C-terminal domain of the Uup ATP-binding cassette ATPase is an essential folding domain that binds to DNA

The Uup protein belongs to a subfamily of soluble ATP-binding cassette (ABC) ATPases that have been implicated in several processes different from transmembrane transport of molecules, such as transposon precise excision. We have demonstrated previously that Escherichia coli Uup is able to bind DNA. DNA binding capacity is lowered in a truncated Uup protein lacking its C-terminal domain (CTD), suggesting a contribution of CTD to DNA binding. In the present study, we characterize the role of CTD in the function of Uup, on its overall stability and in DNA binding. To this end, we expressed and purified isolated CTD and we investigated the structural and functional role of this domain. The results underline that CTD is essential for the function of Uup, is stable and able to fold up autonomously. We compared the DNA binding activities of three versions of the protein (Uup, UupΔCTD and CTD) by an electrophoretic mobility shift assay. CTD is able to bind DNA although less efficiently than intact Uup and UupΔCTD. These observations suggest that CTD is an essential domain that contributes directly to the DNA binding ability of Uup.     

Significance of the immune response to a major, conformational B-cell epitope on the hepatitis C virus NS3 region defined by a human monoclonal antibody.

The nonstructural protein NS3 of hepatitis C virus (HCV) possesses two enzymatic domains which are thought to be essential for the virus life cycle: an N-terminal serine-type proteinase, responsible for the processing of nonstructural polypeptides, and a C-terminal nucleoside triphosphatase/helicase, presumably involved in the unwinding of the viral genome. The human antibody response to NS3 usually appears early in the course of HCV infection and is predominantly directed against the carboxyl-terminal portion; however, its fine specificity and clinical significance are largely unknown. We have generated a human monoclonal antibody (hMAb), designated CM3.B6, from a cloned B-cell line obtained from the peripheral blood of a patient with chronic HCV infection, which selectively recognized the purified NS3 protein expressed in bacteria or in eukaryotic cells transfected with full-length or NS3 cDNA. Fine-specificity studies revealed that CM3.B6 recognized a 92-amino-acid sequence (clone 8, amino acids 1363 to 1454) selected from an NS3 DNase fragment library but failed to bind to 12-mer peptides synthesized from the same region, suggesting recognition of a conformational B-cell epitope. Experiments using deletion mutants of clone 8 and competitive inhibition studies using a panel of NS3 peptide-specific murine MAbs indicated that limited N-terminal and C-terminal deletions resulted in a significant reduction of hMAb binding to clone 8, thus identifying a minimal antibody binding domain within clone 8. Competition experiments showed that binding of CM3.B6 to the NS3 protein was efficiently inhibited by 39 of 44 (89%) sera from HCV-infected patients, suggesting that the hMAb recognized an immunodominant epitope within the NS3 region. More importantly, recognition of the sequence defined by CM3.B6 appeared to accurately discriminate between viremic and nonviremic anti-HCV positive sera, suggesting potentially relevant clinical applications in the diagnosis and treatment of HCV infection.     

Deadly teamwork: neural cancer stem cells and the tumor microenvironment

Neural cancers display cellular hierarchies with self-renewing tumorigenic cancer stem cells (CSCs) at the apex. Instructive cues to maintain CSCs are generated by both intrinsic networks and the niche microenvironment. The CSC-microenvironment relationship is complex, as CSCs can modify their environment and extrinsic forces induce plasticity in the cellular hierarchy.     

Chelating derivatives of chitosan obtained by reaction with ascorbic acid

Ascorbic acid immediately dissolves Euphausia superba chitosan upon mixing and forms chitosan ascorbate; during the 6-h period after dissolution in water at pH 5–7, ascorbate is oxidized to dehydroascorbate which undergoes Schiff reaction with the amino groups of chitosan, thus yielding a viscous solution of a polymeric ketimine. The latter is characterized by infrared spectrometry, circular dichroism spectropolarimetry, viscometry and alkalimetry. When brought into contact with transition metal ions, the chitosan ascorbate ketimine yields insoluble metal chelates. Upon reduction with sodium cyanoborohydride, the water-insoluble N-[2-(1,2-dihydroxyethyl)tetrahydrofuryl] chitosan (NDTC) is obtained, which shows enhanced capacity for uranium, up to 800 mg U/g from solutions at pH 4·5.     

Validity of the reduced version of the Morningness-Eveningness Questionnaire

Sleep and Biological Rhythms - The aim of the present work was to validate the reduced version of the Morningness-Eveningness Questionnaire (MEQr) using circadian motor activity as external...     

Functional Anatomy of the Ovule in Broad Bean (Vicia faba L.): Ultrastructural Seed Development and Nutrient Pathways

Ovules of broad bean (Vicia faba L.) were studied to discloseultrastructural features, which can facilitate nutrient transportto the embryo sac from 10 d after pollination (DAP) to the matureseed. Fertilization occurs during the first 24 h after pollination.The endosperm is a coenocyte, which is eventually consumed bythe embryo. By 10 DAP the inner integument is degraded and theouter integument adjoins the embryo sac boundary. The heart-shapedembryo approaches the embryo sac boundary at two sites, whichhere are named contact zones. Small integument cells in theneighbourhood of the first formed contact zones become separatedby prominent intercellular spaces. A heterogenous scatteringmaterial, probably representing secretion products accumulatesin these spaces. By 14-16 DAP the integument exudate disappears,and the suspensor degenerates. As the contact zones increasein size, wall ingrowths form a bridging network in the narrowspace between the embryo sac boundary and the extra-embryonicpart of the endosperm wall. The epidermal cells of the embryoseparate adjacent to these zones, and develop conspicuous wallingrowths. At 20 DAP vacuoles showing various stages in formationof protein bodies appear in the cells of the embryo.Copyright1994, 1999 Academic Press Vicia faba, broad beans, ovule, seed, nutrient transport