ANALYSIS OF PHOTOENZYMATIC REPAIR OF UV LESIONS IN DNA BY SINGLE LIGHT FLASHES .3. COMPARISON OF REPAIR EFFECTS AT VARIOUS TEMPERATURES BETWEEN +37 DEGREES AND -196 DEGREES

UV-irradiated transforming DNA of Haemophilus influenzae can be repaired in vitro in the presence of yeast photoreactivating enzyme (PRE) by an intense light flash of about 1 msec duration. Such a flash repaires a fraction > 0.9 of enzyme-complexed photorepairable lesions at all temperatures between +37° and -40°, but only a fraction of about 0.25 or 0.1 at -78° or -196° respectively. A similar decline below -40° is observed for photoenzymatic repair (PR) of E. coli Bs-1 cells and for PR of transforming DNA by continuous illumination with white light. The decreased effects at low temperatures result neither from damage to the reaction mixture by freezing or by the illumination itself, nor do they result from dissociation of the PRE-substrate complexes in the dark. The drastic reduction in the extent of PR is observed already below -2° if the photolytic reaction is limited by using a less intense light flash. These results are best explained by assuming that the relative repair efficiency of incident photons drops substantially below -2°, but this is not noticed with a very intense flash, where the number of incident photons is excessive. It remains open whether the decreasing repair efficiency with decreasing temperature reflects reduced absorbance or a reduced quantum yield of the absorbed photons. Increased PR effects at -78° and -196° can be obtained by sequential flashes or by increased periods of illumination with white light, but the kinetics indicate the presence of 2 or more fractions of PRE-substrate complexes, differing in their probabilities of being repaired. At low temperature as well as at room temperature flashes photolyse a majority of the complexes more efficiently than equivalent doses of continuous illumination, suggesting an enhancement of PR by absorption of 2 or more photons within a limited time interval. © 1969.