In contrast to double-stranded DNA, there has so far been little evidence that double-stranded RNA can undergo major conformational transitions. We have investigated the conformation in different conditions of the double-stranded RNA molecule poly(G-C).poly(G-C), by NMR, circular dichroism and absorbance spectroscopy. We report here evidence obtained by these different spectroscopic techniques that poly(G-C).poly(G-C) undergoes a transition from the A-form to a left-handed Z-form in conditions of high ionic strength and at temperatures above 35 degrees C. This conformational transition may be of relevance to the biological situations in which double-stranded RNA occurs, such as in ribosomes and in some viruses.

The interconversion between the right (R) and left (L) helical forms of poly [d(G-C)] occurs at low concentrations of MgCI2 and EtOH, acting together in a highly synergistic manner. established by X-ray crystallography (Wang et al., 1979;Drew et al., 1980). However, L DNA formed in Mg2+-EtOH (which we designate as Z* DNA) has unique properties: a) it can be sedimented readily out of solution at low speed, indicative of condensation and intermolecular aggregation; b) it supports the binding of several intercalating (ethidium bromide, actinomycin D) and non-intercalating (mithramycin) drugs, although these interact preferentially with the R (i.e., B) form of DNA; and c) it functions as a template for Escherichia coli RNA polymerase. B and Z* DNAs can be generated under idential ionic conditions and compared in a number of biochemical systems. Our results suggest that left-handed DNA may form under physiological conditions and serve a biological function.

Uracil DNA glycosylase excises uracil residues from DNA that can arise as a result of deamination of cytosine or incorporation of dUMP residues by DNA polymerase. We have carried out a detailed study to define the specificities and the kinetic parameters for its substrates by using a number of synthetic oligodeoxyribonucleotides of varying lengths and containing uracil residue(s) in various locations. The results show that the Escherichia coli enzyme can remove a 5'-terminal U from an oligomer only if the 5'-end is phosphorylated. The enzyme does not remove U residues from a 3'-terminal position, but U residues can be excised from oligonucleotides with either pd(UN)p or pd(UNN) 3'-termini. The oligomer d(UUUUT) can have the second or third U residues from the 5'-end excised even when the neighboring site is an abasic site (3' or 5', respectively). On the basis of these findings, pd(UN)p was anticipated to be the smallest size substrate. Results show detectable amounts of U release from the substrate pd(UT)p; however, significantly higher amounts of U release were observed from pd(UT-sugar) or pd(UTT). Determinations of the Km and Vmax values show that the different rates of U excision from oligomers of different sizes (trimeric to pentameric) but containing U in the same position are largely due to the differences in the Km values, whereas the different rates of U excision from the substrates of the same size but containing U in different positions are largely due to different Vmax values.

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