十二指肠贾第虫Rad52蛋白的生化特性及其对DNA损伤的响应

Rosa María Martínez-Miguel, Antonio Sandoval-Cabrera, M. L. Bazán-Tejeda, A. L. Torres-Huerta, Diego A. Martínez-Reyes, R. Bermúdez-Cruz
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引用次数: 4

摘要

十二指肠贾第虫是一种有鞭毛的双核原生动物,在哺乳动物的小肠中定植,引起贾第虫病,急性或慢性腹泻。无论是内源性还是外源性DNA双链断裂都是对DNA的主要损伤,同源重组对DNA的修复对基因组的稳定性至关重要。在HR过程中,Rad52在Rad51重组酶的加载和第二双链断裂端退火到D-loop结构的移位链中起着关键作用。体外在酵母和人Rad52蛋白中发现的功能包括:ssDNA或dsDNA结合活性,退火裸或RPA涂层的ssDNA的能力,以及多聚环的形成。在这项工作中,我们在十二指肠鸡中寻找一个推定的Rad52蛋白(GdRad52)的保守结构域。对其编码序列进行克隆、表达和纯化,研究其生化特性。rGdRad52结合dsDNA和ssDNA,对后者的亲和力更强。同样,rgdrpa52促进未包被和包被GdRPA1的DNA退火。rGdRad52与GdDMC1B和GdRPA1蛋白相互作用。此外,rGdRad52在电子显微镜下形成多聚环。最后,GdRad52在对滋养体DNA损伤的反应中过表达。
本文章由计算机程序翻译,如有差异,请以英文原文为准。
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.
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