Structural basis of transcriptional regulation by the HigA antitoxin.

Bacterial toxin-antitoxin systems are important factors implicated in growth inhibition and plasmid maintenance. Type II toxin-antitoxin pairs are regulated at the transcriptional level by the antitoxin itself. Here, we examined how the HigA antitoxin regulates the expression of the Proteus vulgaris higBA toxin-antitoxin operon from the Rts1 plasmid.

RING E3 mechanism for ubiquitin ligation to a disordered substrate visualized for human anaphase-promoting complex.

For many E3 ligases, a mobile RING (Really Interesting New Gene) domain stimulates ubiquitin (Ub) transfer from a thioester-linked E2∼Ub intermediate to a lysine on a remotely bound disordered substrate. One such E3 is the gigantic, multisubunit 1.2-MDa anaphase-promoting complex/cyclosome (APC), which controls cell division by ubiquitinating cell cycle regulators to drive their timely degradation.

Mechanism of polyubiquitination by human anaphase-promoting complex: RING repurposing for ubiquitin chain assembly.

Polyubiquitination by E2 and E3 enzymes is a predominant mechanism regulating protein function. Some RING E3s, including anaphase-promoting complex/cyclosome (APC), catalyze polyubiquitination by sequential reactions with two different E2s. An initiating E2 ligates ubiquitin to an E3-bound substrate.

Structure of a RING E3 trapped in action reveals ligation mechanism for the ubiquitin-like protein NEDD8.

Most E3 ligases use a RING domain to activate a thioester-linked E2∼ubiquitin-like protein (UBL) intermediate and promote UBL transfer to a remotely bound target protein. Nonetheless, RING E3 mechanisms matching a specific UBL and acceptor lysine remain elusive, including for RBX1, which mediates NEDD8 ligation to cullins and >10% of all ubiquitination. We report the structure of a trapped RING E3-E2∼UBL-target intermediate representing RBX1-UBC12∼NEDD8-CUL1-DCN1, which reveals the mechanism of NEDD8 ligation and how a particular UBL and acceptor lysine are matched by a multifunctional RING E3.

Structure of the Proteus vulgaris HigB-(HigA)2-HigB toxin-antitoxin complex.

Bacterial toxin-antitoxin (TA) systems regulate key cellular processes to promote cell survival during periods of stress. During steady-state cell growth, antitoxins typically interact with their cognate toxins to inhibit activity presumably by preventing substrate recognition. We solved two x-ray crystal structures of the Proteus vulgaris tetrameric HigB-(HigA)2-HigB TA complex and found that, unlike most other TA systems, the antitoxin HigA makes minimal interactions with toxin HigB.