397 research outputs found
Structural change in a B-DNA helix with hydrostatic pressure
Study of the effects of pressure on macromolecular structure improves our understanding of the forces governing structure, provides details on the relevance of cavities and packing in structure, increases our understanding of hydration and provides a basis to understand the biology of high-pressure organisms. A study of DNA, in particular, helps us to understand how pressure can affect gene activity. Here we present the first high-resolution experimental study of B-DNA structure at high pressure, using NMR data acquired at pressures up to 200 MPa (2 kbar). The structure of DNA compresses very little, but is distorted so as to widen the minor groove, and to compress hydrogen bonds, with AT pairs compressing more than GC pairs. The minor groove changes are suggested to lead to a compression of the hydration water in the minor groove
MultiSig: a new high-precision approach to the analysis of complex biomolecular systems
MultiSig is a newly developed mode of analysis of sedimentation equilibrium (SE) experiments in the analytical ultracentrifuge, having the capability of taking advantage of the remarkable precision (~0.1 % of signal) of the principal optical (fringe) system employed, thus supplanting existing methods of analysis through reducing the ‘noise’ level of certain important parameter estimates by up to orders of magnitude. Long-known limitations of the SE method, arising from lack of knowledge of the true fringe number in fringe optics and from the use of unstable numerical algorithms such as numerical differentiation, have been transcended. An approach to data analysis, akin to ‘spatial filtering’, has been developed, and shown by both simulation and practical application to be a powerful aid to the precision with which near-monodisperse systems can be analysed, potentially yielding information on protein-solvent interaction. For oligo- and poly-disperse systems the information returned includes precise average mass distributions over both cell radial and concentration ranges and mass-frequency histograms at fixed radial positions. The application of MultiSig analysis to various complex heterogenous systems and potentially multiply-interacting carbohydrate oligomers is described
Investigation of bovine serum albumin denaturation using ultrasonic spectroscopy
This is the author’s version of a work that was accepted for publication in the journal Food Hydrocolloids. Changes resulting from the publishing process, such as peer review, editing, corrections, structural formatting, and other quality control mechanisms may not be reflected in this document. Changes may have been made to this work since it was submitted for publication. A definitive version was subsequently published at: http://dx.doi.org/10.1016/j.foodhyd.2010.11.011The ability of ultrasound spectroscopy to characterise protein denaturation at relatively high
concentrations and under conditions found in foods, is examined. Measurement of longitudinal sound
velocity against concentration and frequency (20-160 MHz) for the bovine serum albumin monomer at
pH 7.0 gave a frequency independent value for molecular compressibility of at 25 °C, corresponding to
a sound velocity for the BSA molecule of 1920 ms-1. At 160 MHz, the longitudinal sound attenuation in
BSA molecules is ~5200 Npm-1, a factor of 10 higher than in water. The excess attenuation of the
solution over water was nearly 90 Npm-1 at the highest measured volume fraction of 0.03 (or 3% v/v).
Concentration-dependent ultrasound velocity (20 - 160 MHz) and attenuation (2 - 120 MHz) spectra
were obtained over time for heated bovine serum albumin (BSA) solutions up to 40 mg/mL at neutral
pH and at 25 °C. An acoustic scattering model was used which considered the solute molecules as
scatterers of ultrasound, to determine the molecules' sound velocity, compressibility, and attenuation
properties. Mild heat treatment caused the molecule to organise into dimers and trimers, without
change in sound velocity; implying that there is little or no change in secondary structure. Changes in
attenuation spectra correlated with estimated molecular weight as determined through DLS and SEC
measurements. During oligomerisation, the BSA molecules continue to behave acoustically as
monomers.
Under severe heat treatment, BSA rapidly suffered irreversible denaturation and gelation occurred
which affected both ultrasound attenuation spectra and the velocity of sound, consistent with
significant molecular conformation changes and/or molecule-molecule interactions
Folding Thermodynamics of the Hybrid-1 Type Intramolecular Human Telomeric GQuadruplex
Guanine-rich DNA sequences that may form
G-quadruplexes are located in strategic DNA loci with
the ability to regulate biological events. G-quadruplexes
have been under intensive scrutiny owing to their potential
to serve as novel drug targets in emerging anticancer
strategies. Thermodynamic characterization of
G-quadruplexes is an important and necessary step in
developing predictive algorithms for evaluating the conformational
preferences of G-rich sequences in the presence
or the absence of their complementary C-rich
strands. We use a combination of spectroscopic, calorimetric,
and volumetric techniques to characterize the
folding/unfolding transitions of the 26-meric human telomeric
sequence d[A3G3(T2AG3)3A2]. In the presence of
K1 ions, the latter adopts the hybrid-1 G-quadruplex
conformation, a tightly packed structure with an unusually
small number of solvent-exposed atomic groups. The
K1-induced folding of the G-quadruplex at room temperature
is a slow process that involves significant accumulation
of an intermediate at the early stages of the
transition. The G-quadruplex state of the oligomeric
sequence is characterized by a larger volume and compressibility
and a smaller expansibility than the coil state.
These results are in qualitative agreement with each other
all suggesting significant dehydration to accompany the
G-quadruplex formation. Based on our volume data,
432619 water molecules become released to the bulk
upon the G-quadruplex formation. This large number is
consistent with a picture in which DNA dehydration is
not limited to water molecules in direct contact with the
regions that become buried but involves a general
decrease in solute–solvent interactions all over the surface
of the folded structure. VC 2013 Wiley Periodicals, Inc.
Biopolymers 101: 216–227, 2014.
Keywords: G-quadruplexes; conformational transitions;
volume; compressibility; expansibilit
Stability of intramolecular quadruplexes: sequence effects in the central loop
Hundreds of thousands of putative quadruplex sequences have been found in the human genome. It is important to understand the rules that govern the stability of these intramolecular structures. In this report, we analysed sequence effects in a 3-base-long central loop, keeping the rest of the quadruplex unchanged. A first series of 36 different sequences were compared; they correspond to the general formula GGGTTTGGGHNHGGGTTTGGG. One clear rule emerged from the comparison of all sequence motifs: the presence of an adenine at the first position of the loop was significantly detrimental to stability. In contrast, adenines have no detrimental effect when present at the second or third position of the loop. Cytosines may either have a stabilizing or destabilizing effect depending on their position. In general, the correlation between the Tm or ΔG° in sodium and potassium was weak. To determine if these sequence effects could be generalized to different quadruplexes, specific loops were tested in different sequence contexts. Analysis of 26 extra sequences confirmed the general destabilizing effect of adenine as the first base of the loop(s). Finally, analysis of some of the sequences by microcalorimetry (DSC) confirmed the differences found between the sequence motifs
Hydrostatic and osmotic pressure study of the RNA hydration
The tertiary structure of nucleic acids results from an equilibrium between electrostatic interactions of phosphates, stacking interactions of bases, hydrogen bonds between polar atoms and water molecules. Water interactions with ribonucleic acid play a key role in its structure formation, stabilization and dynamics. We used high hydrostatic pressure and osmotic pressure to analyze changes in RNA hydration. We analyzed the lead catalyzed hydrolysis of tRNAPhe from S. cerevisiae as well as hydrolytic activity of leadzyme. Pb(II) induced hydrolysis of the single phosphodiester bond in tRNAPhe is accompanied by release of 98 water molecules, while other molecule, leadzyme releases 86
Energetic signatures of single base bulges: thermodynamic consequences and biological implications
DNA bulges are biologically consequential defects that can arise from template-primer misalignments during replication and pose challenges to the cellular DNA repair machinery. Calorimetric and spectroscopic characterizations of defect-containing duplexes reveal systematic patterns of sequence-context dependent bulge-induced destabilizations. These distinguishing energetic signatures are manifest in three coupled characteristics, namely: the magnitude of the bulge-induced duplex destabilization (ΔΔGBulge); the thermodynamic origins of ΔΔGBulge (i.e. enthalpic versus entropic); and, the cooperativity of the duplex melting transition (i.e. two-state versus non-two state). We find moderately destabilized duplexes undergo two-state dissociation and exhibit ΔΔGBulge values consistent with localized, nearest neighbor perturbations arising from unfavorable entropic contributions. Conversely, strongly destabilized duplexes melt in a non-two-state manner and exhibit ΔΔGBulge values consistent with perturbations exceeding nearest-neighbor expectations that are enthalpic in origin. Significantly, our data reveal an intriguing correlation in which the energetic impact of a single bulge base centered in one strand portends the impact of the corresponding complementary bulge base embedded in the opposite strand. We discuss potential correlations between these bulge-specific differential energetic profiles and their overall biological implications in terms of DNA recognition, repair and replication
Stacking Interactions in Denaturation of DNA Fragments
A mesoscopic model for heterogeneous DNA denaturation is developed in the
framework of the path integral formalism. The base pair stretchings are treated
as one-dimensional, time dependent paths contributing to the partition
function. The size of the paths ensemble, which measures the degree of
cooperativity of the system, is computed versus temperature consistently with
the model potential physical requirements. It is shown that the ensemble size
strongly varies with the molecule backbone stiffness providing a quantitative
relation between stacking and features of the melting transition. The latter is
an overall smooth crossover which begins from the \emph{adenine-thymine} rich
portions of the fragment. The harmonic stacking coupling shifts, along the
-axis, the occurrence of the multistep denaturation but it does not change
the character of the crossover. The methods to compute the fractions of open
base pairs versus temperature are discussed: by averaging the base pair
displacements over the path ensemble we find that such fractions signal the
multisteps of the transition in good agreement with the indications provided by
the specific heat plots.Comment: European Physical Journal E (2011) in pres
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