Temperature Effect on Ionic Current and ssDNA Transport through Nanopores.

We have investigated the role of electrostatic interactions in the transport of nucleic acids and ions through nanopores. The passage of DNA through nanopores has so far been conjectured to involve a free-energy barrier for entry, followed by a downhill translocation where the driving voltage accelerates the polymer. We have tested the validity of this conjecture by using two toxins, α-hemolysin and aerolysin, which differ in their shape, size, and charge. The characteristic timescales in each toxin as a function of temperature show that the entry barrier is ∼15 kBT and the translocation barrier is ∼35 kBT, although the electrical force in the latter step is much stronger. Resolution of this fact, using a theoretical model, reveals that the attraction between DNA and the charges inside the barrel of the pore is the most dominant factor in determining the translocation speed and not merely the driving electrochemical potential gradient.

Thermal unfolding of proteins probed at the single molecule level using nanopores.

The nanopore technique has great potential to discriminate conformations of proteins. It is a very interesting system to mimic and understand the process of translocation of biomacromolecules through a cellular membrane. In particular, the unfolding and folding of proteins before and after going through the nanopore are not well understood. We study the thermal unfolding of a protein, probed by two protein nanopores: aerolysin and α-hemolysin. At room temperature, the native folded protein does not enter into the pore. When we increase the temperature from 25 to 50 °C, the molecules unfold and the event frequency of current blockade increases. A similar sigmoid function fits the normalized event frequency evolution for both nanopores, thus the unfolding curve does not depend on the structure and the net charge of the nanopore. We performed also a circular dichroism bulk experiment. We obtain the same melting temperature (around 45 °C) using the bulk and single molecule techniques.

Stability and structure of long intramolecular G-quadruplexes.

G-quadruplexes are formed from guanine-rich sequences of DNA and RNA. They consist of stacks of square arrangements of guanines called G-quartets. Increasing evidence suggests that these structures are involved in cellular processes such as transcription or translation. Knowing their structure and their stability in vitro should help to predict their formation in vivo and to understand their biological functions. Many studies have been performed on isolated G-quadruplexes, but little attention has been given to their interactions. Here, we present non-denaturing gel electrophoresis, UV melting, and circular dichroism data obtained for long sequences of DNA which are capable of forming two simultaneous G-quadruplexes, namely, d(TG(3)T(3)G(3)T(3)G(3)T(3)G(3)T(n)G(3)T(3)G(3)T(3)G(3)T(3)G(3)T), with n varying from one to seven. These sequences can form up to two separate G-quadruplexes. We also study mutated versions of these sequences designed to form one G-quadruplex at specific positions on the strand. Comparing results from the original sequences and their mutated versions, we show that for the former different folded states coexist: either with six stacked G-quartets or only three, in various combinations. Which ones are favored depends on n. Moreover, for n greater than three, the thermodynamic stability stays constant, contrary to an expected decrease in stability if the six G-quartets were stacked together in a single structure. This result agrees with a beads-on-a-string folding model for long sequences of G-quadruplexes, where two adjacent G-quadruplexes fold independently.

Predicting and understanding the stability of G-quadruplexes.

MOTIVATION: G-quadruplexes are stable four-stranded guanine-rich structures that can form in DNA and RNA. They are an important component of human telomeres and play a role in the regulation of transcription and translation. The biological significance of a G-quadruplex is crucially linked with its thermodynamic stability. Hence the prediction of G-quadruplex stability is of vital interest. RESULTS: In this article, we present a novel Bayesian prediction framework based on Gaussian process regression to determine the thermodynamic stability of previously unmeasured G-quadruplexes from the sequence information alone. We benchmark our approach on a large G-quadruplex dataset and compare our method to alternative approaches. Furthermore, we propose an active learning procedure which can be used to iteratively acquire data in an optimal fashion. Lastly, we demonstrate the usefulness of our procedure on a genome-wide study of quadruplexes in the human genome. AVAILABILITY: A data table with the training sequences is available as supplementary material. Source code is available online at http://www.inference.phy.cam.ac.uk/os252/projects/quadruplexes. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Function and targeting of G-quadruplexes.

Guanine-rich sequences of DNA and RNA may fold in vitro and in vivo into G-quadruplexes, four-stranded helical structures held together by a guanine core. G-quadruplexes have various biological functions, including inhibition of telomerase and the regulation of gene transcription and translation, and have become an active target for drug development, particularly for novel anticancer therapies. The physiological functions of G-quadruplexes are discussed in this review and the current knowledge of G-quadruplex ligand and drug development is outlined.

Emulsification and stabilization mechanisms of o/w emulsions in the presence of chitosan.

We study emulsification of paraffin oil in aqueous solutions of chitosan without adding any other surfactant. By monitoring the surface tension of the water-paraffin interface, we show that chitosan itself has only a weak surface activity. Nevertheless, chitosan dissolved in the aqueous phase allows the dispersion of oil by increasing the matrix viscosity and provides stabilization of the oil-water interface by forming a dense polyelectrolitic brush on the water side of this interface. We characterize emulsions with varying oil content, and concentrations of chitosan, and follow their long-term stability. Finally, we show that by precipitating the chitosan the rigid elastic network is formed in the aqueous phase, making a very stable suspension.