Luo Yu, Jiang Chen, Yu Lihua, Yang Aimin
School of Life Sciences, Chongqing University, Chongqing, China.
Front Chem. 2020 Apr 3;8:233. doi: 10.3389/fchem.2020.00233. eCollection 2020.
Macroautophagy (hereafter referred to as autophagy) is an evolutionarily conserved lysosomal degradation pathway in all eukaryotic cells, which is critical for maintaining cell homeostasis. A series of autophagy-related (ATG) proteins are involved in the regulation of autophagy. The activities of ATG proteins are mainly modulated by posttranslational modifications (PTMs), such as phosphorylation, lipidation, acetylation, ubiquitination, and sumoylation. To tackle molecular mechanisms of autophagy, more and more researches are focusing on the roles of PTMs in regulation of the activity of ATG proteins and autophagy process. The protein ligation techniques have emerged as powerful tools for the chemical engineering of proteins with PTMs, and provided effective methods to elucidate the molecular mechanism and physiological significance of PTMs. Recently, several ATG proteins with PTM were prepared by protein ligation techniques such as native chemical ligation (NCL), expressed protein ligation (EPL), peptide hydrazide-based NCL, and Sortase A-mediated ligation (SML). More importantly, the synthesized ATG proteins are successfully used to probe the mechanism of autophagy. In this review, we summarize protein ligation techniques for the preparation of ATG proteins with PTMs. In addition, we highlight the biological applications of synthetic ATG proteins to probe the autophagy mechanism.
巨自噬（以下简称自噬）是所有真核细胞中一种进化保守的溶酶体降解途径，对维持细胞内稳态至关重要。一系列自噬相关（ATG）蛋白参与自噬的调控。ATG蛋白的活性主要通过翻译后修饰（PTM）来调节，如磷酸化、脂化、乙酰化、泛素化和类泛素化。为了探究自噬的分子机制，越来越多的研究聚焦于PTM在调控ATG蛋白活性和自噬过程中的作用。蛋白质连接技术已成为对蛋白质进行PTM化学工程改造的有力工具，并为阐明PTM的分子机制和生理意义提供了有效方法。最近，通过天然化学连接（NCL）、表达蛋白连接（EPL）、基于肽酰肼的NCL和分选酶A介导的连接（SML）等蛋白质连接技术制备了几种具有PTM的ATG蛋白。更重要的是，合成的ATG蛋白已成功用于探究自噬机制。在本综述中，我们总结了用于制备具有PTM的ATG蛋白的蛋白质连接技术。此外，我们重点介绍了合成ATG蛋白在探究自噬机制方面的生物学应用。
