Neutron-encoded diubiquitins to profile linkage selectivity of deubiquitinating enzymes.
Nat Commun
; 14(1): 1661, 2023 03 25.
Article
em En
| MEDLINE
| ID: mdl-36966155
ABSTRACT
Deubiquitinating enzymes are key regulators in the ubiquitin system and an emerging class of drug targets. These proteases disassemble polyubiquitin chains and many deubiquitinases show selectivity for specific polyubiquitin linkages. However, most biochemical insights originate from studies of single diubiquitin linkages in isolation, whereas in cells all linkages coexist. To better mimick this diubiquitin substrate competition, we develop a multiplexed mass spectrometry-based deubiquitinase assay that can probe all ubiquitin linkage types simultaneously to quantify deubiquitinase activity in the presence of all potential diubiquitin substrates. For this, all eight native diubiquitins are generated and each linkage type is designed with a distinct molecular weight by incorporating neutron-encoded amino acids. Overall, 22 deubiquitinases are profiled, providing a three-dimensional overview of deubiquitinase linkage selectivity over time and enzyme concentration.
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Poliubiquitina
/
Enzimas Desubiquitinantes
Idioma:
En
Ano de publicação:
2023
Tipo de documento:
Article