A spectrophotometric study of the secondary structure of ribonucleic acid based on a method for diminishing single-stranded base-‘stacking’ without affecting multi-helical structures

Abstract

On the basis of studies with model compounds it was concluded that in 8M-urea-M-potassium chloride (or 4M-guanidinium chloride) in 0.01 M-potassium phosphate buffer, pH 7.0, multi-helical structures have about the same stability as in 0. 1M-potassium phosphate buffer, pH 7.0, whereas the tendency of base residues to ''stack'' along a single polynucleotide chain is much decreased. Base-pairing was eliminated whereas base-''stacking'' persisted after RNA in 1% formaldehyde-0.1M-potassium phosphate buffer, pH 7.0, was heated to 95[degree]. From a study of the thermal de-naturation of unfractionated transfer RNA from Escherichia coli and of RNA from the fractionated sub-units of rabbit reticulocyte ribo-somes in 8M-urea-M-potassium chloride (or 4M-guanidinium chloride) in 0.01M-potassium phosphate buffer, pH 7.0, it was inferred that ''stacked'' residues may account for up to 25% of the increase in E260 found on heating RNA in solvents such as 0.1M-potassium phosphate buffer, pH 7.0. Changes in the spectrum with temperature were analysed on the basis of the assumptions that the polynucleotide chain is amorphous on denaturation (which is probable in 8M-urea-M-potassium chloride-0.01 M-potassium phosphate buffer, pH 7.0) and that the polynucleotide chain adopts a single-stranded ''stacked'' con-formation on denaturation (which is probable when ordinary solvents such as 0.1 M-potassium phosphate buffer, pH 7.0, are used).