MECHANISMS OF INACTIVATION OF DEOXYRIBONUCLEIC ACIDS BY HEAT

Abstract

The inactivation by heat of two preparations of the transforming principle of D. pneumoniae has been studied quantitatively with the object of elucidating the mechanisms of heat inactivation. These preparations were both deproteinated by the phenol method, which gives preparations showing simple first-order kinetics. Two processes are found, in accordance with previous workers. The first, or "collapse," process is apparently the actual separation of the complementary strands of the DNA and leads to a rapid and kinetically complex loss of from 93 to 98 % of the activity. The remaining few per cent of activity is apparently inherent in the denatured DNA and does not depend on "renaturation." The other process, the "single-hit" process, is a first-order reaction that apparently proceeds only at those base-pairs that are open as a result of thermal excitation, and hence its rate increases anomalously fast with temperature in the temperature region just below that at which the collapse occurs. It seems likely that the first-order reaction is the loss of a purine from one of the strands. With this assumption and the known rate of depurination, we estimate the size of the streptomycin resistance maker to be about 430 base-pairs, although this estimate must be considered as very tentative in view of differences between the two samples. The collapse temperature is different with different markers in the same sample, while the dependence of the single-hit process on temperature is nearly the same for all markers as long as the collapse process does not intrude. Following other workers, we attribute the differences in the collapse temperatures to heterogeneities in the guanine-cytosine content of the different markers, but in order to reconcile this idea with the rate data, we must assume that the heterogeneities are very local and that the over-all compositions of all the markers are effectively the same.