Deuteration kinetics of deoxyguanosine, deoxycytidine, and their polynucleotides by means of ultraviolet spectrophotometry.

Abstract

The kinetics of the hydrogen-deuterium exchange reactions of deoxyguanosine (dG), deoxycytidine (dC), double-helical poly[d(G-C)] X poly[d(G-C], and double-helical poly(dG) X poly(dC) have been examined at 20 degrees C, pH 7.0, and in low-salt (0.15 M NaCl) medium by stopped-flow ultraviolet spectrophotometry, in the spectral region of 260 to 320 nm. The rate constant was found to be 78.9 s-1 for dG-NH, 2.2 s-1 for dG-NH2, 39.3 s-1 for dC-NH2, 2.4 s-1 (fast) and 0.94 s-1 (slow) for poly[d(G-C)] X poly[d(G-C)], and 2.2 s-1 (fast) and 0.92 s-1 (slow) for poly(dG) X poly(dC). From these values, the probability of base-pair opening of the G X C containing B-form double helix is estimated to be (3 +/- 1) X 10(-3). This is much greater than what is expected from an extrapolation of the van't Hoff plot at the helix-coil transition region, i.e. at about 110 degrees C. The mechanism of these base-pair openings at 20 degrees C (as well as the mechanism of base-pair reformation) is suggested to be totally different from those in the melting temperature range.