We describe a simple approach and present a straightforward numerical algorithm to compute the best fit shot-noise limited proximity ratio histogram (PRH) in single-molecule fluorescence resonant energy transfer diffusion experiments. The key ingredient is the use of the experimental burst size distribution, as obtained after burst search through the photon data streams. We show how the use of an alternated laser excitation scheme and a correspondingly optimized burst search algorithm eliminates several potential artifacts affecting the calculation of the best fit shot-noise limited PRH. This algorithm is tested extensively on simulations and simple experimental systems. We find that dsDNA data exhibit a wider PRH than expected from shot noise only and hypothetically account for it by assuming a small Gaussian distribution of distances with an average standard deviation of 1.6 Å. Finally, we briefly mention the results of a future publication and illustrate them with a simple two-state model system (DNA hairpin), for which the kinetic transition rates between the open and closed conformations are extracted.

We introduce an extended version of oxDNA, a coarse-grained model of deoxyribonucleic acid (DNA) designed to capture the thermodynamic, structural, and mechanical properties of single-and double-stranded DNA. By including explicit major and minor grooves and by slightly modifying the coaxial stacking and backbone-backbone interactions, we improve the ability of the model to treat large (kilobase-pair) structures, such as DNA origami, which are sensitive to these geometric features. Further, we extend the model, which was previously parameterised to just one salt concentration ([Na + ] = 0.5M), so that it can be used for a range of salt concentrations including those corresponding to physiological conditions. Finally, we use new experimental data to parameterise the oxDNA potential so that consecutive adenine bases stack with a different strength to consecutive thymine bases, a feature which allows a more accurate treatment of systems where the flexibility of single-stranded regions is important. We illustrate the new possibilities opened up by the updated model, oxDNA2, by presenting results from simulations of the structure of large DNA objects and by using the model to investigate some salt-dependent properties of DNA. C 2015 AIP Publishing LLC. [http://dx

Prebiotic chemistry presumably took place before formation of an oxygen-rich atmosphere and thus under conditions of intense short wavelength UV irradiation. Therefore, the UV photochemical stability of the molecular building blocks of life may have been an important selective factor in determining the eventual chemical makeup of critical biomolecules. To investigate the role of UV irradiation in base-pairing we have studied guanine (G) and cytosine (C) base pairs in the absence of the RNA backbone. We distinguished base-pair structures by IR-UV hole-burning spectroscopy as well as by high-level correlated ab initio calculations. The Watson-Crick structure exhibits broad UV absorption, in stark contrast to other GC structures and other base-pair structures. This broad absorption may be explained by a rapid internal conversion that makes this specific base pair arrangement uniquely photochemically stable.ab initio computation ͉ DNA base pairs ͉ IR-UV spectroscopy ͉ jet cooling ͉ photochemistry T he DNA bases involved in reproduction have short S 1 excited state lifetimes, of the order of 1 ps or less (1-7). It has been argued that this phenomenon serves to protect these bases against photochemical damage, because after excitation they do not cross to the reactive triplet state; instead, they rapidly internally convert to the electronic ground state (8). This mechanism may have been particularly significant under the conditions of the early earth, when purines and pyrimidines presumably were assembled into the first macromolecular structures, producing RNA. At that time, the earth was exposed to shorter wavelength UV radiation than it is today. For an analysis of possible prebiotic chemistry, it is necessary not only to consider the individual bases but also to study them as they interact and to do so without the RNA backbone (9). We achieve such an analysis by studying clusters of guanine (G) and cytosine (C) in the gas phase by using double resonant laser spectroscopy techniques. The experimental studies are accompanied by stateof-the-art quantum chemical and molecular dynamics calculations of the pairing of G and C. These two bases can form many different hydrogen-bonded structures, of which at least 20 are within 12 kcal͞mol of the global minimum. Thus, the familiar Watson-Crick (WC) structure may not be unique, based on energies alone. Here, we report a remarkable difference in excited state properties among the WC structure and other structures. The former exhibits broad UV absorption features; this is in contrast with the sharp UV spectra exhibited by non-WC structures. The broad absorption can be explained by recent theoretical results, predicting a pathway for rapid internal conversion (10). If the difference is solely due to lifetime broadening, these results correspond to a lifetime for the WC structures that is at least 2 orders of magnitude shorter than those of observed non-WC structures. Thus, it appears possible that the WC recognition mechanism involves a structure that was significantly more stab...

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