Drivers of i-DNA Formation in a Variety of Environments Revealed by Four-Dimensional UV Melting and Annealing
Mingpan Cheng, Jielin Chen, Huangxian Ju, Jun Zhou, Jean‐Louis Mergny
Abstract
studies have often been performed under different experimental conditions, making comparisons difficult. To overcome this, we developed a four-dimensional UV melting and annealing (4DUVMA) approach to analyze i-DNA formation under a variety of conditions (e.g., pH, temperature, salt, crowding). Analysis of 25 sequences provided a global understanding of i-DNA formation under disparate conditions, which should ultimately allow the design of accurate prediction tools. For example, we found reliable linear correlations between the midpoint of pH transition and temperature (-0.04 ± 0.003 pH unit per 1.0 °C temperature increment) and between the melting temperature and pH (-23.8 ± 1.1 °C per pH unit increment). In addition, by analyzing the hysteresis between denaturing and renaturing profiles in both pH and thermal transitions, we found that loop length, nature of the C-tracts, pH, temperature, and crowding agents all play roles in i-DNA folding kinetics. Interestingly, our data indicate which conformer is more favorable for the sequences with an odd number of cytosine base pairs. Then the thermal and pH stabilities of "native" i-DNAs from human promoter genes were measured under near physiological conditions (pH 7.0, 37 °C). The 4DUVMA method can become a universal resource to analyze the properties of any i-DNA-prone sequence.