A proton magnetic resonance study of the interaction of adenosine with polyuridylic Acid: evidence for both adenine-uracil base-stacking and base-pairing.

We report here a proton magnetic resonance (pmr) study of the interaction of adenosine with polyuridylic acid in aqueous solution. The results of this study indicate that the mode of interaction is adenosine intercalation and adenine-uracil base-stacking above 260C, and verifies that a triple-stranded complex which is stabilized by both adenine-uracil hydrogen-bonding and adenineadenine base-stacking is formed below this temperature. Some information regarding the dynamics of these processes has also been obtained. Several studies of the interactions between biological bases and nucleosides in aqueous solution'-3 have shown extensive association by vertical basestacking, but no evidence for base-pairing by horizontal hydrogen-bonding has been found at the monomer level. In nonaqueous solvents such as dimethylsulfoxide (DMSO) and CHC13, however, interaction by horizontal hydrogenbonding is apparently favored.44 The incorporation of a particular base in a polynucleotide has been demonstrated to affect its mode of interaction with monomeric bases and nucleosides in aqueous solution, favoring hydrogenbonding over base-stacking.7-9 Recently, we studied the binding of unsubstituted purine to polyuridylic acid by pmr spectroscopy,'0 and found that at 290C, purine interacts with the polymer by base-stacking and intercalation only. The adenosine-polyuridylic acid system is of more direct biological interest because of the involvement of adenine-uracil base-pairs in the Watson-Crick bonding scheme. Studies of this system by infrared spectroscopy,8 and by equilibrium dialysis, solubility, optical rotation, and hydrodynamic methods9 have shown that adenosine binds to polyuridylic acid in a cooperative manner with a "melting temperature" near 20'C. The complex involves 1 adenosine per 2 uracil bases, and is presumably a rigid triple helix. Materials and Methods.-Polyuridylic acid (poly U) of mol wt 100,000 was obtained from Schwarz BioResearch, Inc. The poly U was converted from the original potassium salt to the sodium salt by passing the sample through a column of Dowex 50-W-X8 cation exchange resin and lyophilizing the resulting solution. Adenosine (A grade) was obtained from Calbiochem and was used without further purification. The poly U and adenosine were dried over P205 at _250C, and solution concentrations in D20 were determined from the weight of the dry materials. A small amount of tetramethylammonium chloride (-0.01 M) was added to the solution to provide an internal reference for the chemical shift measurements. No other salt or buffer was added. The 100 Mc/sec pmr spectra were obtained on a Varian HA-100 nuclear magnetic reso nance (NMR) spectrometer operated in the frequency sweep mode. Tetramethylsilane (TMS) in a sealed capillary was used as an external reference and provided the field/frequency lock signal. Chemical shifts reported are referred to the TMS capillary