Evidence for hydrogen bonding between 5'-ribonucleotides in water has been obtained from a 220-MHz proton magnetic resonance study of nitrogenous protons. The amino groups of GMP, AMP, and CMP exhibit proton resonance lines which are somewhat broadened by proton exchange with the solvent at 00; their downfield shifts in mixtures of mononucleotides provide the basis for the following order of base-pairing tendencies: GMP CMP > AMP. UMP. Hydrogen bonding is also observed in other pairs of mononucleotides, notably GMP UMP, AMP-CMP, and CMP-UMP, to a lesser extent in GMP-IMP, CMPS XMP, and possibly in CMP-IMP. In agreement with previous reports, hydrophobic interactions of mononucleotides have also been observed; base pairing occurs in addition to vertical stacking of these bases, their hydrogen bonding to water, or self-association. Only CMP shows clear evidence of self-association via hydrogen bonding in water; the evidence for GMP is less direct, and that for AMP is negative. This lack of observable self-association may occur as a result of competition from strong stacking interactions.Only CMP shows restricted rotation of the amino group at 00 and neutral pH. As expected, higher temperatures increase the rate of rotation of the amino group for CMP, as well as accelerate the rate of proton exchange between water and the amino protons of mononucleotides. High-resolution proton magnetic resonance spectroscopy could prove to be a valuable tool in mapping out the specificities conferred by hydrogen bonding between biomolecules in aqueous solution.Hydrogen bonding between the monomers of nucleic acids has been the object of numerous investigations. The initial success of infrared spectroscopy in demonstrating hydrogen bonding between A and U derivatives in deuterochloroform (1) was quickly followed by more extensive infrared studies (2, 3), as well as by two proton magnetic resonance (PMR) reports (4, 5) that showed the remarkable Watson-Crick specificity of G * C, and the much weaker A * U interaction in organic solvents. Other possible combinations of bases showed little or no association, as analyzed by infrared and PMR spectroscopy.Although interactions occurring in aqueous solvents are more applicable to biological systems, little work has been done on hydrogen bonding of nucleotide bases in water. Several workers (6-8) have interpreted their results in terms of base pairing, but these chromatographic and solubility studies, as well as an extensive osmometric study (9), were incapable of distinguishing between hydrogen bonding and stacking interactions. Raman spectroscopy studies of complementary mononucleotides failed to detect any spectral changes (10), whereas PMR spectroscopy studies generally revealed upfield shifts of nondissociable ring protons; this result is consistent with the view that stacking of mononucleotides is predominant in D20 (11-13).We believe that we have PMR evidence of hydrogen bonding between mononucleotides in aqueous solution. This evidence is based on the detection and measurem...

Data obtained by means of proton magnetic resonance spectroscopy indicate that specific association of FMN Proton magnetic resonance (PMR) spectroscopy has been widely applied to detect the specific interactions of molecules in solution. By the use of water as the solvent, we have recently applied the technique to monitoring those hydrogen bonding interactions that involve the amino groups of mononucleotides (1). We have examined the specificity conferred by hydrogen bonding between FMN and AMP, and extended this study to the intramolecular interaction between the flavin and adenine moieties of FAD. Earlier PMR studies of FAD and its components have established the assignment of proton resonances and proposed various models of intramolecular stacking as the predominant mode of association in aqueous media (2-4).It is generally believed that nonpolar solvents obliterate stacking interactions. Using infrared spectroscopy, Kyogoku and Yu (5, 6) have reported the specific association of riboflavin with adenine derivatives in chloroform, and indicated that a hydrogen bonded dimer is not formed between the derivatives of riboflavin and guanosine, cytidine, or uracil. Furthermore, Voet and Rich (7)

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The aromatic amino acids tryptophan, phenylalanine, and histidine interact with singlestranded polyadenylic acid [poly(A)i as observed by proton magnetic resonance spectroscopy. The chemical shift of the C2 and C8 protons of the adenine moiety of poly(A) is consistent with a destacking of the initially partly-stacked polynucleotide chain by the intercalation of the planar ring structure. The relative magnitude of this interaction is tryptophan>phenylalanine>histidine.The measurement of specific interactions (binding constants) between amino acids and nucleic acids are of interest not only as a basis for a stereochemical theory of the origin of the genetic code (1, 2) but also to understand the nature of protein-nucleic acid interactions in general (for example, the cases of repressors, initiators, and polymerases). Zubay and Doty (3) used equilibrium dialysis to study the binding to calf-thymus DNA of those amino acids that are predominantly found in thymus tissue with essentially negative results. By the same technique, a preference of AT-rich DNA for polylysine over polyarginine has been reported by Leng and Felsenfeld (4); recent experiments by Fox et al. (5) indicate that polyamino acids rich in arginine tend to react preferentially with polymers of nucleotides of adenine and guanine, while polyamino acids rich in lysine tend to react preferentially with polymers of nucleotides rich in uridine and cytidine. Further, qualitative evidence has been presented for the specificity of interaction between a few dipeptides and tRNA (6) as monitored by line broadening in proton magnetic resonance (PMR) spectroscopy. We wish to report here some evidence, obtained by PMR spectroscopy, of the interaction of aromatic amino acids with neutral poly(A), and the effect of this interaction on the melting temperature of single-stranded poly(A). shown to interact minimally with polynucleotides (7). Intensities of the poly(A) aromatic region were integrated by cutting out and weighing the spectral areas.Binding constants were obtained with the help of a computer program described elsewhere (8, 9). The feature of this method is that in the limit case of fast exchange, the chemical shift of a nucleus that results from binding can be quantitated

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