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Research was conducted to describe the characterization of the aes gene of Escherichia coli that encodes an enzyme with esterase activity. A Qiagen kit was utilized to prepare the plasmids while the mRNA start point was mapped by the reverse transcriptase method of primer extension. Moreover, the beta-galactosidase activity of lacZ fusion strains was evaluated in permeabilized cells. Results showed that the plasmid-encoded protein is a cytoplasmic enzyme exhibiting esterase activity that counteracts maltose sensitivity.

Research has been conducted on the composite transposon Tn10. The determination of Tn10 complete nucleotide sequence has been performed.

An unexpected property of a ligand is described that has disrupted the structure of two different DNA G-quadruplex structures, each derived from sequence motifs in the promoter of the proto-oncogene c-kit. The studies have shown that the functional consequence of small molecule ligands interacting with G-quadruplex structures has depended on the specific mode of interaction.

The design, synthesis, biophysical evaluation with primary biological data on 3,8,10-trisubstituted isoalloxazines are described. 3,8,10-trisubstituted isoalloxazines are considered to be a new class of G-quadruplex binding ligands that have shown selective binding to a c-kit promoter quadruplex.

It has been hypothesized that the formation of G-quadruplex structures in the DNA of gene promoters may be functionally linked to transcription and consequently that small molecules that interact with such G-quadruplexes may modulate transcription. We previously reported that triarylpyridines are a class of small molecules that selectively interact with G-quadruplex DNA. Here we describe an unexpected property of one such ligand that was found to disrupt the structure of two different DNA G-quadruplex structures, each derived from sequence motifs in the promoter of the proto-oncogene c-kit. Furthermore, cell-based experiments in a cell line that expresses c-kit (HGC-27) showed that the same ligand increased the expression of c-kit. This contrasts with G-quadruplex-inducing ligands that have been previously found to inhibit gene expression. It would thus appear that the functional consequence of small molecule ligands interacting with G-quadruplex structures may depend on the specific mode of interaction. These observations provide further evidence to suggest that G-quadruplex forming sequence motifs play a role that relates to transcription. [ABSTRACT FROM AUTHOR]

DNA processing reactions often involve multiple components acting in concert to achieve the desired outcome. However, it is usually difficult to know how the components communicate and cooperate to orchestrate an ordered series of events. We address this question in the context of the Tn 10 transposition reaction, in which the DNA cleavage and joining events occur within a higher-order complex containing a transposase dimer, two transposon ends and the DNA-bending host-factor IHF (Integration Host Factor). Previously it was shown that the complex is asymmetric. The α side consists of an IHF protomer initially immobilized by a DNA-loop, but subsequently used to promote conformational changes required for the cleavage steps. The β side of the complex was considered to fulfil a more passive role. Here we show that the α side of the complex promotes coupled conformational changes at both transposon ends, while the α and β sides communicate and cooperate to dominate different phases of the transposition reaction. Together, these effects provide for a robust response to critical changes in the transposon end. These findings also explain the intriguing genetic phenotypes of a series of previously reported Tn 10 mutants and have consequences for the evolution of new elements. [ABSTRACT FROM AUTHOR]

The linear and three-dimensional arrangement and composition of chromatin in eukaryotic genomes underlies the mechanisms directing gene regulation. Understanding this organization requires the integration of many data types and experimental results. Here we describe the approach of integrating genome-wide protein-DNA binding data to determine chromatin states. To investigate spatial aspects of genome organization, we present a detailed description of how to run stochastic simulations of protein movements within a simulated nucleus in 3D. This systems level approach enables the development of novel questions aimed at understanding the basic mechanisms that regulate genome dynamics.

DNA processing reactions often involve multiple components acting in concert to achieve the desired outcome. However, it is usually difficult to know how the components communicate and cooperate to orchestrate an ordered series of events. We address this question in the context of the Tn 10 transposition reaction, in which the DNA cleavage and joining events occur within a higher-order complex containing a transposase dimer, two transposon ends and the DNA-bending host-factor IHF (Integration Host Factor). Previously it was shown that the complex is asymmetric. The a side consists of an IHF protomer initially immobilized by a DNA-loop, but subsequently used to promote conformational changes required for the cleavage steps. The beta side of the complex was considered to fulfil a more passive role. Here we show that the a side of the complex promotes coupled conformational changes at both transposon ends, while the a and beta sides communicate and cooperate to dominate different phases of the transposition reaction. Together, these effects provide for a robust response to critical changes in the transposon end. These findings also explain the intriguing genetic phenotypes of a series of previously reported Tn10 mutants and have consequences for the evolution of new elements.