Functional evaluation of the C-terminal region of bacteriophage T4 Rad50.
Biochem Biophys Res Commun
; 526(2): 485-490, 2020 05 28.
Article
em En
| MEDLINE
| ID: mdl-32238267
ABSTRACT
Bacteriophage T4 encodes orthologs of the proteins Rad50 (gp46) and Mre11 (gp47), which form a heterotetrameric complex (MR) that participates in the processing of DNA ends for recombination-dependent DNA repair. Crystal and high-resolution cryo-EM structures of Rad50 have revealed DNA binding sites near the dimer interface of Rad50 opposite of Mre11, and near the base of the coiled-coils that extend out from the globular head domain. An analysis of T4-Rad50 using sequenced-based algorithms to identify DNA binding residues predicts that a conserved region of positively charged residues near the C-terminus, distal to the observed binding sites, interacts with DNA. Mutant proteins were generated to test this prediction and their enzymatic and DNA binding activities were evaluated. Consistent with the predictions, the Rad50 C-terminal mutants had reduced affinity for DNA as measured by Rad50 equilibrium DNA binding assays and an increased Km-DNA as determined in MR complex nuclease assays. Moreover, the allosteric activation of ATP hydrolysis activity due to DNA binding was substantially reduced, suggesting that these residues may be involved in the communication between the DNA and ATP binding sites.
Palavras-chave
Texto completo:
1
Base de dados:
MEDLINE
Assunto principal:
Proteínas Virais
/
Bacteriófago T4
/
Proteínas de Ligação a DNA
Tipo de estudo:
Prognostic_studies
Idioma:
En
Revista:
Biochem Biophys Res Commun
Ano de publicação:
2020
Tipo de documento:
Article
País de afiliação:
Estados Unidos