2,242 research outputs found

    Solution structure of the DNA-binding domain of RPA from Saccharomyces cerevisiae and its interaction with single-stranded DNA and SV40 T antigen

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    Replication protein A (RPA) is a three-subunit complex with multiple roles in DNA metabolism. DNA-binding domain A in the large subunit of human RPA (hRPA70A) binds to single-stranded DNA (ssDNA) and is responsible for the species-specific RPA–T antigen (T-ag) interaction required for Simian virus 40 replication. Although Saccharomyces cerevisiae RPA70A (scRPA70A) shares high sequence homology with hRPA70A, the two are not functionally equivalent. To elucidate the similarities and differences between these two homologous proteins, we determined the solution structure of scRPA70A, which closely resembled the structure of hRPA70A. The structure of ssDNA-bound scRPA70A, as simulated by residual dipolar coupling-based homology modeling, suggested that the positioning of the ssDNA is the same for scRPA70A and hRPA70A, although the conformational changes that occur in the two proteins upon ssDNA binding are not identical. NMR titrations of hRPA70A with T-ag showed that the T-ag binding surface is separate from the ssDNA-binding region and is more neutral than the corresponding part of scRPA70A. These differences might account for the species-specific nature of the hRPA70A–T-ag interaction. Our results provide insight into how these two homologous RPA proteins can exhibit functional differences, but still both retain their ability to bind ssDNA

    Inhibition of poly(ADP-ribose)polymerase binding to DNA by thymidine dimer

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    AbstractThe ability of poly(ADP-ribose)polymerase to bind damaged DNA was assessed by electrophoretic mobility shift assay. DNA binding domain of poly(ADP-ribose)polymerase (PARPDBD) binds to synthetic deoxyribonucleotide duplex 10-mer. However, the synthetic deoxyribonucleotide duplex containing cys-syn thymidine dimer which produces the unwinding of DNA helix structure lost its affinity to PARPDBD. It was shown that the binding of PARPDBD to the synthetic deoxyribonucleotide duplex was not affected by O6-Me-dG which causes only minor distortion of DNA helix structure. This study suggests that the stabilized DNA helix structure is important for poly(ADP-ribose)polymerase binding to DNA breaks, which are known to stimulate catalytic activity of poly(ADP-ribose)polymerase

    Arterial Injury Associated with Tension-Free Vaginal Tapes-SECUR Procedure Successfully Treated by Radiological Embolization

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    Various postoperative complications have been reported after the use of tension-free vaginal tapes (TVT). The transobturator approach was introduced to minimize the potential complications. The next generation of recently introduced TVT-SECUR is intended to minimize the incidence of complications. Herein we report a case of internal pudendal artery injury sustained during this procedure that was successfully treated by radiological embolization. Angiography with vessel embolization, when available, should be considered when the arterial injury is suspected

    Extraction and separation of hexavalent molybdenum from acidic sulfate solutions using Alamine 336 as an extractant

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    Extraction and separation of hexavalent molybdenum from acidic sulfate solutions using Alamine 336 as an extractant and kerosene as diluent is described. Variation of acid concentration influences the percentage of extraction of hexavalent molybdenum indicating the ion exchange type mechanism. Effect of concentration of Alamine 336 on the extraction of hexavalent molybdenum in the presence of divalent copper and trivalent iron is also presented. The upper limit of concentration of extractant for extraction of hexavalent molybdenum free from divalent copper and trivalent iron is observed with 0.1 mol/L of Alamine 336. However, when the concentration of Alamine 336 is increased to 1.0 mol/L, about 48% of copper is extracted along with molybdenum but without any iron. The method is suitable for the separation of molybdenum from copper and iron contained leach liquor. The optimized experimental parameters such as phase contact time, effect of extractant, metal, loading capacity of extractant and followed by strip ping studies with ammonia, hydrogen peroxide, sodium hydroxide, sodium thiosulfate and thiourea is presented. From the above experimental data we proposed the aqueous mechanism for hexavalent molybdenum extraction processes

    Insight into highly conserved H1 subtype-specific epitopes in influenza virus hemagglutinin

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    Influenza viruses continuously undergo antigenic changes with gradual accumulation of mutations in hemagglutinin (HA) that is a major determinant in subtype specificity. The identification of conserved epitopes within specific HA subtypes gives an important clue for developing new vaccines and diagnostics. We produced and characterized nine monoclonal antibodies that showed significant neutralizing activities against H1 subtype influenza viruses, and determined the complex structure of HA derived from a 2009 pandemic virus A/Korea/01/2009 (KR01) and the Fab fragment from H1-specific monoclonal antibody GC0587. The overall structure of the complex was essentially identical to the previously determined KR01 HA-Fab0757 complex structure. Both Fab0587 and Fab0757 recognize readily accessible head regions of HA, revealing broadly shared and conserved antigenic determinants among H1 subtypes. The beta-strands constituted by Ser110-Glu115 and Lys169-Lys170 form H1 epitopes with distinct conformations from those of H1 and H3 HA sites. In particular, Glu112, Glu115, Lys169, and Lys171 that are highly conserved among H1 subtype HAs have close contacts with HCDR3 and LCDR3. The differences between Fab0587 and Fab0757 complexes reside mainly in HCDR3 and LCDR3, providing distinct antigenic determinants specific for 1918 pdm influenza strain. Our results demonstrate a potential key neutralizing epitope important for H1 subtype specificity in influenza virus
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