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At least 757 records · Page 42Linked to original sources
PubMed2006

Direct observation of DNA distortion by the RSC complex.

The Snf2 family represents a functionally diverse class of ATPase sharing the ability to modify DNA structure. Here, we use a magnetic trap and an atomic force microscope to monitor the activity of a member of this class: the RSC complex. This enzyme caused transient shortenings in DNA length involving translocation of typically 400 bp within 2 s, resulting in the formation of a loop whose size depended on both the force applied to the DNA and the ATP concentration. The majority of loops then decrease in size within a time similar to that with which they are formed, suggesting that the motor has the ability to reverse its direction. Loop formation was also associated with the generation of negative DNA supercoils. These observations support the idea that the ATPase motors of the Snf2 family of proteins act as DNA translocases specialized to generate transient distortions in DNA structure.

Adenosine Triphosphatases↗
PubMed2003

Patterns of antipsychotic and anticholinergic prescribing for hospital inpatients.

The development of atypical antipsychotics has not only given the prescriber more options, but also increased the complexity of decision making. We examined current prescribing practice for antipsychotic and anticholinergic drugs, which involved a 1-day census of all antipsychotic and anticholinergic drugs prescribed for 4191 inpatients in 49 UK mental health services. Eighty-five percent of inpatients were prescribed antipsychotics, 48% of whom were prescribed more than one. Atypical antipsychotics were widely prescribed and combined with typicals in over 60% of cases. Large doses of antipsychotics were frequently prescribed 'as required'. The dose administered was always much less than the dose prescribed and nothing at all was administered against 79% of prescriptions. Antipsychotic prescribing often deviates from recommended practice. Nursing staff have considerable discretion to administer large doses of antipsychotics on an 'as required' basis.

Adolescent↗
PubMed2001

Magnetic resonance imaging of the wrist: diagnostic performance statistics.

AIM: To review the published diagnostic performance statistics for magnetic resonance imaging (MRI) of the wrist for tears of the triangular fibrocartilage complex, the intrinsic carpal ligaments, and for osteonecrosis of the carpal bones. MATERIALS AND METHODS: We used Medline and Embase to search the English language literature. Studies evaluating the diagnostic performance of MRI of the wrist in living patients with surgical confirmation of MR findings were identified. RESULTS: We identified 11 studies reporting the diagnostic performance of MRI for tears of the triangular fibrocartilage complex for a total of 410 patients, six studies for the scapho-lunate ligament (159 patients), six studies for the luno-triquetral ligament (142 patients) and four studies (56 patients) for osteonecrosis of the carpal bones. CONCLUSIONS: Magnetic resonance imaging is an accurate means of diagnosing tears of the triangular fibrocartilage and carpal osteonecrosis. Although MRI is highly specific for tears of the intrinsic carpal ligaments, its sensitivity is low. The diagnostic performance of MRI in the wrist is improved by using high-resolution T2* weighted 3D gradient echo sequences. Using current imaging techniques without intra-articular contrast medium, magnetic resonance imaging cannot reliably exclude tears of the intrinsic carpal ligaments. Hobby, J. L. (2001). Clinical Radiology, 56, 50-57.

Carpal Bones↗
PubMed2003

Non-toxic antifouling activity of polymeric 3-alkylpyridinium salts from the Mediterranean sponge Reniera sarai (Pulitzer-Finali).

The antifouling activity and toxicity of polymeric 3-alkylpyridinium salts (poly-APS) isolated from the Mediterranean sponge Reniera sarai were studied. The activity of these natural products was compared to that of zinc and copper complexes of pyrithione, two non-persistent booster biocides successfully used in current antifouling coatings. Larvae of Balanus amphitrite (cyprids and nauplii) were used to monitor settlement inhibition and the extent to which inhibition was due to toxicity. The microalga Tetraselmis suecica and larvae of the mussel Mytilus galloprovincialis were used in toxicity bioassays. Compared to the booster biocides, poly-APS were less effective at inhibiting cyprid settlement, but their effects were non toxic and reversible, with very low toxicity against the organisms used in the toxicity bioassays. Although encouraging, these results are not enough to warrant the use of poly-APS as a potential commercial antifoulant. They however justify possible future efforts to chemically synthesize poly-APS analogues for further tests.

Animals↗
PubMed2004

Exploring the complexities of prostate cancer screening with a view to supporting informed consent.

Men request to be screened for prostate cancer because they believe they are exhibiting responsible health promotion behaviour and there are definite benefits from the early detection of the disease. This belief about the benefits is in contrast to several national guidelines recommending that screening for prostate cancer not be done. Despite the guidelines men continue to request to be screened and doctors continue to supply screening tests to asymptomatic males. The lack of an appropriate screening test has been a key factor in supporting recommendations not to screen. However, recent studies show improved accuracy in the use of serum prostate specific antigens (PSA) as a screening tool. This implies that a revision of the guidelines might soon be appropriate. It is important that nurses and other health professionals are kept abreast of developments in prostate screening to assist men with their screening options to ensure their fully informed consent.

Evidence-Based Medicine↗
PubMed2004

Unifying temperature effects on the growth rate of bacteria and the stability of globular proteins.

The specific growth rate constant for bacterial growth does not obey the Arrhenius-type kinetics displayed by simple chemical reactions. Instead, for bacteria, steep convex curves are observed on an Arrhenius plot at the low- and high-temperature ends of the biokinetic range, with a region towards the middle of the growth range loosely approximating linearity. This central region has been considered by microbiologists to be the "normal physiological range" for bacterial growth, a concept whose meaningfulness we now question. We employ a kinetic model incorporating thermodynamic terms for temperature-induced enzyme denaturation, central to which is a term to account for the large positive heat capacity change during unfolding of the proteins within the bacteria. It is now widely believed by biophysicists that denaturation of complex proteins and/or other macromolecules is due to hydrophobic hydration of non-polar compounds. Denaturation is seen as the process by which enthalpic and entropic forces becomes imbalanced both at high and at low temperatures resulting in conformational changes in the enzyme structure that expose hydrophobic amino acid groups to the surrounding water molecules. The "thermodynamic" rate model, incorporating the heat capacity change and its effect on the enthalpy and entropy of the system, fitted 35 sets of data for psychrophilic, psychrotrophic, mesophilic and thermophilic bacteria well, resulting in biologically meaningful estimates for the important thermodynamic parameters. As these results mirror those obtained by biophysicists for globular proteins, it appears that the same or a similar mechanism applies to bacteria as applies to proteins.

Bacteria↗
PubMed2003

Meconium is a potent activator of complement in human serum and in piglets.

Meconium aspiration syndrome (MAS) is a clinical condition in the newborn infant with a significant morbidity and mortality. The complex pathophysiology of MAS, leading to both pulmonary and systemic complications, is characterized by an incompletely understood inflammatory reaction. Treatment is symptomatic, mainly limited to airway cleaning and ventilatory support. In this study, we show for the first time that meconium is a potent activator of complement, a key mediator of inflammation. In vitro, meconium activated the alternative complement pathway in human umbilical cord serum as judged by a substantial increase in the alternative pathway convertase C3bBbP. The activation proceeded through C3 (C3bc) and the terminal C5-9 pathway (terminal SC5b-9 complement complex), whereas the classical and lectin pathways were not activated (C1rs-C1-inhibitor complexes and C4bc). The lipid fraction, containing, e.g. free fatty acids, and the water fraction, containing, e.g. bile acids, contributed equally to the complement activation. A blocking antibody to factor D (alternative pathway) completely inhibited the meconium-induced complement activation, whereas blocking antibodies to mannose-binding lectin (lectin pathway) and C2 (classical and lectin pathway) had no effect. In vivo, meconium induced systemic complement activation in a piglet model of MAS, paralleling the increase in lung dysfunction. In conclusion, meconium is a potent activator of the complement system both in vitro and in vivo. Complement may be important in the pathogenesis of MAS, and specific complement inhibition might be a possible treatment approach in MAS.

Age Factors↗
PubMed2005

A unique RNA Fold in the RumA-RNA-cofactor ternary complex contributes to substrate selectivity and enzymatic function.

A single base (U1939) within E. coli 23S ribosomal RNA is methylated by its dedicated enzyme, RumA. The structure of RumA/RNA/S-adenosylhomocysteine uncovers the mechanism for achieving unique selectivity. The single-stranded substrate is "refolded" on the enzyme into a compact conformation with six key intra-RNA interactions. The RNA substrate contributes directly to catalysis. In addition to the target base, a second base is "flipped out" from the core loop to stack against the adenine of the cofactor S-adenosylhomocysteine. Nucleotides in permuted sequence order are stacked into the site vacated by the everted target U1939 and compensate for the energetic penalty of base eversion. The 3' hairpin segment of the RNA binds distal to the active site and provides binding energy that contributes to enhanced catalytic efficiency. Active collaboration of RNA in catalysis leads us to conclude that RumA and its substrate RNA may reflect features from the earliest RNA-protein era.

Bacterial Proteins↗
PubMed2004

Principles of droplet electrohydrodynamics for lab-on-a-chip.

Electrically controlled droplet-based labs-on-a-chip operate under the principles of electro-capillarity and dielectrophoresis. The microfluidic mechanics of manipulating electrified droplets are complex and not entirely understood. In this article, we analyse these operating principles, especially electrowetting on dielectric (a form of electro-capillarity) and dielectrophoresis, under a unified framework of droplet electrohydrodynamics. We differentiate them by their electric origins and their energy transduction mechanisms. Our study shows that both electrowetting on dielectric and dielectrophoresis are effective for droplet generation and manipulation. In addition, our study demonstrates: (1) the presence of a wetting contribution to dielectrophoresis; and (2) contact angle reduction is merely an observable consequence of, not a condition for, the occurrence of electrowetting on dielectric. Simulations are used extensively in this article to illustrate device operation, to expose underlying physics, and to validate our conclusions. Simulations of electrically driven droplet generation, droplet translocation, droplet fusion, and droplet fission are presented.

Electrophoresis, Capillary↗
PubMed2003

Crystal structures of 3-methyladenine DNA glycosylase MagIII and the recognition of alkylated bases.

DNA glycosylases catalyze the excision of chemically modified bases from DNA. Although most glycosylases are specific to a particular base, the 3-methyladenine (m3A) DNA glycosylases include both highly specific enzymes acting on a single modified base, and enzymes with broader specificity for alkylation-damaged DNA. Our structural understanding of these different enzymatic specificities is currently limited to crystal and NMR structures of the unliganded enzymes and complexes with abasic DNA inhibitors. Presented here are high-resolution crystal structures of the m3A DNA glycosylase from Helicobacter pylori (MagIII) in the unliganded form and bound to alkylated bases 3,9-dimethyladenine and 1,N6-ethenoadenine. These are the first structures of a nucleobase bound in the active site of a m3A glycosylase belonging to the helix-hairpin-helix superfamily. MagIII achieves its specificity for positively-charged m3A not by direct interactions with purine or methyl substituent atoms, but rather by stacking the base between two aromatic side chains in a pocket that excludes 7-methylguanine. We report base excision and DNA binding activities of MagIII active site mutants, together with a structural comparison of the HhH glycosylases.

Amino Acid Sequence↗
PubMed2005

Absence of thrombin-activatable fibrinolysis inhibitor protects against sepsis-induced liver injury in mice.

Thrombin-activatable fibrinolysis inhibitor (TAFI), also known as carboxypeptidase R, has been implicated as an important negative regulator of the fibrinolytic system. In addition, TAFI is able to inactivate inflammatory peptides such as complement factors C3a and C5a. To determine the role of TAFI in the hemostatic and innate immune response to abdominal sepsis, TAFI gene-deficient (TAFI-/-) and normal wild-type mice received an i.p. injection with Escherichia coli. Liver TAFI mRNA and TAFI protein concentrations increased during sepsis. In contrast to the presumptive role of TAFI as a natural inhibitor of fibrinolysis, TAFI-/- mice did not show any difference in E. coli-induced activation of coagulation or fibrinolysis, as measured by plasma levels of thrombin-anti-thrombin complexes and D-dimer and the extent of fibrin depositions in lung and liver tissues. However, TAFI-/- mice were protected from liver necrosis as indicated by histopathology and clinical chemistry. Furthermore, TAFI-/- mice displayed an altered immune response to sepsis, as indicated by an increased neutrophil recruitment to the peritoneal cavity and a transiently increased bacterial outgrowth together with higher plasma TNF-alpha and IL-6 levels. These data argue against an important part for TAFI in the regulation of the procoagulant-fibrinolytic balance in sepsis and reveals a thus far unknown role of TAFI in the occurrence of hepatic necrosis.

Animals↗
PubMed2004

Molecular mimicry of the NF-kappaB DNA target site by a selected RNA aptamer.

During the past two decades, structural and biophysical studies of DNA-protein and RNA-protein complexes have enhanced our understanding of the physico-chemical basis of nucleic acid recognition by proteins. However, it remains unclear what protein surface features are most important for nucleic acid binding and whether the same protein surface could bind specifically to both DNA and RNA. The recently described X-ray crystal structure of the transcription factor NF-kappaB p50 homodimer bound to a high-affinity RNA aptamer allows the direct comparison of NF-kappaB-RNA and NF-kappaB-DNA binding modes. The RNA aptamer, which bears no sequence homology to natural NF-kappaB DNA targets, adopts a structure with similar physico-chemical properties to kappaB DNA and contacts a common nucleic-acid-binding 'consensus surface' on the p50 homodimer.

Base Sequence↗
PubMed2002

The citrus methoxyflavone tangeretin affects human cell-cell interactions.

Two effects of the citrus methoxyflavone tangeretin on cell-cell interactions are biologically relevant. Firstly, tangeretin upregulates the function of the E-cadherin/catenin complex in human MCF-7/6 breast carcinoma cells. This leads to firm cell-cell adhesion and inhibition of invasion in vitro. Secondly, tangeretin downregulates the interleukin-2 receptor on T-lymphocytes and natural killer cells. This leads to a decrease in the cytotoxic competence of these immunocytes against cancer cells. The second effect can become important when high doses of tangeretin are combined with adjuvant tamoxifen treatment for breast cancer. Experiments with nude mice bearing MCF-7/6 tumors showed that tangeretin given orally at high doses, abrogated the therapeutic suppression of tumor growth exerted by tamoxifen. No evidence for a tumor promoting effect of tangeretin by itself was found in these experiments. Tangeretin may be an interesting molecule for application in cases where immunosuppression could be clinically beneficial.

Animals↗
PubMed2006

Safety and pharmacokinetics of recombinant factor XIII-A2 administration in patients with congenital factor XIII deficiency.

Congenital factor XIII (FXIII) deficiency is associated with a tendency for severe bleeding, a risk for spontaneous abortion, and a high rate of spontaneous intracranial hemorrhage. This phase 1 escalating-dose study was developed to evaluate the safety and pharmacokinetics of a single administration of human recombinant FXIII-A2 (rFXIII-A2) homodimer in adults with congenital FXIII deficiency. Pharmacokinetics and activity of rXIII and changes in endogenous B subunit levels were assessed. Recombinant FXIII-A2 homodimer were complexed with endogenous FXIII-B subunits to form an FXIII-A2B2 heterotetramer with a half-life of 8.5 days, similar to that of endogenous FXIII. The median dose response was a 2.4% increase in FXIII activity based on unit per kilogram rFXIII administered. After the administration of rFXIII-A2, clot solubility normalized as measured by clot lysis in urea. Clot strength and resistance to fibrinolysis, as assessed by thromboelastography, also improved. Safety reviews were conducted before each dose escalation; no serious adverse events, including bleeding or thrombosis, were noted during the study. In addition, there was no evidence of the generation of specific antibodies to rFXIII or yeast proteins. Recombinant FXIII appears to be a safe and potentially effective alternative for FXIII replacement in patients with FXIII deficiency.

Adult↗
PubMed2002

Epitope mapping of the immunodominant antigen TB10.4 and the two homologous proteins TB10.3 and TB12.9, which constitute a subfamily of the esat-6 gene family.

The human T-cell recognition of the low-molecular-mass culture filtrate antigen TB10.4 was evaluated in detail. The molecule was strongly recognized by T cells isolated from tuberculosis (TB) patients and from BCG-vaccinated donors. The epitopes on TB10.4 were mapped with overlapping peptides and found to be distributed throughout the molecule. The broadest response was found in TB patients, whereas the response in BCG-vaccinated donors was focused mainly toward a dominant epitope located in the N terminus (amino acids 1 to 18). The gene encoding TB10.4 was found to belong to a subfamily within the esat-6 family that consists of the three highly homologous proteins TB10.4, TB10.3, and TB12.9 (Rv0288, Rv3019c, and Rv3017c, respectively). Southern blot analysis combined with database searches revealed that the three members of the TB10.4 family were present only in strains of the Mycobacterium tuberculosis complex, including BCG, and M. kansasii, whereas other atypical mycobacteria had either one (M. avium, M. intracellulare, and M. marinum) or none (M. scrofulaceum, M. fortuitum, and M. szulgai) of the genes. The fine specificity of the T-cell response to the three closely related esat-6 family members was markedly different, with only a few epitopes shared between the molecules. Minimal differences in the amino acid sequence translated into large differences in recognition by T cells and secretion of gamma interferon. In general, the peptides from TB10.4 stimulated the largest responses, but epitopes unique to both TB10.3 and TB12.9 were found. The relevance of the findings for TB vaccine development and as a potential mechanism for immune evasion is discussed.

Amino Acid Sequence↗
PubMed2005

Crystallographic refinement by knowledge-based exploration of complex energy landscapes.

Although X-ray crystallography remains the most versatile method to determine the three-dimensional atomic structure of proteins and much progress has been made in model building and refinement techniques, it remains a challenge to elucidate accurately the structure of proteins in medium-resolution crystals. This is largely due to the difficulty of exploring an immense conformational space to identify the set of conformers that collectively best fits the experimental diffraction pattern. We show here that combining knowledge-based conformational sampling in RAPPER with molecular dynamics/simulated annealing (MD/SA) vastly improves the quality and power of refinement compared to MD/SA alone. The utility of this approach is highlighted by the automated determination of a lysozyme mutant from a molecular replacement solution that is in congruence with a model prepared independently by crystallographers. Finally, we discuss the implications of this work on structure determination in particular and conformational sampling and energy minimization in general.

Amino Acid Sequence↗
PubMed2005

Statins affect cell-surface expression of major histocompatibility complex class II molecules by disrupting cholesterol-containing microdomains.

Statins, the main therapy for hypercholesterolemia, are currently considered as possible immunomodulatory agents. Statins inhibit the production of proinflammatory cytokines and reduce the expression of several immunoregulatory molecules, including major histocompatibility complex class II (MHC-II) molecules. In this study, we investigated the mechanism by which simvastatin reduces the membrane expression of MHC-II molecules on several human cell types. We demonstrate that the reduction of MHC-II membrane expression by simvastatin correlates with disruption of cholesterol-containing microdomains, which transport and concentrate MHC-II molecules to the cell surface. In addition, we demonstrate that statins reduce cell-surface expression of other immunoregulatory molecules, which include MHC-I, CD3, CD4, CD8, CD28, CD40, CD80, CD86, and CD54. Our observations indicate that the downregulation of MHC-II at the cell surface contributes to the immunomodulatory properties of statins and is achieved through disruption of cholesterol-containing microdomains, which are involved in their intracellular transport.

Cell Line, Tumor↗
PubMed2006

Crystal structure of the mitochondrial chaperone TIM9.10 reveals a six-bladed alpha-propeller.

Import of proteins into mitochondria occurs by coordinated actions of preprotein translocases in the outer and inner membranes. Tim9 and Tim10 are translocase components of the intermembrane space, related to deafness-dystonia peptide 1 (DDP1). They coassemble into a hexamer, TIM9.10, which captures and chaperones precursors of inner membrane metabolite carriers as they exit the TOM channel in the outer membrane. The crystal structure of TIM9.10 reveals a previously undescribed alpha-propeller topology in which helical "blades" radiate from a narrow central pore lined with polar residues. The propeller blades are reminiscent of "tentacles" in chaperones Skp and prefoldin. In each TIM9.10 subunit, a signature "twin CX3C" motif forms two intramolecular disulfides. There is no obvious binding pocket for precursors, which we suggest employ the chaperone-like tentacles of TIM9.10 as surrogate lipid contacts. The first reported crystal structure of a mitochondrial translocase assembly provides insights into selectivity and regulation of precursor import.

Amino Acid Sequence↗