A kinetic study of the alkaline transition of DNA, in clearly defined physico-chemical conditions, is presented, which allows us to identify, within the alkaline transition region, different pH ranges, corresponding to different ratelimiting factors. This analysis brings into consideration three distinct intervals of time which characterize the whole process, namely the time necessary for full hyperchromicity to be reached, the time required for strand separation in the case of a single DNA molecule, and the time for complete denaturation to be reached in the case of a DNA solution.THE RESULTS OBTAINED FROM ULTRACENTRIFUGAL, AND SPECTROPHOTOMETRIC MEASUREMENTS, INVOLVING RAPID MIXING EXPERIMENTS, SEEM TO INDICATE THE FOLLOWING CONCLUSIONS: whereas, in the lower pH ranges considered within the transition region, the denaturation process is limited by the first time constant, this same constant becomes extremely short at higher pH. On the other hand the fact that, in the higher pH range, the second and third time constants do not coincide (the time to unwind a single T2 DNA molecule being at least one order of magnitude shorter than the time required for bulk denaturation to be reached) suggests that in this pH range the overall denaturation rate is limited by a statistical process governing the initiation of unwinding.These observations are discussed in terms of a model in which the unwinding energy is given by the electrostatic repulsions which originate in the deprotonated DNA molecule. The model itself suggests some experiment which seem to confirm it.
A kinetic study of the alkaline transition of DNA, in clearly defined physico-chemical conditions, is presented, which allows us to identify, within the alkaline transition region, different pH ranges, corresponding to different ratelimiting factors. This analysis brings into consideration three distinct intervals of time which characterize the whole process, namely the time necessary for full hyperchromicity to be reached, the time required for strand separation in the case of a single DNA molecule, and the time for complete denaturation to be reached in the case of a DNA solution.THE RESULTS OBTAINED FROM ULTRACENTRIFUGAL, AND SPECTROPHOTOMETRIC MEASUREMENTS, INVOLVING RAPID MIXING EXPERIMENTS, SEEM TO INDICATE THE FOLLOWING CONCLUSIONS: whereas, in the lower pH ranges considered within the transition region, the denaturation process is limited by the first time constant, this same constant becomes extremely short at higher pH. On the other hand the fact that, in the higher pH range, the second and third time constants do not coincide (the time to unwind a single T2 DNA molecule being at least one order of magnitude shorter than the time required for bulk denaturation to be reached) suggests that in this pH range the overall denaturation rate is limited by a statistical process governing the initiation of unwinding.These observations are discussed in terms of a model in which the unwinding energy is given by the electrostatic repulsions which originate in the deprotonated DNA molecule. The model itself suggests some experiment which seem to confirm it.
Authors: Corbin J England; Tanner C Gray; Shubha R L Malla; Samantha A Oliveira; Benjamin R Martin; Gary W Beall; L Kevin Lewis Journal: Anal Biochem Date: 2020-12-31 Impact factor: 3.365