Formation of ternary complexes involving zinc or copper ions, polynucleotides, and polypeptides containing glutamic acid and tyrosine residues.
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The formation of triple-stranded nucleic acid helices is studied by molecular mechanics and molecular dynamics calculations. Using standard TAT and CGG homopolymers, single, triple, and quintuple molecular replacements are made. Some of these replacements are expected to form Hoogsteen bonds and some are not. While the electrostatic and total energetic differences for base triplet mismatches were dependent on the electrostatic model chosen, clear trends in the local geometric distortions were apparent. Relationships between these model-built strand geometries and chemical probe experiments are discussed.
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Polymer complexes formed by schizophyllan and poly(C) showed a unique antihydrolysis effect when poly(C) was subjected to hydrolysis under basic conditions. The complexation reduced the hydrolysis rate to 80% of the control (i.e., poly(C) itself). However, when we added oligoamines with the intention of catalyzing the hydrolysis, the oligoamines induced dissociation of the complex instead of acceleration of the hydrolysis.
We have improved a sandwich hybridization assay to detect single base substitutions in polymerase chain reaction (PCR) amplified DNA sequences. The target DNA was captured by an immobilized oligonucleotide and revealed using a second oligonucleotide coupled to an enzyme. Short oligonucleotides (13, 15 bases) were used to obtain specific hybridization at 37 degrees C. We developed two different assay formats for rapid identification of PCR products: a microtitration plate format with oligonucleotides bound to polystyrene and a channelling assay using oligonucleotides immobilized on Sepharose, which did not require any separation step. The specificity and advantages of both methods are described.
As a model for interaction of steroid receptors with DNA, the binding of estradiol receptor complexes (E2R) to oligodeoxynucleotides, covalently linked to cellulose, was studied in detail. Binding was optimal at concentrations of monovalent cationic salts at, or near, isotonic levels and was selective for intracellular receptors in contrast to extracellular steroid binding proteins. Among the oligomers, the order of affinity was oligo dG greater than oligo dT greater than oligo dC greater than oligo dA greater than oligo dI. The binding to oligo dG was stable to 37 degrees C exposure and the processes of adsorption and desorption, while reactivity with oligo dT, oligo dC and oligo dA was labile. The decrease in binding following purification was restored by histone 2B. Oligo dG binding was the most resistant to inhibition by cibacron blue F3GA (CB) and pyridoxal-5-phosphate. On the basis of these data, a hypothesis is proposed for the interaction of mouse uterine cytosol E2R with prevalent nonspecific and putative specific sequences of DNA.