Reaction of formaldehyde with nucleotides and ribonucleic acid

Abstract

1. The products of formaldehyde condensation with purine nucleotides and RNA have been separated and isolated chromatographically. 2. Adenylic acid, like adenine and adenosine, affords a methylene bis-derivative on treatment with formaldehyde. Three isomers of methylene bis-(adenosine 2′(3′)-phosphate) have been separated by chromatography on Dowex 1 (formate); the isomers differ from each other in the positions of the phosphate residues. On enzymic dephosphorylation, all three isomers produced a compound identified chromatographically and spectrophotometrically as methylene bis-adenosine. 3. The reaction of nucleotides with formaldehyde yield, besides adenylic methylene dinucleotide, guanylic methylene dinucleotide and methylene adenine-guanine dinucleotide. The latter compounds were identified kinetically by the bell-shaped curve of the dependence of the product formation rate upon the logarithm of formaldehyde concentration, peculiar to methylene bis-derivatives. Other indirect evidence for the formation of methylene bis-guanylic acid and methylene adenine-guanine dinucleotide has also been obtained. No condensation products of pyrimidine nucleotides with formaldehyde have been detected by chromatography. 4. All three above-mentioned methylene dinucleotides ( P-Ado-CH 2-Ado- P, P-Guo-CH 2-Guo- P and P-Ado-CH 2-Guo- P) have been isolated from the alkaline hydrolysate of formaldehyde-treated RNA. No indications of the formation of bridges between pyrimidine residues of RNA have been obtained. 5. A method for the quantitative estimation of methylene derivatives in formaldehyde-treated RNA has been developed. At pH 4.8 and 20° the reaction of RNA and formaldehyde is complete in about 15 days; at a low ionic strength, from 15–19 % of the purine bases of RNA form methylene dimers. When treated with formaldehyde under similar conditions RNA was found to contain twice as many bridges as are present in the mixture of purine nucleotides. 6. The possibility of the intramolecular formation of cross-links in RNA and their use in biochemical investigations and for the interpretation of certain biological effects of formaldehyde is discussed.