Chemical Biology of Autophagy-Related Proteins With Posttranslational Modifications: From Chemical Synthesis to Biological Applications.

Yu Luo,Aimin Yang,Lihua Yu,Chen Jiang

doi:10.3389/fchem.2020.00233

Abstract

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.

Highlights

Macroautophagy, hereafter referred to as autophagy, is an evolutionarily conserved lysosomal degradation pathway that removes cytoplasmic components including protein aggregates, damaged organelles and intracellular pathogens in all eukaryotic cells (Feng et al, 2014; Ohsumi, 2014)
Surface plasmon resonance (SPR) and microscale thermophoresis (MST) were used to quantify the binding affinity between Atg3 and Atg8, and the results showed that acetylation did not alter the interaction between Atg3 and Atg8 in vitro
Autophagy plays an important role in maintaining cell homeostasis and regulating cellular energy metabolism

Summary

INTRODUCTION

Macroautophagy, hereafter referred to as autophagy, is an evolutionarily conserved lysosomal degradation pathway that removes cytoplasmic components including protein aggregates, damaged organelles and intracellular pathogens in all eukaryotic cells (Feng et al, 2014; Ohsumi, 2014). Microtubule-associated protein light chain 3 (LC3), a mammalian homolog of yeast Atg, is involved in both biogenesis of autophagosomes and recruitment of autophagic cargos. The activities of ATG proteins are mainly modulated by posttranslational modifications (PTMs), such as phosphorylation, lipidation, acetylation, ubiquitination, and sumoylation (Figure 1) (Wani et al, 2015; Xie et al, 2015). These modifications play an important role in controlling the fate of the ATG proteins and the process of autophagy. Uncovering the roles of PTMs in regulating the function of ATG proteins is crucial for understanding the mechanisms of autophagy. Protein ligation techniques for the preparation of ATG proteins are summarized and biological application of synthetic ATG proteins for the elucidation of autophagy mechanism are highlighted

PROTEIN LIGATION

CHEMICAL SYNTHESIS OF ATG PROTEINS

PROBING THE AUTOPHAGY USING SYNTHETIC ATG PROTEINS

CONCLUSIONS AND PERSPECTIVES

AUTHOR CONTRIBUTIONS