;Mehan M;Miller YE;Pass HI;Rom WN;Siegfried JM;Stewart A;Walker JJ;Weissfeld JL;Williams S;Zichi D;Brody EN
通讯作者：
Brody EN
影响因子：
3.7
作者：
Gold L;Ayers D;Bertino J;Bock C;Bock A;Brody EN;Carter J;Dalby AB;Eaton BE;Fitzwater T;Flather D;Forbes A;Foreman T;Fowler C;Gawande B;Goss M;Gunn M;Gupta S;Halladay D;Heil J;Heilig J;Hicke B;Husar G;Janjic N;Jarvis T;Jennings S;Katilius E;Keeney TR;Kim N;Koch TH;Kraemer S;Kroiss L;Le N;Levine D;Lindsey W;Lollo B;Mayfield W;Mehan M;Mehler R;Nelson SK;Nelson M;Nieuwlandt D;Nikrad M;Ochsner U;Ostroff RM;Otis M;Parker T;Pietrasiewicz S;Resnicow DI;Rohloff J;Sanders G;Sattin S;Schneider D;Singer B;Stanton M;Sterkel A;Stewart A;Stratford S;Vaught JD;Vrkljan M;Walker JJ;Watrobka M;Waugh S;Weiss A;Wilcox SK;Wolfson A;Wolk SK;Zhang C;Zichi D
通讯作者：
Zichi D
DOI：
10.1073/pnas.97.17.9367
发表时间：
2000-08-15
影响因子：
11.1
作者：
Erlanson, DA;Braisted, AC;Wells, JA
通讯作者：
Wells, JA
影响因子：
16.6
作者：
Agnew, Heather D.;Rohde, Rosemary D.;Millward, Steven W.;Nag, Arundhati;Yeo, Woon-Seok;Hein, Jason E.;Pitram, Suresh M.;Tariq, Abdul Ahad;Burns, Vanessa M.;Krom, Russell J.;Fokin, Valery V.;Sharpless, K. Barry;Heath, James R.
通讯作者：
Heath, James R.
影响因子：
15
作者：
Meyer, Scott C.;Shomin, Carolyn D.;Ghosh, Indraneel
通讯作者：
Ghosh, Indraneel