biostudies-literatureNguyen NChina Scholarship CouncilHHS | NIH | National Institute of General Medical SciencesHHS | NIH | National Institute of Neurological Disorders and StrokeNINDS NIH HHSNIGMS NIH HHS13584-13600https://www.ebi.ac.uk/biostudies/studies/S-EPMC7521654biostudies-literatureUnknown295(39)During autophagy, LC3 and GABARAP proteins become covalently attached to phosphatidylethanolamine on the growing autophagosome. This attachment is also reversible. Deconjugation (or delipidation) involves the proteolytic cleavage of an isopeptide bond between LC3 or GABARAP and the phosphatidylethanolamine headgroup. This cleavage is carried about by the ATG4 family of proteases (ATG4A, B, C, and D). Many studies have established that ATG4B is the most active of these proteases and is sufficient for autophagy progression in simple cells. Here we examined the second most active protease, ATG4A, to map out key regulatory motifs on the protein and to establish its activity in cells. We utilized fully <i>in vitro</i> reconstitution systems in which we controlled the attachment of LC3/GABARAP mThe Journal of biological chemistryThe insufficiency of ATG4A in macroautophagy.PMC7521654GM100930R01 NS063973T32 GM007223NS063973R01 GM100930201406180015Yu SMelia TJMires ANguyen NOlivas TJLuan LKauffman KJJin JNag SfalseThe insufficiency of ATG4A in macroautophagy.During autophagy, LC3 and GABARAP proteins become covalently attached to phosphatidylethanolamine on the growing autophagosome. This attachment is also reversible. Deconjugation (or delipidation) involves the proteolytic cleavage of an isopeptide bond between LC3 or GABARAP and the phosphatidylethanolamine headgroup. This cleavage is carried about by the ATG4 family of proteases (ATG4A, B, C, and D). Many studies have established that ATG4B is the most active of these proteases and is sufficient for autophagy progression in simple cells. Here we examined the second most active protease, ATG4A, to map out key regulatory motifs on the protein and to establish its activity in cells. We utilized fully <i>in vitro</i> reconstitution systems in which we controlled the attachment of LC3/GABARAP m2020-01-01T00:00:00Z2020 Sep2026-05-02T11:43:30.434Z2022-02-11T16:03:40.625ZS-EPMC75216543273229010.1074/jbc.ra120.01389710.1074/jbc.RA120.013897