Articles by Marie-Michelle Genois in JoVE
GST-Su purificación: A dos pasos Affinity Protocolo Purificación Cediendo largos proteínas purificadas Ranjan Maity*1, Joris Pauty*1, Jana Krietsch*1, Rémi Buisson1, Marie-Michelle Genois1, Jean-Yves Masson1 1Genome Stability Laboratory, Oncology Axis, Hôtel-Dieu de Québec En el presente protocolo, se demuestra un método de purificación de proteínas a pequeña escala altamente eficiente y rentable, que permite la purificación de proteínas recombinantes, combinando de forma única una etiqueta de GST escindible y una pequeña cola de His.
Other articles by Marie-Michelle Genois on PubMed
Lactococcal Phage P2 ORF35-Sak3 is an ATPase Involved in DNA Recombination and AbiK Mechanism Molecular Microbiology. Apr, 2011 | Pubmed ID: 21276096 Virulent phages of the Siphoviridae family are responsible for milk fermentation failures worldwide. Here, we report the characterization of the product of the early expressed gene orf35 from Lactococcus lactis phage p2 (936 group). ORF35(p2), also named Sak3, is involved in the sensitivity of phage p2 to the antiviral abortive infection mechanism AbiK. The localization of its gene upstream of a gene coding for a single-strand binding protein as well as its membership to a superfamily of single-strand annealing proteins (SSAPs) suggested a possible role in homologous recombination. Electron microscopy showed that purified ORF35(p2) form a hexameric ring-like structure that is often found in proteins with a conserved RecA nucleotide-binding core. Gel shift assays and surface plasmon resonance data demonstrated that ORF35(p2) interacts preferentially with single-stranded DNA with nanomolar affinity. Atomic force microscopy showed also that it preferentially binds to sticky DNA substrates over blunt ends. In addition, in vitro assays demonstrated that ORF35(p2) is able to anneal complementary strands. Sak3 also stimulates Escherichia coli RecA-mediated homologous recombination. Remarkably, Sak3 was shown to possess an ATPase activity that is required for RecA stimulation. Collectively, our results demonstrate that ORF35(p2) is a novel SSAP stimulating homologous recombination.
Interactions Between BRCA2 and RAD51 for Promoting Homologous Recombination in Leishmania Infantum Nucleic Acids Research. Aug, 2012 | Pubmed ID: 22505581 In most organisms, the primary function of homologous recombination (HR) is to allow genome protection by the faithful repair of DNA double-strand breaks. The vital step of HR is the search for sequence homology, mediated by the RAD51 recombinase, which is stimulated further by proteins mediators such as the tumor suppressor BRCA2. The biochemical interplay between RAD51 and BRCA2 is unknown in Leishmania or Trypanosoma. Here we show that the Leishmania infantum BRCA2 protein possesses several critical features important for the regulation of DNA recombination at the genetic and biochemical level. A BRCA2 null mutant, generated by gene disruption, displayed genomic instability and gene-targeting defects. Furthermore, cytological studies show that LiRAD51 can no longer localize to the nucleus in this mutant. The Leishmania RAD51 and BRCA2 interact together and the purified proteins bind single-strand DNA. Remarkably, LiBRCA2 is a recombination mediator that stimulates the invasion of a resected DNA double-strand break in an undamaged template by LiRAD51 to form a D-loop structure. Collectively, our data show that LiBRCA2 and LiRAD51 promote HR at the genetic and biochemical level in L. infantum, the causative agent of visceral leishmaniasis.