Protein semisynthesis reveals plasticity in HECT E3 ubiquitin ligase mechanisms.
Nat Chem
; 2024 Jul 19.
Article
en En
| MEDLINE
| ID: mdl-39030419
ABSTRACT
Lys ubiquitination is catalysed by E3 ubiquitin ligases and is central to the regulation of protein stability and cell signalling in normal and disease states. There are gaps in our understanding of E3 mechanisms, and here we use protein semisynthesis, chemical rescue, microscale thermophoresis and other biochemical approaches to dissect the role of catalytic base/acid function and conformational interconversion in HECT-domain E3 catalysis. We demonstrate that there is plasticity in the use of the terminal side chain or backbone carboxylate for proton transfer in HECT E3 ubiquitin ligase reactions, with yeast Rsp5 orthologues appearing to be possible evolutionary intermediates. We also show that the HECT-domain ubiquitin covalent intermediate appears to eject the E2 conjugating enzyme, promoting catalytic turnover. These findings provide key mechanistic insights into how protein ubiquitination occurs and provide a framework for understanding E3 functions and regulation.
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Banco de datos:
MEDLINE
Idioma:
En
Revista:
Nat Chem
Asunto de la revista:
QUIMICA
Año:
2024
Tipo del documento:
Article
País de afiliación:
Estados Unidos