In Vitro Assay to Measure APE1 Enzymatic Activity on Ribose Monophosphate Abasic Site.
Methods Mol Biol
; 2701: 21-38, 2023.
Article
en En
| MEDLINE
| ID: mdl-37574473
ABSTRACT
APE1 (apurinic/apyrimidinic endodeoxyribonuclease 1) is a central enzyme of the base excision repair (BER) pathway playing a pivotal role in protecting mammalian cells against genotoxins and in safeguarding genome stability. Recently, we demonstrated the APE1 ability to process abasic ribonucleotides embedded in DNA. Here, we provide a pipeline of protocols to quantify endodeoxyribonuclease activity by APE1 on these substrates, by using recombinant protein and whole-cell extracts. The repair capacity is measured by using fluorescent oligonucleotide substrates, which are then separated by polyacrylamide gel electrophoresis and detected by imaging scanning. The specificity of APE1 action is demonstrated using specific APE1 enzymatic inhibitors.
Palabras clave
Texto completo:
1
Colección:
01-internacional
Base de datos:
MEDLINE
Asunto principal:
ADN-(Sitio Apurínico o Apirimidínico) Liasa
/
Pruebas de Enzimas
Idioma:
En
Revista:
Methods Mol Biol
Asunto de la revista:
BIOLOGIA MOLECULAR
Año:
2023
Tipo del documento:
Article
País de afiliación:
Italia