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PubMed2004

The aging of a clinical information system.

The senescence of a clinical information system is more likely to have administrative than technical bases. Supporting this claim is a case study of one aging oncology information system. The case study is qualitative, as behooves the subject matter. Content analysis of several documents suggests that the change in job description of the data coordinator led to a workflow breakdown. Next, twenty-two individuals were interviewed. Notes from the interviews were coded, and the resulting patterns led to partial support for the workflow breakdown conjecture, refutation of the hypothesis that users disliked the character-based, human-computer interface, support of the conjecture that political rather than technical factors drive the usage patterns of the system, and evidence that 'political' activity will determine the future of the information system. A stakeholder matrix is proposed that addresses administrative concerns. Also, the issue of the uniqueness of any oncology clinical information system is linked to the plans for this legacy system.

Databases, Factual↗

PubMed2002

A smart adhesive joint: entropic control of adhesion at a polymer/metal interface.

The study of adhesion has a long and rich history, with theory, experiments, and applications bridging numerous disciplines, including physics, chemistry, engineering, and medicine. This diverse interest has led to the development of a large number of methods for both enhancing and inhibiting adhesion at specific interfaces of interest. We report herein "smart" adhesion at a polymer/metal (oxide) interface that responds reversibly to changes in temperature by increasing or decreasing in magnitude. The temperature dependence in this system arises from the rubber elasticity of the polymer, 1,4-polybutadiene, and mirrors the interfacial behavior of the same polymer against water. Such systems offer unique opportunities for designing responsive materials whose properties can be actively controlled.

Journal Article↗

PubMed1996

Identification of residues of the epidermal growth factor receptor proximal to residue 45 of bound epidermal growth factor.

A triple mutant of murine epidermal growth factor (mEGF), N1Q/H22Y/R45K-mEGF, was constructed by site-directed mutagenesis, expressed, purified, and characterized for use in an affinity cross-linking study to identify aminoacyl residues of the EGF receptor adjacent to a residue in the carboxyl-terminal domain of bound EGF thought to be important in distinguishing between EGF and transforming growth factor-alpha in their recognition by the receptor. Cyclization of Gln1 to form pyroglutamate (pE) limited the site of cross-linking in the mutant to Lys45, permitting identification of receptor residues that are proximal to this residue of bound EGF. The resulting N1pE/H22Y/R45K-mEGF was shown to be comparable to wild-type mEGF in receptor binding and stimulation of receptor autophosphorylation. 125I-Labeled N1pE/H22Y/R45K-mEGF was reacted with the heterobifunctional cross-linking reagent sulfo-N-succinimidyl-4-(fluorosulfonyl)benzoate, and the resulting modified EGF was incubated with A431 membrane vesicles bearing EGF receptors. Incubation resulted in specific cross-linking of the labeled N1pE/H22Y/R45K-mEGF to EGF receptors. The resulting cross-linked complex was then partially purified, denatured, reduced, and carboxyamidomethylated. Digestion with endoprotease LysC resulted in a unique radiolabeled peptide that could be immunoprecipitated using antibodies to mEGF. This immunoprecipitated fragment was purified by gel electrophoresis and subjected to microsequencing. The resulting sequence was matched to that of a LysC fragment of the receptor, which begins with Thr464 and is near the interface of receptor subdomains III and IV. Loss of signal at cycle 2 suggests that the point of attachment of cross-linked N1pE/H22Y/R45K is Lys465 of the receptor.

Amino Acid Sequence↗

PubMed2001

Genetic control of peripheral TCRAV usage by representation in the preselection repertoire and MHC allele-specific overselection.

TCRAV segments contribute significantly to MHC restriction as illustrated by their general preference for either the CD4 or CD8 T cell subset and additional, MHC allele-specific overselection during T cell differentiation. The 10-fold over-representation of the TCRAV8S2 (VA8S2) segment in CD8 over CD4 T cells by the RT1(f) haplotype of LEW.1F rats provides the most striking example of MHC allele-specific overselection of a VA segment reported so far. Also in alloreactivity, VA8S2(+) CD8 cells from RT1(f-) rats are preferentially expanded by RT1(f+) stimulators. We have identified the class I molecule, A(f), mediating VA8S2 overselection and report that it differs only in four amino acids at the MHC-TCR interface from the class I molecule A(a), which is neutral with regard to selection of VA8S2. We also provide an extensive survey of the TCRAV8 family and show that among 14 functional VA8 segments in LEW rats, the dramatic A(f)-dependent overselection is unique for VA8S2. Surprisingly, VA8S2 expression in CD8 T cells of RT1(f+) rats derived from a Sprague-Dawley stock was only 3% as compared to the 12% observed in LEW.1F. The VA8S2 segment of Sprague-Dawley (VA8S2(SD)) differs from VA8S2 of the LEW background (VA8S2(l)) in only two amino acids, one of which is located in CDR2 and could thus participate in allele-specific recognition of A(f). However, analysis of the pre- and postselection thymic repertoires of Sprague-Dawley and LEW.1F rats and of the repertoire of CD8 cells from both strains expanded in the alloreactive response to RT1(f) revealed that the difference in VA8S2 representation between the two backgrounds is explained by differential availability in the preselection repertoires and not by a difference in overselection. Sequence comparisons of A(f) and A(a) and of both VA8S2 segments suggest a predominant role of CDR1 in hyper-reactivity to A(f). Thus, the VA composition of the mature TCR repertoire is influenced by TCRA: locus polymorphisms at two levels: the regulation of VA usage in the preselection repertoire and the composition of structural elements which contribute to specific VA-MHC interactions during thymic selection.

Alleles↗

PubMed2000

Peptide nucleic acid (PNA): its medical and biotechnical applications and promise for the future.

Synthetic molecules that can bind with high sequence specificity to a chosen target in a gene sequence are of major interest in medicinal and biotechnological contexts. They show promise for the development of gene therapeutic agents, diagnostic devices for genetic analysis, and as molecular tools for nucleic acid manipulations. Peptide nucleic acid (PNA) is a nucleic acid analog in which the sugar phosphate backbone of natural nucleic acid has been replaced by a synthetic peptide backbone usually formed from N-(2-amino-ethyl)-glycine units, resulting in an achiral and uncharged mimic. It is chemically stable and resistant to hydrolytic (enzymatic) cleavage and thus not expected to be degraded inside a living cell. PNA is capable of sequence-specific recognition of DNA and RNA obeying the Watson-Crick hydrogen bonding scheme, and the hybrid complexes exhibit extraordinary thermal stability and unique ionic strength effects. It may also recognize duplex homopurine sequences of DNA to which it binds by strand invasion, forming a stable PNA-DNA-PNA triplex with a looped-out DNA strand. Since its discovery, PNA has attracted major attention at the interface of chemistry and biology because of its interesting chemical, physical, and biological properties and its potential to act as an active component for diagnostic as well as pharmaceutical applications. In vitro studies indicate that PNA could inhibit both transcription and translation of genes to which it has been targeted, which holds promise for its use for antigene and antisense therapy. However, as with other high molecular mass drugs, the delivery of PNA, involving passage through the cell membrane, appears to be a general problem.

Animals↗

PubMed2006

Expressed sequence-tag analysis in Casuarina glauca actinorhizal nodule and root.

The present study aimed to identify and assess the frequency and tissue specificity of plant genes in the actinorhizal Casuarina glauca-Frankia symbiosis through expressed sequence tag (EST) analysis. Using a custom analysis pipeline for raw sequences of C. glauca uninfected roots and nodules, we obtained an EST databank web interface. Gene expression was studied in nodules vs roots using comparative quantitative real-time reverse transcription-polymerase chain reaction (qRT-PCR). From roots and nodules, 2028 ESTs were created and clustered in 242 contigs and 1429 singletons, giving a total of 1616 unique genes. Half the nodule transcripts showed no similarity to previously identified genes. Genes of primary metabolism, protein synthesis, cell division and defence were highly represented in the nodule library. Differential expression was observed between roots and nodules for several genes linked to primary metabolism and flavonoid biosynthesis. This comparative EST-based study provides the first picture of the set of genes expressed during actinorhizal symbiosis. We consider our database to be a flexible tool that can be used for the management of EST data from other actinorhizal symbioses.

Cluster Analysis↗

PubMed1994

Aerosol characteristics of 99mTc-pentetic acid (DTPA) and synthetic surfactant (Exosurf).

This study evaluated the feasibility of using 99mTc-pentetic acid (DTPA) as a radioactive tracer for aerosolized synthetic surfactant (DPPC, cetyl alcohol, tyloxapol). The 99mTc-DTPA was admixed with surfactant and aerosolized using a nebulizer system interfaced to a ventilator with a cascade impactor attached to the endotracheal tube. Particle size distribution for DPPC, cetyl alcohol, and 99mTc-DTPA were almost identical during the 0- to 15-, 15- to 30-, and 0- to 30-min collection periods. Tyloxapol exhibited a unique distribution pattern with increased deposition in large (> 10 microns) and small (0.65 to 1.1 microns) particles. The mass median aerodynamic diameter for all aerosolized components was in the respirable range of 2.1 to 2.5 microns. A mixture of 99mTc-DTPA with synthetic surfactant appears to be a reasonable method to evaluate surfactant deposition.

Aerosols↗

PubMed2002

A concordance-based study of metaphoric expressions used by general practitioners and patients in consultation.

BACKGROUND: All languages use metaphoric expressions; some deliberately chosen, some (for example, 'digesting information') not usually perceived as metaphoric. Increasingly, it is suggested metaphoric expressions constrain the way we conceptualise the world, as well as being a means of achieving stylistic effect. AIM: To study metaphoric expressions used by doctors and patients in general practice. DESIGN OF STUDY: Concordance-based language analysis of spoken data. METHOD: A database containing transcriptions of 373 consultations with 40 doctors in a UK general practice setting was scrutinised for metaphoric expressions, using 'concordancing' software. Concordancing enables identification of strings of text with similar lexical properties. Comparators (for example, 'like'), selected verb-types (for example, of feeling), and the verb 'to be' were used as starting points for systematically exploring the data. Quantitative and qualitative thematic methods were used in analysis. RESULTS: Doctors and patients use different metaphors. Doctors use mechanical metaphors to explain disease and speak of themselves as problem-solvers' and 'controllers of disease'. Patients employ a range of vivid metaphors, but fewer metaphors of machines and problem/solution. Patients use metaphors to describe symptoms and are more likely to use metaphoric language at the interface of physical and psychological symptoms ('tension, 'stress'). CONCLUSION: The different patterns of metaphoric expression suggest that doctors make limited attempts to enter the patients' conceptual world. This may not be a bad thing. One function of the consultation may be to reinterpret vivid and unique descriptions as accounts of the familiar and systemically comprehensible. Doctors may use different conceptual metaphors as a reassuring signal of expertise.

Communication↗

PubMedDate not supplied

Quartz crystal microbalance: a useful tool for studying thin polymer films and complex biomolecular systems at the solution-surface interface.

The quartz crystal microbalance (QCM) is a simple, cost effective, high-resolution mass sensing technique, based upon the piezoelectric effect. As a methodology, the QCM evolved a solution measurement capability in largely analytical chemistry and electrochemistry applications due to its sensitive solution-surface interface measurement capability. The technique possesses a wide detection range. At the low mass end, it can detect monolayer surface coverage by small molecules or polymer films. At the upper end, it is capable of detecting much larger masses bound to the surface. These can be complex arrays of biopolymers and biomacromolecules, even whole cells. In addition, the QCM can provide information about the energy dissipating properties of the bound surface mass. Another important and unique feature of the technique is the ability to measure mass and energy dissipation properties of films while simultaneously carrying out electrochemistry on solution species or upon film systems bound to the upper electrode on the oscillating quartz crystal surface. These measurements can describe the course of electropolymerization of a film or can reveal ion or solute transport within a film during changes in the film environment or state, including the oxidation state for an electroactive film driven by the underlying surface potential. The past decade has witnessed an explosive growth in the application of the QCM technique to the study of a wide range of molecular systems at the solution-surface interface, in particular, biopolymer and biochemical systems. In this report, we start with a brief historical and technical overview. Then we discuss the application of the QCM technique to measurements involving micellar systems, self-assembling monolayers and their phase transition behavior, molecularly imprinted polymers, chemical sensors, films formed using the layer-by-layer assembly technique, and biopolymer films and point out the utility of the electrochemical capabilities of the technique to characterizing film properties, especially electroactive polymer films. We also describe the wide range of surface chemistries and attachment strategies used by investigators to bring about surface attachment and multi-layer interactions of these thin film systems. Next we review the wide range of recent applications of the technique to: studies of complex biochemical and biomimetic systems, the creation of protein and nucleic acid biosensors, studies of attached living cells and whole cell biosensor applications. Finally, we discuss future technical directions and applications of the QCM technique to areas such as drug discovery.

Biopolymers↗

PubMed1990

Regulation of proteolysis at the neutrophil-substrate interface by secretory leukoprotease inhibitor.

Human neutrophils can initiate the rapid degradation of extracellular matrix macromolecules by localizing the destructive process to sites of cell-substrate contact. Although plasma and its filtrates contain multiple proteinase inhibitors, these inhibitors did not prevent neutrophils from attacking either underlying fibronectin or elastin. However, subjacent substrates could be protected from neutrophils by recombinant secretory leukoprotease inhibitor, a structurally unique serine proteinase inhibitor whose natural counterpart is normally confined to human mucous secretions. The identification of this extravascular proteinase inhibitor as a potent regulator of subjacent proteolysis could lead to the development of a new class of anti-inflammatory therapeutics.

Cell Adhesion↗

PubMed1995

Crystal structure of a pyrimidine dimer-specific excision repair enzyme from bacteriophage T4: refinement at 1.45 A and X-ray analysis of the three active site mutants.

Crystallographic study of bacteriophage T4 endonuclease V, which is involved in the initial step of the pyrimidine dimer-specific excision repair pathway, has been carried out with respect to the wild-type and three different mutant enzymes. This enzyme catalyzes the cleavage of the N-glycosyl bond at the 5'-side of the pyrimidine dimer, and subsequently incises the phosphodiester bond at the apyrimidinic site through a beta-elimination reaction. The structure of the wild-type enzyme refined at 1.45 A resolution reveals the detailed molecular architecture. The enzyme is composed of a single compact domain classified as an all-alpha structure. The molecule is stabilized mainly by three hydrophobic cores, two of which include many aromatic side-chain interactions. The structure has a unique folding motif, where the amino-terminal segment penetrates between two major alpha-helices and prevents their direct contact, and it is incompatible with the close-packing category of helices for protein folding. The concave surface, covered with many positive charges, implies an interface for DNA binding. The glycosylase catalytic center, which comprises Glu23 and the surrounding basic residues Arg3, Arg22 and Arg26, lie in this basic surface. The crystal structures of the three active-site mutants, in which Glu23 was replaced by Gln(E23Q) and Asp (E23D), respectively, and Arg3 by Gln (R3Q), have been determined at atomic resolution. The backbone structures of the E23Q and R3Q mutants were almost identical with that of the wild-type, while the E23D mutation induces a small, but significant, change in the backbone structure, such as an increase of the central kink of the H1 helix at Pro25. In the catalytic center of the glycosylase, however, these three mutations do not generate notable movements of protein atoms, except for significant shifts of some bound water molecules. Thus, the structural differences between the wild-type and each mutant are confined to the remarkably small region around their replaced chemical groups. Combined with the biochemical studies and the difference circular dichroism measurements, these results allow us to conclude that the negatively charged carboxyl group of Glu23 is essential for the cleavage of the N-glycosyl bond, and that the positively charged guanidino group of Arg3 is crucial to bind the substrate, a DNA duplex containing a pyrimidine dimer. The amino terminal alpha-amino group is located at a position approximately 4.4 A away from the carboxyl group of Glu23. These structural features are generally consistent with the reaction scheme proposed by Dodson and co-workers.

Amino Acid Sequence↗

PubMed2000

The extracellular domain of BPAG2 has a loop structure in the carboxy terminal flexible tail in vivo.

The 180 kDa bullous pemphigoid antigen is a hemidesmosome-associated transmembranous protein with a molecule length estimated to be 190-230 nm, which is much longer than the transverse length of the lamina lucida and lamina densa. The purpose of this study was to clarify the precise in vivo structure of the 180 kDa bullous pemphigoid antigen in normal human skin. We used three monoclonal antibodies directed to (i) the intracellular globular head of the 180 kDa bullous pemphigoid antigen, (ii) the mid-portion of the flexible tail of the antigen, corresponding approximately to amino acids 1000-1320, and (iii) the carboxyl terminal end, corresponding approximately to amino acids 1320-1500 of the antigen. Using low temperature postembedding immunoelectron microscopy, we quantitated the distribution of immunogold labeling of these monoclonal antibodies in normal human skin. The results showed that the monoclonal antibodies (i) bound to the intracellular portion of the hemidesmosome at a mean distance of 20 nm from the plasma membrane, (ii) bound to the lamina densa beneath the hemidesmosome at a mean distance of 65 nm from the plasma membrane, and (iii) bound to the lamina densa-lamina lucida interface at a mean distance of 39 nm from the plasma membrane. Considering the reported size of the 180 kDa bullous pemphigoid antigen, our results indicate that the extracellular domain of the antigen has at least one loop structure in the lamina densa in vivo. This unique structure of the antigen is thought to contribute to dermo- epidermal adhesion by intertwining with other basement membrane components.

Amino Acid Sequence↗

PubMed2006

Clusters, surfaces, and catalysis.

The surface science of heterogeneous metal catalysis uses model systems ranging from single crystals to monodispersed nanoparticles in the 1-10 nm range. Molecular studies reveal that bond activation (C-H, H-H, C-C, CO) occurs at 300 K or below as the active metal sites simultaneously restructure. The strongly adsorbed molecules must be mobile to free up these sites for continued turnover of reaction. Oxide-metal interfaces are also active for catalytic turnover. Examples using C-H and CO activation are described to demonstrate these properties. Future directions include synthesis, characterization, and reaction studies with 2D and 3D monodispersed metal nanoclusters to obtain 100% selectivity in multipath reactions. Investigations of the unique structural, dynamic, and electronic properties of nanoparticles are likely to have major impact in surface technologies. The fields of heterogeneous, enzyme, and homogeneous catalysis are likely to merge for the benefit of all three.

Catalysis↗

PubMed1998

Crystallographic comparison of the estrogen and progesterone receptor's ligand binding domains.

The 2.8-A crystal structure of the complex formed by estradiol and the human estrogen receptor-alpha ligand binding domain (hERalphaLBD) is described and compared with the recently reported structure of the progesterone complex of the human progesterone receptor ligand binding domain, as well as with similar structures of steroid/nuclear receptor LBDs solved elsewhere. The hormone-bound hERalphaLBD forms a distinctly different and probably more physiologically important dimer interface than its progesterone counterpart. A comparison of the specificity determinants of hormone binding reveals a common structural theme of mutually supported van der Waals and hydrogen-bonded interactions involving highly conserved residues. The previously suggested mechanism by which the estrogen receptor distinguishes estradiol's unique 3-hydroxy group from the 3-keto function of most other steroids is now described in atomic detail. Mapping of mutagenesis results points to a coactivator-binding surface that includes the region around the "signature sequence" as well as helix 12, where the ligand-dependent conformation of the activation function 2 core is similar in all previously solved steroid/nuclear receptor LBDs. A peculiar crystal packing event displaces helix 12 in the hERalphaLBD reported here, suggesting a higher degree of dynamic variability than expected for this critical substructure.

Amino Acid Sequence↗

PubMed1999

Integrating geriatrics into clinical training, research training, board certification, and continuing education in infectious diseases: meeting review and commentary.

Although adults aged 65 years and older constitute the most rapidly growing segment of the U.S. population, geriatric issues have not typically been a focus of training in infectious diseases (ID). Underrecognition of the unique aspects of geriatric care, apathy toward this population, and the feeling that "we're all geriatricians" (and thus know geriatric medicine) all contribute to this problem. This article summarizes the recent meeting focused on integrating geriatric principles within ID training at all levels. The ID/geriatric interface as an attractive area for basic and clinical research is emphasized.

Adult↗

PubMed1998

Composite tumor consisting of dermatofibrosarcoma protuberans and myxofibrosarcoma.

A unique case of a composite tumor consisting of dermatofibrosarcoma protuberans and low-grade myxofibrosarcoma is described. The myxofibrosarcoma component formed 70% of the lesion volume. Both components protruded above the skin, and equally both infiltrated the subcutaneous tissue in the base of the lesion. The interface between them was mostly sharp but some small areas of gradual transition were also seen. Beside fibrosarcoma arising in the dermatofibrosarcoma protuberans, myxofibrosarcoma seems thus to be another possible manifestation of the change into a more malignant tumor rarely seen in the dermatofibrosarcoma protuberans.

Aged↗

PubMed2005

Potential nectin-1 binding site on herpes simplex virus glycoprotein d.

Four glycoproteins (gD, gB, gH, and gL) are essential for herpes simplex virus (HSV) entry into cells. An early step of fusion requires gD to bind one of several receptors, such as nectin-1 or herpesvirus entry mediator (HVEM). We hypothesize that a conformational change in gD occurs upon receptor binding that triggers the other glycoproteins to mediate fusion. Comparison of the crystal structures of gD alone and gD bound to HVEM reveals that upon HVEM binding, the gD N terminus transitions from a flexible stretch of residues to a hairpin loop. To address the contribution of this transition to the ability of gD to trigger fusion, we attempted to "lock" the gD N terminus into a looped conformation by engineering a disulfide bond at its N and C termini. The resulting mutant (gD-A3C/Y38C) failed to trigger fusion in the absence of receptor, suggesting that formation of the loop is not the sole fusion trigger. Unexpectedly, although gD-A3C/Y38C bound HVEM, it failed to bind nectin-1. This was due to the key role played by Y38 in interacting with nectin-1. Since tyrosines are often "hot spot" residues at the center of protein-protein interfaces, we mutated residues that surround Y38 on the same face of gD and tested their binding and functional properties. Our results suggest that this region of gD is important for nectin-1 interaction and is distinct from but partially overlaps the site of HVEM binding. Unique gD mutants with altered receptor usage generated in this study may help dissect the roles played by various HSV receptors during infection.

Animals↗

PubMed2026

Microbiome Datahub: an open-access platform integrating environmental metadata, taxonomy, and functional annotation for comprehensive metagenome-assembled genome datasets.

BACKGROUND: Metagenome-assembled genomes (MAGs) provide crucial insights into the genomic diversity of uncultured microbes. However, MAG datasets deposited in public repositories such as INSDC are often difficult to reuse due to heterogeneous quality, inconsistent taxonomic and functional annotations, and insufficiently curated environmental metadata. While secondary MAG databases such as MGnify, IMG/M, and SPIRE provide standardized resources, they reconstruct MAGs de novo from public metagenomic reads and therefore do not represent the original MAGs reported in publications. RESULTS: To address this gap, we developed Microbiome Datahub, an open-access platform that systematically aggregates and re-annotates original MAGs from INSDC. We collected 214,427 MAGs, predicted genes by DFAST, performed quality assessment with CheckM, standardized taxonomic assignments with GTDB-Tk, inferred 27 phenotypic traits using Bac2Feature, assigned proteins to MBGD ortholog clusters and KEGG Orthology IDs using PZLAST, and annotated environmental metadata with the Metagenome and Microbes Environmental Ontology. Across these MAGs, the average completeness was 80.5% and contamination 1.8%; notably, the most frequent values were >95% completeness and <1% contamination, indicating that the majority of MAGs are of high quality. Comparative analyses showed that Microbiome Datahub provides phylogenetically and environmentally diverse MAGs: while the majority originated from vertebrate gut environments, a substantial number were also recovered from other habitats such as groundwater, including nearly 10,000 MAGs from the Patescibacteria. Inference of 27 phenotypic traits, including optimum growth temperature, further revealed ecological differentiation across phyla. Protein clustering revealed 56 million identity 40% clusters, with the majority unique compared with MGnify and GlobDB, and ~19% of proteins unassigned to MBGD ortholog clusters, underscoring their novelty. CONCLUSIONS: Microbiome Datahub integrates MAG genome sequences, gene and protein predictions, quality metrics, environmental and taxonomic annotations, ortholog cluster assignments, and phenotype predictions, all accessible via a web interface, API, and bulk downloads. By combining original MAGs with curated metadata and functional annotations, Microbiome Datahub constitutes a comprehensive and reusable resource that will accelerate microbiome and microbial genomics research. Video Abstract.

Metagenome↗