Biomedical subjects

J Crain

Publications and source records attributed to J Crain.

At least 19 recordsLinked to original sources

PubMed2006

Time-resolved FRET and FLIM of four-way DNA junctions.

Conformational transitions in a 4-way DNA junction when titrated with ionic solutions are studied using time-resolved fluorescence resonance energy transfer. Parameters characterising the transition in terms of critical ion concentration (c1/2) and the Hill coefficient for ion binding are obtained by fitting a simple two-state model using steady-state spectra. Data obtained from a fluorescence lifetime plate reader and analysed by fitting a single exponential to donor fluorescence lifetime decays are shown to be in good agreement with the parameters obtained from steady-state measurements. Fluorescence lifetimes, however, offer advantages, particularly in being independent of fluorophore concentration, output intensity, inhomogeneity in the excitation source and output wavelength. We demonstrate preliminary FRET-FLIM images of DNA junction solutions obtained using a picosecond gated CCD which are in agreement with results from a fluorescence lifetime plate reader. The results suggest that time-resolved FRET-FLIM is sensitive to subtle structural changes and may be useful in assays based on 4-way DNA junctions.

DNA, Cruciform↗

PubMed2006

The stability and characteristics of a DNA Holliday junction switch.

A Holliday junction (HJ) consists of four DNA double helices, with a branch point discontinuity at the intersection of the component strands. At low ionic strength, the HJ adopts an open conformation, with four widely spaced arms, primarily due to strong electrostatic repulsion between the phosphate groups on the backbones. At high ionic strength, screening of this repulsion induces a switch to a more compact (closed) junction conformation. Fluorescent labelling with dyes placed on the HJ arms allows this conformational switch to be detected optically using fluorescence resonance energy transfer (FRET), producing a sensitive fluorescent output of the switch state. This paper presents a systematic and quantitative survey of the switch characteristics of such a labelled HJ. A short HJ (arm length 8 bp) is shown to be prone to dissociation at low switching ion concentration, whereas an HJ of arm length 12 bp is shown to be stable over all switching ion concentrations studied. The switching characteristics of this HJ have been systematically and quantitatively studied for a variety of switching ions, by measuring the required ion concentration, the sharpness of the switching transition and the fluorescent output intensity of the open and closed states. This stable HJ is shown to have favourable switch characteristics for a number of inorganic switching ions, making it a promising candidate for use in nanoscale biomolecular switch devices.

DNA, Cruciform↗

PubMed2006

Structural properties of liquid N-methylacetamide via ab initio, path integral, and classical molecular dynamics.

In order to better understand the physical interactions that stabilize protein secondary structure, the neat liquid state of a peptidic fragment, N-methylacetamide (NMA), was studied using computer simulation. Three different descriptions of the molecular liquid were examined: an empirical force field treatment with classical nuclei, an empirical force field treatment with quantum mechanical nuclei, and an ab initio density functional theory (DFT) treatment. The DFT electronic structure was evaluated using the BLYP approximate functional and a plane wave basis set. The different physical effects probed by the three models, such as quantum dispersion, many-body polarization, and nontrivial charge distributions on the liquid properties, were compared. Much of the structural ordering in the liquid is characterized by hydrogen bonded chains of NMA molecules. Modest structural differences are present among the three models of liquid NMA. The average molecular dipole in the liquid under the ab initio treatment, however, is enhanced by 60% over the gas phase value.

Acetamides↗

PubMed2006

Structure and hydrogen bonding in neat N-methylacetamide: classical molecular dynamics and Raman spectroscopy studies of a liquid of peptidic fragments.

The results of classical molecular dynamics (MD) simulations and Raman spectroscopy studies of neat liquid N-methylacetamide (NMA), the simplest model system relevant to the peptides, are reported as a function of temperature and pressure. The MD simulations predict that near ambient conditions, the molecules form a hydrogen bond network consisting primarily of linear chains. Both the links between molecules within the hydrogen-bonded chains and the associations between chains are stabilized by weak methyl-donated "improper" hydrogen bonds. The three-dimensional structural motifs observed in the liquid show some similarity to protein beta-sheets. The temperature and pressure dependence of the hydrogen bond network, as probed by the mode frequency of the experimentally determined amide-I Raman band, blue shifts on heating and red shifts under compression, respectively, suggesting weakened and enhanced hydrogen bonding in response to temperature and pressure increases. Disruption of the hydrogen-bonding network is clearly observed in the simulation data as temperature is increased, whereas the improper hydrogen bonding is enhanced under compression to reduce the energetic cost of increasing the packing fraction. Because of the neglect of polarizability in the molecular model, the computed dielectric constant is underestimated compared to the experimental value, indicating that the simulation may underestimate dipolar coupling in the liquid.

Acetamides↗

PubMed2005

Segregation in aqueous methanol enhanced by cooling and compression.

Molecular segregation in methanol-water mixtures is studied across a wide concentration range as a function of temperature and pressure. Cluster distributions obtained from both neutron diffraction and molecular dynamics simulations point to significantly enhanced segregation as the mixtures are cooled or compressed. This evolution toward greater molecular heterogenity in the mixture accounts for the observed changes in the water-water radial distribution function and there are indications also of a change in the topology of the water clusters. The observed behavior is consistent with an approach to an upper critical solution point. Such a point would appear to be "hidden" below the freezing line, thereby precluding observation of the two-fluid region.

Journal Article↗

PubMed2004

Methanol-water solutions: a bi-percolating liquid mixture.

An extensive series of neutron diffraction experiments and molecular dynamics simulations has shown that mixtures of methanol and water exhibit extended structures in solution despite the components being fully miscible in all proportions. Of particular interest is a concentration region (methanol mole fraction between 0.27 and 0.54) where both methanol and water appear to form separate, percolating networks. This is the concentration range where many transport properties and thermodynamic excess functions reach extremal values. The observed concentration dependence of several of these material properties of the solution may therefore have a structural origin.

Journal Article↗

PubMed2004

Single molecule fluorescence imaging and its application to the study of DNA condensation.

Single molecule fluorescence imaging incorporated with optical tweezers and a laminar flow cell has been used to monitor the kinetic process of DNA condensation induced by spermidine. It was found that at least two steps were involved in the condensation process of the hydrodynamically-stretched linear DNA; a lag period followed by a rapid collapse of DNA. The lag time increased with the flow speed and the collapse time remained short within the range of the flow speed studied. The effect of salt concentration on the condensation process was examined, and the results suggest that the longer lag time observed in the higher salt buffer probably results from the displacement of bound cations and rearrangement of spermidine on the DNA. The flow-speed dependence of the lag time suggests that a nucleation event at the free end of the DNA, i.e. formation of a loop, may play a vital role in the kinetic process of condensation.

DNA↗

PubMed2002

Molecular segregation observed in a concentrated alcohol-water solution.

When a simple alcohol such as methanol or ethanol is mixed with water, the entropy of the system increases far less than expected for an ideal solution of randomly mixed molecules. This well-known effect has been attributed to hydrophobic headgroups creating ice-like or clathrate-like structures in the surrounding water, although experimental support for this hypothesis is scarce. In fact, an increasing amount of experimental and theoretical work suggests that the hydrophobic headgroups of alcohol molecules in aqueous solution cluster together. However, a consistent description of the details of this self-association is lacking. Here we use neutron diffraction with isotope substitution to probe the molecular-scale structure of a concentrated alcohol water mixture (7:3 molar ratio). Our data indicate that most of the water molecules exist as small hydrogen-bonded strings and clusters in a 'fluid' of close-packed methyl groups, with water clusters bridging neighbouring methanol hydroxyl groups through hydrogen bonding. This behaviour suggests that the anomalous thermodynamics of water alcohol systems arises from incomplete mixing at the molecular level and from retention of remnants of the three-dimensional hydrogen-bonded network structure of bulk water.

Journal Article↗

PubMed2000

Colloid-liquid-crystal composites: An unusual soft solid

Mixing model colloidal particles with a thermotropic nematic liquid crystal results in a soft solid with significant storage modulus (G' approximately 10(3)-10(5) Pa). The soft solid comprises a network of particle aggregates, formed by the exclusion of particles from emergent nematic domains as the mixture is cooled below the isotropic-nematic transition. The unusually high storage modulus of the colloid-liquid-crystal composites may be due to the local frustration of nematic order within the particle aggregates. The birefringent soft solid is potentially important as a switchable electro-optical material that can be readily handled and processed.

Journal Article↗

PubMed1999

Mitral regurgitation after atrioventricular node catheter ablation for atrial fibrillation and heart failure: acute hemodynamic features.

BACKGROUND: Radiofrequency catheter ablation of the atrioventricular node and pacemaker insertion have been associated with occasional development of mitral regurgitation (MR). Ventricular pacing might result in MR if (1) left ventricular (LV) compliance is decreased and/or (2) mitral valve leaflet apposition is disturbed. We studied acute hemodynamic changes resulting from initiation of ventricular pacing in patients undergoing ablation. METHODS AND RESULTS: Thirteen patients (10 men and 3 women) with a mean age of 73.4 +/- 8. 6 years, with chronic atrial fibrillation and congestive heart failure, had permanent ventricular pacemaker insertion with lead placement at the right ventricular (RV) apex. The following hemodynamic recordings were obtained before ablation (during atrial fibrillation) and then immediately after ablation (during RV pacing): heart rate, mean arterial pressure, LV end-diastolic pressure (LVEDP), mean pulmonary capillary wedge pressure, V-wave amplitude, and cardiac index. Presence of MR was assessed by V-wave amplitude and the results of LV angiography. In patients who had MR, recordings were also obtained during temporary ventricular pacing from the RV outflow tract (RVOT). As a group there were no significant changes in any hemodynamic measurement. Before ablation, mild MR by LV angiogram was present in 5 patients, but none had large V-wave amplitude. After ablation, mild MR was present by LV angiogram in 6 patients, and in 3 of these patients large V-wave amplitude developed (mean amplitude 42.7 +/- 2.2 mm Hg; assigned to group 1). This was associated with a rise in LVEDP in 1 patient (consistent with reduced LV compliance), but LVEDP was unchanged in the other 2 patients (suggesting abnormal mitral valve leaflet apposition). All patients in group 1 exhibited a fall in V-wave amplitude when the pacing site was moved to the RVOT. CONCLUSIONS: Both reduced LV compliance and disturbed mitral valve leaflet apposition contribute to MR after ablation. MR is reduced by pacing from the RVOT.

Aged↗

PubMed1997

Effect of antiphospholipid antibodies in women undergoing in-vitro fertilization: role of heparin and aspirin.

To describe the prevalence of antiphospholipid antibodies in women undergoing in-vitro fertilization (IVF) and to determine if heparin and aspirin affect implantation rates, 191 women with a history of infertility undergoing IVF were prospectively tested for antiphospholipid antibodies. This was a two-centre, non-randomized comparison of women with positive antiphospholipid antibodies receiving heparin and aspirin versus standard treatment. An enzyme-linked immunosorbent assay, with referenced standards and known positive and negative sera on each plate, was utilized to measure antibodies to cardiolipin, phosphatidylinositol, phosphatidylglycerol, phosphatidylserine and phosphatidylethanolamine. Statistical analyses of results included analysis of variance and Fisher's two-tailed exact test. Antiphospholipid antibodies were detected in 18.8% of patients undergoing IVF compared with only 5.5% in the 200 normal controls, 26% in 200 women with recurrent pregnancy loss, and 32% in 200 women with systemic lupus erythematosus. In conclusion, antiphospholipid antibodies were found more frequently in women undergoing IVF than in the normal control population. Although implantation rates appeared higher in the group of women treated with heparin and aspirin, no statistically significant differences were detected in implantation, pregnancy and ongoing pregnancy rates between those who received standard therapy and those treated with heparin and aspirin.

Adult↗

PubMed1996