Giardia duodenalis Rad52 protein: biochemical characterization and response upon DNA damage.
Giardia duodenalis is a flagellated binucleated protozoan that colonizes the small intestine in mammals, causing giardiasis, acute or chronic diarrhea. DNA double strand break either endogenously or exogenously generated is a major insult to DNA and its repair by homologous recombination (HR) is crucial for genomic stability. During HR, Rad52 plays key roles in the loading of the Rad51 recombinase, and the annealing of the second double-strand break end to the displaced strand of the D-loop structure. Among the functions found in vitro in yeast and human Rad52 protein are: ssDNA or dsDNA binding activity, ability to anneal bare or RPA coated-ssDNA, as well as multimeric ring formation. In this work, we searched for conserved domains in a putative Rad52 protein from G. duodenalis (GdRad52). Its coding sequence was cloned, expressed and purified to study its biochemical properties. rGdRad52 binds to dsDNA and ssDNA, with greater affinity for the latter. Likewise, rGdRad52 promotes annealing of DNA uncoated and coated with GdRPA1. rGdRad52 interacts with GdDMC1B and with GdRPA1 protein as shown in far western blotting assay. Additionally, rGdRad52 formed multimeric rings as observed by electronic microscopy. Finally, GdRad52 is over expressed in response upon DNA damage inflicted on trophozoites.
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Ciudad de México, Mexico.
Rosa María
RM
Martínez-Miguel
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Ciudad de México, Mexico.
Antonio
A
Sandoval-Cabrera
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Ciudad de México, Mexico.
María Luisa
ML
Bazán-Tejeda
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Ciudad de México, Mexico.
Ana Laura
AL
Torres-Huerta
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Ciudad de México, Mexico.
Diego A
DA
Martínez-Reyes
Departamento de Genética y Biología Molecular, Centro de Investigación y de Estudios Avanzados del IPN, Ciudad de México, Mexico.
Rosa María
RM
Bermúdez-Cruz
D019295:Computational Biology; D004247:DNA / Q000737:chemistry / Q000378:metabolism*; D004249:DNA Damage; D016829:Giardia lamblia / Q000737:chemistry* / Q000166:cytology / Q000378:metabolism; D008854:Microscopy, Electron; D008958:Models, Molecular; D051721:Rad52 DNA Repair and Recombination Protein / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*
D051721:Rad52 DNA Repair and Recombination Protein; D004247:DNA
DNA double strand break; DNA repair; Giardia duodenalis; Rad52; homologous recombination