Structural characterization of the mammalian autophagy conjugation systems Atg12-Atg5 and LC3

Autophagy is a conserved eukaryotic process, where cytoplasmic content is taken up by an autophagosomal vesicle, which is then degraded in the lysosome. Among different autophagy-related proteins and complexes the Atg12-Atg5 and LC3II conjugation systems are crucial for autophagosome formation.

Two ubiquitin-like proteins are involved in autophagy, namely Atg12 and Atg8 (mammalian MAP1LC3). Similar to the canonical ubiquitination pathway these two proteins are activated and conjugated to their respective targets by E1- and E2-like enzymes. These two conjugation systems are essential for autophagosome formation. To obtain the Atg12-Atg5 conjugate and lipidated LC3 for further studies, Atg12/5/7/10 and LC3II/Atg3/7 were co-expressed in insect cells. So far, conjugated Atg12-Atg5 could be purified in small yields and was subjected to mass spectrometric analysis.

This work is supported by the German Research Foundation (Collaborative Research Centre 860).