Biomedical subjects

J Wouters

Publications and source records attributed to J Wouters.

At least 19 recordsLinked to original sources

PubMed2005

Differential scanning calorimetry in life science: thermodynamics, stability, molecular recognition and application in drug design.

All biological phenomena depend on molecular recognition, which is either intermolecular like in ligand binding to a macromolecule or intramolecular like in protein folding. As a result, understanding the relationship between the structure of proteins and the energetics of their stability and binding with others (bio)molecules is a very interesting point in biochemistry and biotechnology. It is essential to the engineering of stable proteins and to the structure-based design of pharmaceutical ligands. The parameter generally used to characterize the stability of a system (the folded and unfolded state of the protein for example) is the equilibrium constant (K) or the free energy (deltaG(o)), which is the sum of enthalpic (deltaH(o)) and entropic (deltaS(o)) terms. These parameters are temperature dependent through the heat capacity change (deltaCp). The thermodynamic parameters deltaH(o) and deltaCp can be derived from spectroscopic experiments, using the van't Hoff method, or measured directly using calorimetry. Along with isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC) is a powerful method, less described than ITC, for measuring directly the thermodynamic parameters which characterize biomolecules. In this article, we summarize the principal thermodynamics parameters, describe the DSC approach and review some systems to which it has been applied. DSC is much used for the study of the stability and the folding of biomolecules, but it can also be applied in order to understand biomolecular interactions and can thus be an interesting technique in the process of drug design.

Calorimetry, Differential Scanning↗

PubMed2004

Ultralow-loss 3-dB photonic crystal waveguide splitter.

A photonic crystal waveguide splitter that exhibits ultralow-loss 3-dB splitting for TE-polarized light is fabricated in silicon-on-insulator material by use of deep UV lithography. The high performance is achieved by use of a Y junction, which is designed to ensure single-mode operation, and low-loss 60 degrees bends. Zero-loss 3-dB output is experimentally obtained in the range 1560-1585 nm. Results from three-dimensional finite-difference time-domain modeling with no adjustable parameters are found to be in excellent agreement with the experimental results.

Journal Article↗

PubMed2004

Crystal structure of the C67A mutant of isopentenyl diphosphate isomerase complexed with a mechanism-based irreversible inhibitor.

Isopentenyl diphosphate:dimethylallyl diphosphate (IPP:DMAPP) isomerase is a key enzyme in the biosynthesis of isoprenoids. The mechanism of the isomerization reaction involves protonation of the unactivated carbon-carbon double bond in the substrate. Analysis of the 1.97 A crystal structure of the inactive C67A mutant of E. coli isopentenyl diphosphate:dimethylallyl diphosphate isomerase complexed with the mechanism-based inactivator 3,4-epoxy-3-methyl-1-butyl diphosphate is in agreement with an isomerization mechanism involving Glu 116, Tyr 104, and Cys 67. In particular, the results are consistent with a mechanism where Glu116 is involved in the protonation step and Cys67 in the elimination step.

Binding Sites↗

PubMed2003

Catalytic mechanism of Escherichia coli isopentenyl diphosphate isomerase involves Cys-67, Glu-116, and Tyr-104 as suggested by crystal structures of complexes with transition state analogues and irreversible inhibitors.

Isopentenyl diphosphate (IPP):dimethylallyl diphosphate (DMAPP) isomerase is a key enzyme in the biosynthesis of isoprenoids. The reaction involves protonation and deprotonation of the isoprenoid unit and proceeds through a carbocationic transition state. Analysis of the crystal structures (2 A) of complexes of Escherichia coli IPP.DMAPPs isomerase with a transition state analogue (N,N-dimethyl-2-amino-1-ethyl diphosphate) and a covalently attached irreversible inhibitor (3,4-epoxy-3-methyl-1-butyl diphosphate) indicates that Glu-116, Tyr-104, and Cys-67 are involved in the antarafacial addition/elimination of protons during isomerization. This work provides a new perspective about the mechanism of the reaction.

Carbon-Carbon Double Bond Isomerases↗

PubMed2003

Crystal structure of Enterobacter cloacae 908R class C beta-lactamase bound to iodo-acetamido-phenyl boronic acid, a transition-state analogue.

The structures of the class C beta-lactamase from Enterobacter cloacae 908R alone and in complex with a boronic acid transition-state analogue were determined by X-ray crystallography at 2.1 and 2.3 A, respectively. The structure of the enzyme resembles those of other class C beta-lactamases. The structure of the complex with the transition-state analogue, iodo-acetamido-phenyl boronic acid, shows that the inhibitor is covalently bound to the active-site serine (Ser64). Binding of the inhibitor within the active site is compared with previously determined structures of complexes with other class C enzymes. The structure of the boronic acid adduct indicates ways to improve the affinity of this class of inhibitors. This structure of 908R class C beta-lactamase in complex with a transition-state analogue provides further insights into the mechanism of action of these hydrolases.

Apoenzymes↗

PubMed2003

Crystallography of membrane proteins, major targets in drug design.

Protein crystallography has the potential to accelerate drug discovery greatly. High-resolution structures of membrane proteins of pharmaceutical interest open new perspectives in drug design. Recent structural data obtained for cyclooxygenases, monoamine oxidase, squalene cyclase, rhodopsine, porins, aquaporins, and ABC transporters are presented and briefly discussed.

Crystallography, X-Ray↗

PubMed2002

Polymorphism of N-stearoylethanolamine: differential scanning calorimetric, vibrational spectroscopic (FTIR), and crystallographic studies.

Based on a series of physicochemical properties (differential scanning calorimetry, powder X-ray crystallographic studies and Fourier-transform infra red spectroscopic analysis) determined for N-stearoylethanolamine (NSEA) (C18:0) at different temperatures, evidence has been given that this compound can exist in (at least) three polymorphic forms. Powder X-ray crystallography clearly demonstrates the presence of three distinct molecular packings at distinct temperatures while spectral changes in the vibrational spectra reveal that the geometry of the CH(2)z.sbnd;CO functional group of the molecule is affected during the polymorphic transitions. Rationalization of the thermal physicochemical behavior of NSEA in terms of molecular packing is also proposed. It supposes rearrangement of the hydrocarbon chains upon heating of the molecule.

Calorimetry, Differential Scanning↗

PubMed2002

Digital hearing aids and future directions for hearing aids.

In recent years many technical evolutions have been applied in hearing aids. In this paper differences between analog, programmable and fully digital hearing aids, the basic and supplementary functions of a hearing aid, and some important issues and future directions for digital hearing aids will be mentioned.

Computers↗

PubMed2001

Crystallographic analysis of family 11 endo-beta-1,4-xylanase Xyl1 from Streptomyces sp. S38.

Family 11 endo-beta-1,4-xylanases degrade xylan, the main constituent of plant hemicelluloses, and have many potential uses in biotechnology. The structure of Xyl1, a family 11 endo-xylanase from Streptomyces sp. S38, has been solved. The protein crystallized from ammonium sulfate in the trigonal space group P321, with unit-cell parameters a = b = 71.49, c = 130.30 A, gamma = 120.0 degrees. The structure was solved at 2.0 A by X-ray crystallography using the molecular-replacement method and refined to a final R factor of 18.5% (R(free) = 26.9%). Xyl1 has the overall fold characteristic of family 11 xylanases, with two highly twisted beta-sheets defining a long cleft containing the two catalytic residues Glu87 and Glu177.

Amino Acid Sequence↗

PubMed2001

Psychophysical evidence for a general temporal processing deficit in children with dyslexia.

The hypothesis of a general (i.e. cross-modal) temporal processing deficit in dyslexia was tested by examining rapid processing in both the auditory and the visual system in the same children with dyslexia. Participants were 10- to 12-year-old dyslexic readers and age-matched normal reading controls. Psychophysical thresholds were estimated for auditory gap and visual double flash detection, using a two-interval, two-alternative forced-choice paradigm. Significant group differences were found for the auditory and the visual test. Furthermore, temporal processing measures were significantly related to word and pseudo-word reading skills. As 70% of the dyslexic readers had significantly higher thresholds than controls for both auditory and visual temporal processing, the evidence tends to support the hypothesis of a general temporal processing deficit in children with dyslexia.

Auditory Perception↗

PubMed2001

Structure determination and comparison of BM567, a sulfonylurea, with terbogrel, two compounds with dual action, thromboxane receptor antagonism and thromboxane synthase inhibition.

BM567, a sulfonylurea compound whose crystal structure is here discussed and terbogrel, are both thromboxane receptor antagonists and thromboxane synthase inhibitors. In this paper, their crystallographic and electronic structures are compared and lead to new synthesis prospects among the sulfonylurea series.

Crystallography↗

PubMed2001

Synthesis and anticonvulsant activity of N,N-phthaloyl derivatives of central nervous system inhibitory amino acids.

In order to study the influence of the length of the amino acid chain of N,N-phthaloyl-amino acid amides as analogues of the former anticonvulsant taltrimide on the seizure-antagonizing activity glycine, beta-alanine and gamma-aminobutyric acid (GABA) derivatives were synthesized. The corresponding taurine derivatives were also included. Generally, the glycine-derived amides showed a higher activity than the beta-alanine and GABA derivatives in the maximal electroshock seizure (MES) test in mice upon intraperitoneal administration. The activity was comparable to the respective taurine derivatives. The N,N-phthaloyl-glycine amides were also active in the MES test upon oral administration to rats. No significant activity was noted in the seizure threshold test with subcutaneous pentylene-tetrazole. The ED50 of N,N-phthaloyl-glycine ethyl amide (4b) in the MES test upon intraperitoneal administration to mice was 19.1 mg/kg. On a molar basis this activity is comparable to the activity of phenytoin with little toxicity in the rotorod test. In conclusion, N,N-phthaloyl-glycine amides might represent promising antiepileptic drugs.

Amino Acids↗

PubMed2001

Substrate binding and catalytic mechanism of class B beta-lactamases: a molecular modelling study.

Increased resistance to beta-lactam antibiotics is mainly due to beta-lactamases whose production by pathogenic bacteria makes their broad activity spectrum especially frightening. X-ray structures of several zinc beta-lactamases have revealed the coordination of the two metal ions, but their mode of action remains unclear. Geometry optimisation of stable complexes along the reaction pathway of benzylpenicillin hydrolysis highlighted a proton shuttle occurring from D120 of the Bacillus cereus beta-lactamase to the beta-lactam nitrogen via Zn2 which is central to the network. First, the Zn1 ion has a structural role maintaining Zn-bound waters, WAT1 and WAT2, either directly or through the Zn1 tetrahedrally coordinated histidine ligands. The Zn2 ion has a more catalytic role, stabilising the tetrahedral intermediate, accepting the beta-lactam nitrogen atom as a ligand. The role of Zn2 and the flexibility in the coordination geometry of both Zn ions is of crucial importance for catalysis.

Bacillus cereus↗

PubMed2001

Speech recognition in noise for cochlear implantees with a two-microphone monaural adaptive noise reduction system.

OBJECTIVE: In this study the performance of a noise reduction strategy applied to cochlear implants is evaluated. The noise reduction strategy is based on a 2-channel adaptive filtering strategy using two microphones in a single behind-the-ear hearing aid. DESIGN: Four adult LAURA cochlear implant users (Peeters et al., 1993) took part in the experiments. The tests included identification of monosyllabic CVC (consonant-vowel-consonant) words and measurements of the speech reception threshold (SRT) of lists of numbers, in background noise presented at 90 degrees relative to the 0 degrees frontal direction of the speech. Percent correct phoneme scores for the CVC words at signal to noise ratios (SNRs) of -5, 0, and +5 dB in steady speech-weighted noise at 60 dB SPL and SRTs for numbers in speech-weighted steady and nonsteady ICRA noise were both obtained in conditions with and without the noise reduction pre-processing. Physical SNR improvements of the noise reduction system are evaluated as well, as a function of the direction of the noise source. RESULTS: Highly significant improvements in speech understanding, corresponding on average to an SNR improvement of about 10 dB, were observed with this 2-channel adaptive filtering noise reduction strategy using both types of speech-noise test materials. These perceptual evaluations agree with physical evaluations and simulations of this noise reduction strategy. Taken together, these data demonstrate that cochlear implantees may increase their speech intelligibility in noisy environments with the use of multimicrophone noise reduction systems.

Adult↗

PubMed2001

Comparison of procedures to determine electrical stimulation thresholds in cochlear implant users.

OBJECTIVE: This study was designed to evaluate threshold estimation procedures for cochlear implantees, both for clinical and research purposes. Precision and testing time of eight procedures were evaluated. These procedures included three variations of two adaptive tasks and two variations of an adjustment procedure. DESIGN: Electrical thresholds were obtained with eight different procedures. For each procedure, one of three tasks was combined with one of four decision strategies. With the adjustment task, current level was adjusted from "inaudible" to "just audible" or from "clearly audible" to "just audible." With the two adaptive tasks (i.e., "count the number of pulses" and "choose the interval with the pulse"), current level was adjusted during the sequence of trials by subjects' preceding responses according to three decision criteria (Levitt, 1971). Experimental data were collected on both an apical and a basal channel of six Laura cochlear implantees, and test-retest measures were obtained for each procedure, channel and subject. Moreover, total testing time was recorded in each test, to examine the trade-off with precision, and to investigate the feasibility of the procedures in a clinical setting. RESULTS: In general, the "count the pulses" procedures yield higher threshold values than the "choose the interval with the pulse" and adjustment procedures. Precision was not a statistically significant factor, although the test-retest variation was approximately a factor of 2 larger in the adjustment procedures than in the adaptive procedures. Test duration differences were large. Moreover, analyses of the adaptive procedures showed that a stable estimate requires at least six reversals and that the first four reversals should not be taken into account for threshold estimation. CONCLUSIONS: In a clinical setting, where precision and time are important parameters, precise thresholds can be obtained with the adjustment procedure. However, for research purposes, it is advised to use the "choose the interval with the pulse" task together with one of the transformed up-down decision criteria.

Adolescent↗

PubMed2001

Expression, purification, crystallization and preliminary X-ray analysis of the native class C beta-lactamase from Enterobacter cloacae 908R and two mutants.

Crystals have been obtained of the Enterobacter cloacae 908R beta-lactamase and two point mutants by the vapour-diffusion method using similar conditions [pH 9.0, polyethylene glycol (M(r) = 6000) as precipitant]. The three crystal forms belong to the orthorhombic space group P2(1)2(1)2, with roughly the same unit-cell parameters; i.e. for the wild-type crystals a = 46.46, b = 82.96, c = 95.31 A. In the best cases, the crystals diffract to about 2.1 A resolution on a rotating-anode X-ray source at room temperature. Co-crystallization experiments of poor substrates with the wild-type protein and the active-site serine mutant (S64C) are planned and should lead to a better understanding of the catalytic mechanism of class C beta-lactamases.

Crystallization↗

PubMed2001

Coding of the fundamental frequency in continuous interleaved sampling processors for cochlear implants.

In this study the perception of the fundamental frequency (F0) of periodic stimuli by cochlear implant users is investigated. A widely used speech processor is the Continuous Interleaved Sampling (CIS) processor, for which the fundamental frequency appears as temporal fluctuations in the envelopes at the output. Three experiments with four users of the LAURA (Registered trade mark of Philips Hearing Implants, now Cochlear Technology Centre Europe) cochlear implant were carried out to examine the influence of the modulation depth of these envelope fluctuations on pitch discrimination. In the first experiment, the subjects were asked to discriminate between two SAM (sinusoidally amplitude modulated) pulse trains on a single electrode channel differing in modulation frequency ( deltaf = 20%). As expected, the results showed a decrease in the performance for smaller modulation depths. Optimal performance was reached for modulation depths between 20% and 99%, depending on subject, electrode channel, and modulation frequency. In the second experiment, the smallest noticeable difference in F0 of synthetic vowels was measured for three algorithms that differed in the obtained modulation depth at the output: the default CIS strategy, the CIS strategy in which the F0 fluctuations in the envelope were removed (FLAT CIS), and a third CIS strategy, which was especially designed to control and increase the depth of these fluctuations (F0 CIS). In general, performance was poorest for the FLAT CIS strategy, where changes in F0 are only apparent as changes of the average amplitude in the channel outputs. This emphasizes the importance of temporal coding of F0 in the speech envelope for pitch perception. No significantly better results were obtained for the F0 CIS strategy compared to the default CIS strategy, although the latter results in envelope modulation depths at which sub-optimal scores were obtained in some cases of the first experiment. This indicates that less modulation is needed if all channels are stimulated with synchronous F0 fluctuations. This hypothesis is confirmed in a third experiment where subjects performed significantly better in a pitch discrimination task with SAM pulse trains, if three channels were stimulated concurrently, as opposed to only one.

Acoustic Stimulation↗

PubMed2001

Analysis of the role of OpuC, an osmolyte transport system, in salt tolerance and virulence potential of Listeria monocytogenes.

The success of Listeria monocytogenes as a food-borne pathogen owes much to its ability to survive a variety of stresses, both in the external environment prior to ingestion and subsequently within the animal host. Growth at high salt concentrations and low temperatures is attributed mainly to the accumulation of organic solutes such as glycine betaine and carnitine. We utilized a novel system for generating chromosomal mutations (based on a lactococcal pWVO1-derived Ori(+) RepA(-) vector, pORI19) to identify a listerial OpuC homologue. Mutating the operon in two strains of L. monocytogenes revealed significant strain variation in the observed activity of OpuC. Radiolabeled osmolyte uptake studies, together with growth experiments in defined media, linked OpuC to carnitine and glycine betaine uptake in Listeria. We also investigated the role of OpuC in contributing to the growth and survival of Listeria in an animal (murine) model of infection. Altering OpuC resulted in a significant reduction in the ability of Listeria to colonize the upper small intestine and cause subsequent systemic infection following peroral inoculation.

ATP-Binding Cassette Transporters↗