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Conformational dynamics of the interaction of Escherichia coli endonuclease VIII with DNA substrates
Nikita A. Kuznetsov Vladimir V. Koval Dmitry O. Zharkov Olga S. Fedorova 《DNA Repair》2012,11(11):884-891
Endonuclease VIII (Nei) from Escherichia coli is a DNA repair enzyme that removes a wide range of oxidized pyrimidine bases from DNA. As inferred from the crystal structures and biochemical studies, recognition of DNA lesions by Nei involves several conformational changes in both protein and DNA, such as DNA kinking, damaged base eversion into the enzyme's active site, and insertion of a loop of the enzyme into the void formed by the eversion. Excision of the damaged base by Nei also proceeds through several chemical steps: N-glycosidic bond breakage, β-elimination and δ-elimination of the phosphates flanking the lesion. We have used stopped-flow kinetics with fluorescence detection to follow conformational changes in the Nei molecule when the enzyme binds normal DNA, damaged but uncleavable DNA, or several cleavable damaged DNA substrates. Binding normal or damaged uncleavable DNA proceeded in two fluorescently discernible reversible stages, while processing of cleavable substrates involved three reversible stages followed by and irreversible stage and equilibrium with the reaction product. Individual rate constants were calculated for each reaction step. Based on the stopped-flow data, crystal structure, and a comparison with the stopped-flow kinetics of E. coli formamidopyrimidine-DNA glycosylase, a homolog of Nei, we propose the nature of some of the steps that may be involved into the recognition and excision of damaged bases by Nei. 相似文献
434.
Two strains belonging to a novel anamorphic species, Pseudozyma graminicola, were isolated from the leaves of herbaceous plants in the Moscow region (Russia). This species was genetically distinct
from all known Pseudozyma species, based on sequence divergence in the D1/D2 domains of the large subunit rDNA and the ITS region. It is related phylogenetically
to species of the genus Sporisorium (Ustilaginaceae, Ustilaginales). Physiological characteristics distinguishing this novel species from the other species of
the genus Pseudozyma are presented. 相似文献
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Plasmodium falciparum activates endogenous Cl(-) channels of human erythrocytes by membrane oxidation. 下载免费PDF全文
Stephan M Huber Anne-Catrin Uhlemann Nikita L Gamper Christophe Duranton Peter G Kremsner Florian Lang 《The EMBO journal》2002,21(1-2):22-30
Intraerythrocytic survival of the malaria parasite Plasmodium falciparum requires that host cells supply nutrients and dispose of waste products. This solute transport is accomplished by infection-induced new permeability pathways (NPP) in the erythrocyte membrane. Here, whole-cell patch-clamp and hemolysis experiments were performed to define properties of the NPP. Parasitized but not control erythrocytes constitutively expressed two types of anion conductances, differing in voltage dependence and sensitivity to inhibitors. In addition, infected but not control cells hemolyzed in isosmotic sorbitol solution. Both conductances and hemolysis of infected cells were inhibited by reducing agents. Conversely, oxidation induced identical conductances and hemolysis in non-infected erythrocytes. In conclusion, P.falciparum activates endogenous erythrocyte channels by applying oxidative stress to the host cell membrane. 相似文献