申请人:President and Fellows of Harvard College
公开号:US20130122535A1
公开(公告)日:2013-05-16
Synthesis of many proteins is tightly controlled at the level of translation and plays an essential role in fundamental processes such as cell growth and proliferation, signaling, differentiation or death. Methods that allow imaging and identification of nascent proteins allow for dissecting regulation of translation, both spatially and temporally, including in whole organisms. Described herein are robust chemical methods for imaging and affinity-purifying nascent polypeptides in cells and in animals, based on puromycin analogs. Puromycin analogs of the present invention form covalent conjugates with nascent polypeptide chains, which are rapidly turned over by the proteasome and can be visualized and specifically captured by a bioorthogonal reaction (e.g., [3+2]cycloaddition). The methods of the present invention have broad applicability for imaging protein synthesis and for identifying proteins synthesized under various physiological and pathological conditions in vivo.
许多蛋白质的合成在翻译水平上受到严格的控制,并在细胞生长和增殖、信号传递、分化或死亡等基本过程中发挥着重要作用。允许成像和鉴定新生蛋白质的方法可以在整个生物体内解剖翻译调控的空间和时间,包括在细胞和动物中。本文介绍了一种基于普鲁霉素类似物的成像和亲和纯化新生多肽的强大化学方法。本发明的普鲁霉素类似物与新生多肽链形成共价结合物,这些共价结合物被蛋白酶体迅速降解,并可以通过生物正交反应(例如[3+2]环加成)进行可视化和特异性捕获。本发明的方法具有广泛的适用性,可用于成像蛋白质合成并在体内识别在各种生理和病理条件下合成的蛋白质。