SEARCH · PubMed Health

Results for “Unique interface”

Explore indexed PubMed citations for clinical trials, systematic reviews and public health research. Read source abstracts and follow each citation to its original PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,387 records · Page 77Linked to original sources

PubMedDate not supplied

P-Mod: an algorithm and software to map modifications to peptide sequences using tandem MS data.

The discovery of unanticipated protein modifications is one of the most challenging problems in proteomics. Whereas widely used algorithms such as Sequest and Mascot enable mapping of modifications when the mass and amino acid specificity are known, unexpected modifications cannot be identified with these tools. We have developed an algorithm and software called P-Mod, which enables discovery and sequence mapping of modifications to target proteins known to be represented in the analysis or identified by Sequest. P-Mod matches MS/MS spectra to peptide sequences in a search list. For spectra of modified peptides, P-Mod calculates mass differences between search peptide sequences and MS/MS precursors and localizes the mass shift to a sequence position in the peptide. Because modifications are detected as mass shifts, P-Mod does not require the user to guess at masses or sequence locations of modifications. P-Mod uses extreme value statistics to assign p value estimates to sequence-to-spectrum matches. The reported p values are scaled to account for the number of comparisons, so that error rates do not increase with the expanded search lists that result from incorporating potential peptide modifications. Combination of P-Mod searches from multiple LC-MS/MS analyses and multiple samples revealed previously unreported BSA modifications, including a novel decarboxymethylation or D-->G substitution at position 579 of the protein. P-Mod can serve a unique role in the identification of protein modifications both from exogenous and endogenous sources and may be useful for identifying modified protein forms as biomarkers for toxicity and disease processes.

Algorithms↗

PubMed2005

An integrated approach for the design and synthesis of oligonucleotide probes and their interfacing to a QCM-based RNA biosensor.

The quantitative determination of specific cellular messenger-RNA is extremely important both in basic and applied research, especially in diagnostic and pharmacological fields. In order to perform a direct and easy quantification of transcripts on cell extracts, the feasibility of an analytical device able to selectively detect a defined target RNA in a complex mixture while avoiding labelling, retrotranscription and amplification steps, has been explored. In particular, several aspects necessary to obtain good selectivity in target recognition, stability, reusability and sensitivity of a gene specific biosensor were considered. For the development of suitable probe-receptors, analysis of the nucleotide sequence of the target mRNA was carried out to localise the preferred binding regions. As criteria for optimisation, we selected accessibility and uniqueness. Oligonucleotide probes, designed to specifically bind these sequences, were synthesised by using particular monomers producing nuclease-resistant RNA strands with high affinity towards the target. Quartz crystal microbalance (QCM) technology was selected to realise a microgravimetric sensor able to bind the RNA under investigation through a complementary oligonucleotide probe. Covalent immobilisation of bioreceptor molecules to the transducer sensitive surface ensured a stable integration between the two. The binding ability of immobilised probes was tested evaluating their annealing behaviour with both complementary oligonucleotides and full-length target mRNA. The conditions necessary for the regeneration of biosensor were also assessed. Measurements of shift in QCM resonant frequency, performed by hybridisation experiments in liquido, demonstrate that a label-free RNA-biosensor with high specificity, reusability and the ability to give quantitative information, can be realised.

Biosensing Techniques↗

PubMed1998

The molecular biology of serotonin receptors: therapeutic implications for the interface of mood and psychosis.

This review summarizes the molecular biology of serotonin (5-HT; 5-hydroxytryptamine) receptors and indicates the potential relevance of this information for the treatment of mood and psychotic disorders. At least 15 separate subtypes of 5-HT receptors have been identified by molecular cloning techniques to be distinct genetic entities. Subtle differences in the primary amino acid sequences of these receptors can yield large differences in ligand selectivity. Additionally, it has recently been discovered that drugs such as atypical antipsychotic drugs and serotonin-selective reuptake inhibitors may interact with a large number of heretofore unknown 5-HT receptors. Thus clozapine, for instance, has high affinity for at least four separate 5-HT receptors, and it is unknown which of these receptors is essential for its unique therapeutic efficacy. One way to approach these questions is to test subtype-selective agents, although there are few of these currently available. Approaches to the design of subtype-selective ligands are described, including structure-based drug design and combinatorial approaches. Modes of regulation of 5-HT receptors are also summarized, and it is emphasized that antipsychotic drugs and antidepressants likely exert their effects via nontranscriptional and posttranslational means. Understanding the cellular mechanisms by which 5-HT receptors are regulated by psychopharmacologic agents is likely to yield novel insights into drug action.

Animals↗

PubMed1994

Common structural features of the luxF protein and the subunits of bacterial luciferase: evidence for a (beta alpha)8 fold in luciferase.

The amino acid sequence identity and potential structural similarity between the subunits of bacterial luciferase and the recently determined structure of the luxF molecule are examined. The unique beta/alpha barrel fold found in luxF appears to be conserved in part in the luciferase subunits. From secondary structural predictions of both luciferase subunits, and from structural comparisons between the protein product of the luxF gene, NFP, and glycolate oxidase, we propose that it is feasible for both luciferase subunits to adopt a (beta alpha)8 barrel fold with at least 2 excursions from the (beta alpha)8 topology. Amino acids conserved between NFP and the luciferase subunits cluster together in 3 distinct "pockets" of NFP, which are located at hydrophobic interfaces between the beta-strands and alpha-helices. Several tight turns joining the C-termini of beta-strands and the N-termini of alpha-helices are found as key components of these conserved regions. Helix start and end points are easily demarcated in the luciferase subunit protein sequences; the N-cap residues are the most strongly conserved structural features. A partial model of the luciferase beta subunit from Photobacterium leiognathi has been built based on our crystallographically determined structure of luxF at 1.6 A resolution.

Alcohol Oxidoreductases↗

PubMed1990

Effects of substrata on the polarization of bovine endometrial epithelial cells in vitro.

Epithelial-cell function requires cellular polarity in which apical membrane surfaces have unique characteristics and cellular organelles are stratified. Physiological investigations of endometrial epithelial cells would be enhanced greatly by the ability of a method to polarize cells in culture. This study investigates the effects of different substrata on polarization of cultured bovine endometrial epithelial cells. Fetal bovine endometrial epithelial-cell lines were developed from explant outgrowth. Epithelial monolayers were subcultured onto amniotic membranes, Millicell-HA membranes, or Millicell-CM membranes coated with rat-tail collagen, Matrigel, laminin, Vitrogen, or fibronectin. Cultures on these substrata were maintained at the air/liquid interface. Cells grown on either collagen-coated or uncoated Millicell membranes also were maintained submerged in medium. Excellent polarized morphology was attained in cultures grown at the air/liquid interface on amniotic membranes and rat-tail collagen-coated membranes. Lectin-binding patterns to apical membranes of polarized epithelial cell cultures paralleled patterns of binding to bovine endometrial surfaces in vivo. Cultures on rat-tail collagen were maintained for several weeks. These methods provide a valuable system for studying the endometrium in vitro.

Animals↗

PubMed1995

Rapid application development in small unit settings.

Small units with specialised functions often have difficulty in developing information systems because of their unique and dynamic requirements, the smallness of their applications relative to the large inter-organisational networks, and the sheer numbers of such units, especially in government. Personal experience with the difficulty of establishing a suitable information system has led to the development of a methodology for endusers to develop their own applications with only the guidance of information technology professionals (ITPs)--here entitled 'Small Unit Enduser Rapid Application Development' or SERAD. A data base management system (DBMS) is used to manage data about 'focal objects' with status flags and queries/views at interface points through UDA (UserAction, DataEntry, ApplicationAction) cycles. The system which permits data entry almost immediately, not only eventually creates a full application, but also trains endusers in the process. The methodology leads ITPs and ensuers through a sequence of exercises in systems analysis, tailored to minimise the time and effort input of ITPs despite the relative lack of IT training and concepts in endusers, with a view to rapidly creating dynamic, robust and sound information systems.

Computer Communication Networks↗

PubMed2025

Biochemical and Structural Analyses of the Tardigrade DNA-Damage Suppressor Protein, Dsup.

Tardigrades are extremophiles that withstand harsh environments through unique molecular strategies. One such strategy involves Damage Suppressor (Dsup), a protein shown to protect cells from radiation-induced DNA damage. Little is known about the biochemical and structural characteristics of Dsup that lead to DNA protection. To gain insight into the mechanism of DNA protection by Dsup, we examined its fundamental biochemical and structural properties using mass photometry, biolayer interferometry, small-angle X-ray scattering, and microfluidic modulation spectroscopy. We found that Dsup is largely intrinsically disordered and binds DNA with high affinity via a multi-valent interface. This interaction induced conformational changes in both Dsup and the DNA, suggesting a potential structural mechanism of its DNA protection ability. We propose that Dsup alters DNA structure, possibly by partially unwinding it, to reduce its susceptibility to damage. These findings offer new insights into how a disordered protein such as Dsup functions as radioprotectants in extreme environments.

Tardigrada↗

PubMed2005

Thinking outside the cell: proteases regulate hepatocyte division.

The liver has the unique ability to regenerate after loss of mass and function such as following surgical resection or toxic liver injury. Gene targeting has identified factors crucial to liver development and regeneration. Regeneration occurs through growth-factor- and cytokine-mediated proliferation of differentiated hepatocytes, and extracellular proteases are now recognized to process these molecules. Proteases release cytokines and growth factors that are anchored to the hepatic extracellular matrix or require processing for their bioactivity. Crucial 'start and stop' signals for liver regeneration are regulated by serine proteases and metalloproteases that provide an interface between proteolytic cascades and intracellular signaling during hepatocyte division.

Animals↗

PubMed1993

Cytoplasmic viscosity near the cell plasma membrane: measurement by evanescent field frequency-domain microfluorimetry.

The purpose of this study was to determine whether the unique physical milieu just beneath the cell plasma membrane influences the rheology of fluid-phase cytoplasm. Cytoplasmic viscosity was evaluated from the picosecond rotation of the small fluorophore 2',7'-bis-(2-carboxyethyl)-5-carboxyfluorescein (BCECF) by parallel-acquisition Fourier transform microfluorimetry (Fushimi and Verkman, 1991). Information about viscosity within < 200 nm of cell plasma membranes was obtained by selective excitation of fluorophores in an evanescent field created by total internal reflection (TIR) of impulse-modulated s-plane-polarized laser illumination (488 nm) at a glass-aqueous interface. Measurements of fluorescence lifetime and time-resolved anisotropy were carried out in solutions containing fluorescein or BCECF at known viscosities, and monolayers of BCECF-labeled Swiss 3T3 fibroblasts and Madin-Darby canine kidney (MDCK) cells. Specific concerns associated with time-resolved fluorescence measurements in the evanescent field were examined theoretically and/or experimentally, including variations in lifetime due to fluorophore proximity to the interface, and the use of the s and p polarized excitation. In fluorescein solutions excited with s-plane polarized light, there was a 5-10% decrease in fluorescein lifetime with TIR compared to trans (subcritical) illumination, but no change in rotational correlation time (approximately 98 ps/cP). Intracellular BCECF had a single lifetime of 3.7 +/- 0.1 ns near the cell plasma membrane. Apparent fluid-phase viscosity near the cell plasma membrane was 1.1 +/- 0.2 cP (fibroblast) and 1.0 +/- 0.2 cP (MDCK), not significantly different from the viscosity measured in bulk cytoplasm far from the plasma membrane. The results establish the methodology for time-resolved microfluorimetric measurement of polarization in the evanescent field and demonstrate that the cell plasma membrane has little effect on the fluid-phase viscosity of adjacent cytoplasm.

3T3 Cells↗

PubMed2001

Tetrameric dipeptidyl peptidase I directs substrate specificity by use of the residual pro-part domain.

The crystal structure of mature dipeptidyl peptidase I reveals insight into the unique tetrameric structure, substrate binding and activation of this atypical papain family peptidase. Each subunit is composed of three peptides. The heavy and light chains form the catalytic domain, which adopts the papain fold. The residual pro-part forms a beta-barrel with the carboxylate group of Asp1 pointing towards the substrate amino-terminus. The tetrameric structure appears to stabilize the association of the two domains and encloses a 12700 A3 spherical cavity. The tetramer contains six chloride ions, one buried in each S2 pocket and two at subunit interfaces.

Animals↗

PubMed2003

Structural modeling of the Lhca4 Subunit of LHCI-730 peripheral antenna in photosystem I based on similarity with LHCII.

Peripheral chlorophyll a/b binding antenna of photosystem I (LHCI) from green algae and higher plants binds specific low energy absorbing chlorophylls (red pigments) that give rise to a unique red-shifted emission. A three-dimensional structural model of the Lhca4 polypeptide from the LHCI from higher plants was constructed on the basis of comparative sequence analysis, secondary structure prediction, and homology modeling using LHCII as a template. The obtained model of Lhca4 helps to visualize protein ligands to nine chlorophylls (Chls) and three potential His residues to extra Chls. Central domain of the Lhca4 comprising the first (A) and the third (C) transmembrane (TM) helices that binds 6 Chl molecules and two carotenoids is conserved structurally, whereas the interface between the first and the second TM helices and the outer surface of the second TM helix differ significantly among the LHCI and LHCII polypeptides. The model of Lhca4 predicts a histidine residue in the second TM helix, a potential binding site for extra Chl in close proximity to Chls a5 and b5 (labeling by Kühlbrandt). The interpigment interactions in the formed pigment cluster are suggested to cause a red spectral shift in absorption and emission. Modeling of the LHCI-730 heterodimer based on the model structures of Lhca1 and Lhca4 allowed us to suggest potential sites of pigment-pigment interactions that might be formed upon heterodimerization or docking of the LHCI dimers to the surface of PSI.

Amino Acid Sequence↗

PubMed2005

Structural probing of a pathogenic tRNA dimer.

The A3243G mutation within the human mitochondrial (hs mt) tRNALeuUUR gene is associated with maternally inherited deafness and diabetes (MIDD) and other mitochondrial encephalopathies. One of the most pronounced structural effects of this mutation is the disruption of the native structure through stabilization of a high-affinity dimeric complex. We conducted a series of studies that address the structural properties of this tRNA dimer, and we assessed its formation under physiological conditions. Enzymatic probing was used to directly define the dimeric interface for the complex, and a discrete region of the D-stem and loop of hs mt tRNALeuUUR was identified. The dependence of dimerization on magnesium ions and temperature was also tested. The formation of the tRNA dimer is influenced by temperature, with dimerization becoming more efficient at physiological temperature. Complexation of the mutant tRNA is also affected by the amount of magnesium present, and occurs at concentrations present intracellularly. Terbium probing experiments revealed a specific metal ion-binding site localized at the site of the A3243G mutation that is unique to the dimer structure. This metal ion-binding site presents a striking parallel to dimeric complexes of viral RNAs, which use the same hexanucleotide sequence for complexation and feature a similarly positioned metal ion-binding site within the dimeric structure. Taken together, these results indicate that the unique dimeric complex formed by the hs mt tRNALeuUUR A3243G mutant exhibits interesting similarities to biological RNA dimers, and may play a role in the loss of function caused by this mutation in vivo.

Base Sequence↗

PubMed2003

Genome-wide prediction, display and refinement of binding sites with information theory-based models.

BACKGROUND: We present Delila-genome, a software system for identification, visualization and analysis of protein binding sites in complete genome sequences. Binding sites are predicted by scanning genomic sequences with information theory-based (or user-defined) weight matrices. Matrices are refined by adding experimentally-defined binding sites to published binding sites. Delila-Genome was used to examine the accuracy of individual information contents of binding sites detected with refined matrices as a measure of the strengths of the corresponding protein-nucleic acid interactions. The software can then be used to predict novel sites by rescanning the genome with the refined matrices. RESULTS: Parameters for genome scans are entered using a Java-based GUI interface and backend scripts in Perl. Multi-processor CPU load-sharing minimized the average response time for scans of different chromosomes. Scans of human genome assemblies required 4-6 hours for transcription factor binding sites and 10-19 hours for splice sites, respectively, on 24- and 3-node Mosix and Beowulf clusters. Individual binding sites are displayed either as high-resolution sequence walkers or in low-resolution custom tracks in the UCSC genome browser. For large datasets, we applied a data reduction strategy that limited displays of binding sites exceeding a threshold information content to specific chromosomal regions within or adjacent to genes. An HTML document is produced listing binding sites ranked by binding site strength or chromosomal location hyperlinked to the UCSC custom track, other annotation databases and binding site sequences. Post-genome scan tools parse binding site annotations of selected chromosome intervals and compare the results of genome scans using different weight matrices. Comparisons of multiple genome scans can display binding sites that are unique to each scan and identify sites with significantly altered binding strengths. CONCLUSIONS: Delila-Genome was used to scan the human genome sequence with information weight matrices of transcription factor binding sites, including PXR/RXRalpha, AHR and NF-kappaB p50/p65, and matrices for RNA binding sites including splice donor, acceptor, and SC35 recognition sites. Comparisons of genome scans with the original and refined PXR/RXRalpha information weight matrices indicate that the refined model more accurately predicts the strengths of known binding sites and is more sensitive for detection of novel binding sites.

Binding Sites↗

PubMed1985

A complete system for barcode generation on the HP-41CX hand-held computer using a standard thermal printer.

One of the unique features of the HP-41C hand-held computer is its ability to read program code and alphanumeric data in the form of printed barcode. This greatly facilitates the dissemination of medical software and data used by this system. Initially, the ability to generate HP-41 barcode was restricted to a few centers in possession of specialized microcomputer equipment, and more recently to users able to either obtain a plotter read-only memory (ROM) module and HP7400 series plotter, or a dot-matrix impact printer, for which a special interface was then required. This paper presents a set of twenty programs which allows all nine types of HP-41C barcode to be generated using an HP-41CX, the commonly encountered 82162A thermal printer, an HP-IL ROM, and the plotter ROM. The package includes a number of input/output routines which are of general utility. Complete barcode listings, generated by the system itself, allow complete installation of the system without use of the keyboard. 'Synthetic programming' code permits the use of helpful non-standard characters in the display and allows that part of the system which is concerned with generating program barcode to run in extended memory, freeing main memory (RAM) to hold a large program. This system should make the generation of barcode much more widely available.

Abstracting and Indexing↗

PubMed2003

Preliminary validation of computational procedures for a new atmospheric ionizing radiation (AIR) model.

A new computational procedure to determine particle fluxes in the Earth's atmosphere is presented. The primary cosmic ray spectrum has been modeled through an analysis of simultaneous proton and helium measurements made on high altitude balloon flights and spacecraft. An improved global fit to the data was achieved through applying a unique technique utilizing the Fokker-Plank equation with a non-linear rigidity-dependent diffusion coefficient. The propagation of primary particles through the Earth's atmosphere is calculated with a three-dimensional Monte Carlo transport program called FLUKA. Primary protons and helium nuclei (alphas) are generated within the rigidity range of 0.5 GV-20 TV uniform in cos2 theta. For a given location, primaries above the effective cutoff rigidity are transported through the atmosphere. Alpha particles are initially transported with a separate package called HEAVY to simulate fragmentation. This package interfaces with FLUKA to provide interaction starting points for each nucleon originating from a helium nucleus. Results from this calculation are presented and compared to measurements.

Alpha Particles↗

PubMed2003

Doping control analysis of bovine hemoglobin-based oxygen therapeutics in human plasma by LC-electrospray ionization-MS/MS.

Since January 2000, hemoglobin-based oxygen carriers, such as Hemopure, belong to the list of prohibited substances of the International Olympic Committee. Hemopure is based on bovine hemoglobin, which is intra- and intermolecularly cross-linked by glutaraldehyde units causing an average molecular weight of approximately 250,000. Bovine and human hemoglobins differ by 15% in amino acid sequence; hence, tryptic digestion of these proteins generates species-common and -unique peptides. Those specific fragments originate from the alpha- and beta-subunits of hemoglobin, such as bovine Hb peptides alpha(69-90) (2367.2 Da) or beta(40-58) (2089.9 Da). By means of LC-MS/MS, peptides of human and bovine hemoglobin can be separated and identified, enabling the determination of compounds based on Hb of bovine origin and thus the administration of oxygen carriers such as Hemopure. Blank plasma samples were spiked with Hemopure or human or bovine hemoglobin, filtered, enzymatically digested, and analyzed on an Agilent 1100 Series HPLC interfaced to an Applied Biosystems API 2000 triple quadrupole mass spectrometer. In plasma aliquots of 50 microL containing 50 microg of Hemopure (1 mg/mL), peptides of bovine hemoglobin were confirmed, and blank plasma samples as well as 68 specimens of high-performance athletes were tested with the developed procedure.

Amino Acid Sequence↗

PubMed2001

The C-terminus of glutathione S-transferase A1-1 is required for entropically-driven ligand binding.

Binding of a hydrophobic glutathione product conjugate to rGST A1-1 proceeds via a two-step mechanism, including rapid ligand docking, followed by a slow isomerization to the final [GST.ligand] complex, which involves the localization of the flexible C-terminal helix. These kinetically resolved steps have been observed previously by stopped-flow fluorescence with the wild-type rGST A1-1, which contains a native Trp-21 approximately 20 A from the ligand binding site at the intrasubunit domain-domain interface. To confirm this binding mechanism, as well as elucidate the effects of truncation of the C-terminus, we have further characterized the binding and dissociation of the glutathione-ethacrynic acid product conjugate (GS-EA) to wild-type, F222W:W21F, and Delta209-222 rGST A1-1 and wild-type hGST A1-1. Although modest kinetic differences were observed between the hGST A1-1 and rGST A1-1, stopped-flow binding studies with GS-EA verified that the two-step mechanism of ligand binding is not unique to the GST A1-1 isoform from rat. An F222W:W21F rGST A1-1 double mutant provides a direct fluorescence probe of changes in the environment of the C-terminal residue. The observation of two relaxation times during ligand binding and dissociation to F222W:W21F suggests that the C-terminus has an intermediate conformation following ligand docking, which is distinct from its conformation in the apoenzyme or localized helical state. For the wild-type, Delta209-222, and F222W:W21F proteins, variable-temperature stopped-flow experiments were performed and activation parameters calculated for the individual steps of the binding reaction. Activation parameters for the binding reaction coordinate illustrate that the C-terminus provides a significant entropic contribution to ligand binding, which is completely realized within the initial docking step of the binding mechanism. In contrast, the slow isomerization step is enthalpically driven. The partitioning of entropic and enthalpic components of binding energy was confirmed by isothermal titration calorimetry with wild-type and Delta209-222 rGST A1-1.

Animals↗

PubMed2005

Structural and mechanistic analysis of two prolyl endopeptidases: role of interdomain dynamics in catalysis and specificity.

Prolyl endopeptidases (PEPs) are a unique class of serine proteases with considerable therapeutic potential for the treatment of celiac sprue. The crystal structures of two didomain PEPs have been solved in alternative configurations, thereby providing insights into the mode of action of these enzymes. The structure of the Sphingomonas capsulata PEP, solved and refined to 1.8-A resolution, revealed an open configuration of the active site. In contrast, the inhibitor-bound PEP from Myxococcus xanthus was crystallized (1.5-A resolution) in a closed form. Comparative analysis of the two structures highlights a critical role for the domain interface in regulating interdomain dynamics and substrate specificity. Structure-based mutagenesis of the M. xanthus PEP confirms an important role for several interfacial residues. A salt bridge between Arg-572 and Asp-196/Glu-197 appears to act as a latch for opening or closing the didomain enzyme, and Arg-572 and Ile-575 may also help secure the incoming peptide substrate to the open form of the enzyme. Arg-618 and Asp-145 are responsible for anchoring the invariant proline residue in the active site of this postproline-cleaving enzyme. A model is proposed for the docking of a representative substrate PQPQLPYPQPQLP in the active site, where the N-terminal substrate residues interact extensively with the catalytic domain, and the C-terminal residues stretch into the propeller domain. Given the promise of the M. xanthus PEP as an oral therapeutic enzyme for treating celiac sprue, our results provide a strong foundation for further optimization of the PEP's clinically useful features.

Binding Sites↗