遗传密码扩充技术及其在蛋白质功能研究及标记成像中的应用

Genetic code expansion and its application in characterization and imaging of proteins

遗传密码扩充(genetic code expansion,GCE)技术利用终止密码子将非天然氨基酸掺入到蛋白质中,再结合点击反应对蛋白质实现定点标记。相较于荧光蛋白、标签抗体等其他标记工具,该技术在蛋白标记中使用的化合物分子较小、对蛋白空间结构影响较小,且能通过点击反应实现蛋白分子与染料分子1︰1的化学计量比,从而能够依据荧光强度对蛋白质定量。因此,在活细胞单分子追踪和超分辨率显微成像等需要细胞长时间暴露在高激光功率下的研究中,GCE技术具有极大的优势。同时,该技术也为提高活细胞成像过程中的定位精度和分子计数准确度奠定了基础。文中旨在总结近年来GCE技术在蛋白质研究中的应用进展,特别是在蛋白质标记成像方面的应用进展。

Genetic code expansion(GCE)allows the incorporation of unnatural amino acids into proteins via using stop codons.GCE may achieve site-specific labeling of proteins in combination with the click reaction.Compared with other labeling tools such as fluorescent proteins and tagged antibodies,the compound molecules used in protein labeling by GCE technology are smaller,and therefore,may less interfere the conformational structure of proteins.In addition,through click reaction,GCE allows a 1:1 stoichiometric ratio of the target protein molecule and the fluorescent dye,and the protein can be quantified based on the fluorescence intensity.Thus,GCE technology has great advantages in the researches that require the exposition of living cells under high laser power for longer time,for example,in the context of single molecule tracing and super-resolution microscopic imaging.Meanwhile,this technology lays the foundation for improving the accuracy of positioning and molecule counting in the imaging process of living cells.This review summarized the GCE technology and its recent applications in functionally characterizing,labeling and imaging of proteins.