Structural Basis of Atg8 Activation by a Homodimeric E1, Atg7

Abstract

E1 enzymes activate ubiquitin-like proteins and transfer them to cognate E2 enzymes. Atg7, a noncanonical E1, activates two ubiquitin-like proteins, Atg8 and Atg12, and plays a crucial role in autophagy. Here, we report crystal structures of full-length Atg7 and its C-terminal domain bound to Atg8 and MgATP, as well as a solution structure of Atg8 bound to the extreme C-terminal domain (ECTD) of Atg7. The unique N-terminal domain (NTD) of Atg7 is responsible for Atg3 (E2) binding, whereas its C-terminal domain is comprised of a homodimeric adenylation domain (AD) and ECTD. The structural and biochemical data demonstrate that Atg8 is initially recognized by the C-terminal tail of ECTD and is then transferred to an AD, where the Atg8 C terminus is attacked by the catalytic cysteine to form a thioester bond. Atg8 is then transferred via a trans mechanism to the Atg3 bound to the NTD of the opposite protomer within a dimer.

Mechanistic Study of Uba5 Enzyme and the Ufm1 Conjugation Pathway

“…Various additional domains in these noncanonical E1s could also likely play a role in Ubl activation and E1-E2 transthiolation. For example, the recent structural studies of ATG7 suggest that the unique N-terminal domain of one monomer recruits the incoming E2 (ATG3) and positions the E2 to react in trans with Ubl (ATG8) activated by the other monomer (40,41).…”

The machinery of macroautophagy

“…The C-terminal tail of the ECTD shows no secondary structure and can interact with Atg8. Biochemical data further show that this interaction is important for recognition of Atg8 by Atg7 [158,159]. Based on the structural and biochemical data, Atg8 is first recognized by the ECTD C-terminal tail of Atg7.…”

“…Subsequently, the catalytic Cys507 of Atg7 forms a thioester bond with Atg8. The crystal structure data reveal that Atg7 functions as a homodimer, in which the monomers interact with each other through their AD [158]. Considering that the NTD of Atg7 is responsible for Atg3 binding, this suggests that Atg8 is activated by one Atg7 and then transferred to Atg3 that is bound to another Atg7 [160].…”

“…The Atg7 monomer is composed of three parts: NTD, adenylation domain (AD) and Cterminal Atg7-specific domain (ECTD) [158]. The NTD of Atg7 is highly basic, and Atg3, the cognate E2 enzyme, recognizes this positively charged domain.…”

Autophagy and human diseases

“…Many drugs and compounds that modulate autophagy are currently receiving considerable attention [11,89,108]. These include, for example, autophagy inducers such as the mTORC1 inhibitor rapamycin [109] and its analogues (e.g., CCI-779 [109], RAD001 [110,111], and AP23573 [112]), mTOR kinase inhibitors (e.g., Torin 1 [113], and PP242 [114]), trehalose [115,116], carbamazepine [117], and the newly identified autophagy-inducing peptide Tatbeclin 1 [118]; autophagy inhibitors such as chloroquine [119,120] and hydroxychloroquine [121], Lys05 [122], 3-methyladenine [123] and its derivatives [124], PIK3C3 inhibitors [125], ATG4B inhibitors [126,127], and ATG7 inhibitors [128,129]. Autophagy-modulating drugs that are currently used in clinical trials are summarized in Table 2.…”