Slow exchange of the “outside” amino hydrogens of DNA

Abstract

Rapid hydrogen exchange measurements of native calf thymus DNA, in exchange-out times as short as 7 seconds, confirm the earlier finding of Hanson (1969) that the “outside” (non-interchain hydrogen-bonded) amino hydrogens of bases in double helical DNA exchange anomalously slowly, by a factor of about 10 4, in comparison to expectations from mononucleotide data. Our results demonstrate that all the amino hydrogens of the DNA bases, the 3 arising in dA · T pairs as well as the 5 in dG · dC pairs, participate in this slow exchange phenomenon. The native DNA exchange-out curve (for exchange-out times less than 1000 sec), obtained at 0 °C, pH 7 and 0.1 m-NaCl, can be discriminated into two exchange rate classes. One contains approximately 1.4 hydrogens per nucleotide pair and has a half-time of approximately 25 sec. The other has approximately 2.4 hydrogens per nucleotide pair with a half-time of approximately 330 sec. The former class may represent the outside hydrogens and the latter the inside (internucleotide hydrogen-bonded) hydrogens, though other interpretations could be devised to fit these numbers. Molecular mechanisms which might account for the observed slowing of exchange of the outside hydrogens are considered, and previous interpretations of hydrogen exchange kinetics as a probe of the conformational motility of the native DNA helix are reviewed in the light of these new findings.