Association by Hydrogen Bonding of Mononucleotides in Aqueous Solution

Abstract

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...

Principles of Nucleic Acid Structure

“…The height of the rotation barrier calculated by quantum chemical methods (CNDO/2) is 15 to 25 kcallmole, depending on the N-substituents, hydrogen or methyl groups, in cytosine (200)(201)(202), adenine (203), and guanine (203) derivatives. NMR-spectroscopic data seem to indicate that the barrier to rotation of the amino group in purine bases is lower that that in cytosine (204) for which a value of 15 to 18 kcallmole was derived (205), in close agreement with the theoretical results. Because of the high barrier, the rate of rotation about the exocyclic C-N groups in N4-methylcytosine is very slow, around 90 to 270 sec-I, depending on the temperature in the range between 20° and 30°C, and on the solvent (204)(205)(206).…”

Exploration of factors driving incorporation of unnatural dNTPS into DNA by Klenow fragment (DNA polymerase I) and DNA polymerase

“…The earliest theories on polymerase fidelity proposed that polymerase discrimination arose from the hydrogen bonds between the nascent base pair (4,5). Although it is logical to assume that the proper match of the hydrogen bonds between a correct base pair is the basis for the high fidelity of DNA replication, the difference in thermodynamic stability between a matched and mismatched base pair cannot solely account for the low error rates (6–8). …”

Intra molecular Hydrogen Bonding in Flavin Adenine Dinucleotide

“…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.…”