bluuebunny/crossref_metadata_2025_split huggingface dataset:
Probing flexibility and “induced‐fit” phenomena in aldose reductase by comparative crystal structure analysis and molecular dynamics simulations
Authors: Sotriffer Christoph A. , Krämer Oliver , Klebe Gerhard 
Published: April 2004

AbstractAldose reductase is a promising target for the treatment of diabetic complications, and as such, has become the focus of various drug design projects. As revealed by a survey of available crystal structures, the protein shows pronounced induced‐fit effects upon ligand binding. Although helping to explain the enzyme's substrate promiscuity, phenomena of this kind are still responsible for significant complications in structure‐based design efforts directed to aldose reductase. Accordingly, a deeper understanding of the principles governing conformational alterations in this enzyme would be of utmost practical importance. As a first step in addressing this issue, molecular dynamics (MD) simulations have been carried out. The ultrahigh resolution crystal structure of aldose reductase complexed with inhibitor IDD594 served as ideal starting point for a set of different simulations of nanosecond time scale: the native complexed state with bound inhibitor, the uncomplexed state (after removal of the inhibitor) at standard temperature, and the uncomplexed state at elevated temperature. The reference simulation of the complex exhibits extraordinary stability of the overall fold, whereas two distinct conformational substates are found for the binding‐site region. In contrast, already at standard temperature pronounced changes are observed in the binding region during the simulation of the uncomplexed state. Leu300, for example, closes the access to the pocket opened by IDD594. On the other hand, conformations around the catalytic site are highly conserved, with the His110–Tyr48–NADP+ orientation being stabilized by a water molecule. Detailed analysis of the trajectories allows to reveal a set of distinct conformational substates that may prove useful as alternative structural templates in virtual screening for new aldose reductase inhibitors. Proteins 2004. © 2004 Wiley‐Liss, Inc.
Structural and sequence comparisons arising from the solution structure of the transcription elongation factor NusG from Thermus thermophilus
Authors: Reay Paul , Yamasaki Kazuhiko , Terada Takaho , Kuramitsu Seiki , Shirouzu Mikako , Yokoyama Shigeyuki 
Published: April 2004

AbstractNusG is an essential bacterial protein modulator of transcriptional elongation and termination events, and interacts directly with RNA polymerase and Rho protein. Found also in Archaea, NusG shows stretches of sequence similarity to the eukaryotic transcription elongation factor Spt5. Herein, the three‐dimensional solution structure of the bacterial NusG from Thermus thermophilus, which shows 43% amino acid sequence similarity to the Escherichia coli NusG, is described, and a survey of NusG and Spt5 amino acid sequences is presented. Although there is a clear evolutionary and functional relationship between these proteins, it is evident from the structural, sequence, and biochemical data that their binding specificities to both nucleic acids and other proteins differ. Proteins 2004. © 2004 Wiley‐Liss, Inc.
A detailed comparison of current docking and scoring methods on systems of pharmaceutical relevance
Authors: Perola Emanuele , Walters W. Patrick , Charifson Paul S. 
Published: April 2004

AbstractA thorough evaluation of some of the most advanced docking and scoring methods currently available is described, and guidelines for the choice of an appropriate protocol for docking and virtual screening are defined. The generation of a large and highly curated test set of pharmaceutically relevant protein–ligand complexes with known binding affinities is described, and three highly regarded docking programs (Glide, GOLD, and ICM) are evaluated on the same set with respect to their ability to reproduce crystallographic binding orientations. Glide correctly identified the crystallographic pose within 2.0 Å in 61% of the cases, versus 48% for GOLD and 45% for ICM. In general Glide appears to perform most consistently with respect to diversity of binding sites and ligand flexibility, while the performance of ICM and GOLD is more binding site–dependent and it is significantly poorer when binding is predominantly driven by hydrophobic interactions. The results also show that energy minimization and reranking of the top N poses can be an effective means to overcome some of the limitations of a given docking function. The same docking programs are evaluated in conjunction with three different scoring functions for their ability to discriminate actives from inactives in virtual screening. The evaluation, performed on three different systems (HIV‐1 protease, IMPDH, and p38 MAP kinase), confirms that the relative performance of different docking and scoring methods is to some extent binding site–dependent. GlideScore appears to be an effective scoring function for database screening, with consistent performance across several types of binding sites, while ChemScore appears to be most useful in sterically demanding sites since it is more forgiving of repulsive interactions. Energy minimization of docked poses can significantly improve the enrichments in systems with sterically demanding binding sites. Overall Glide appears to be a safe general choice for docking, while the choice of the best scoring tool remains to a larger extent system‐dependent and should be evaluated on a case‐by‐case basis. Proteins 2004. © 2004 Wiley‐Liss, Inc.
Determination of carnitine and acylcarnitines in plasma by high‐performance liquid chromatography/electrospray ionization ion trap tandem mass spectrometry
Authors: Vernez Laurence , Wenk Markus , Krähenbühl Stephan 
Published: April 2004

AbstractA high‐performance liquid chromatography/mass spectrometry method was developed for the determination of carnitine, its biosynthetic precursor butyrobetaine, and eight acylcarnitines in plasma. The procedure includes a solid‐phase extraction for carnitine and short‐ and medium‐chain acylcarnitines, and a liquid‐liquid extraction for protein‐bound long‐chain acylcarnitines, followed by separation on a reversed‐phase column in the presence of a volatile ion‐pairing reagent. Detection was achieved using an ion‐trap mass spectrometer run in the tandem mass spectrometry (MS/MS) mode. The choice of the matrix for calibrators, used for quantification of these endogenous compounds, was also investigated. Validation was performed for standard quality controls diluted with 4% bovine serum albumin solution and for spiked plasma quality control samples at concentrations between 0.5 and 80 μmol/L, depending on the compound. Intra‐ and inter‐day precisions for the determination of carnitine were below 3.4% and accuracies were between 95.2 and 109.0%. Application of the method to the diagnosis of pathological acylcarnitine profiles of metabolic disorders in a patient suffering from methylmalonic aciduria is presented. The method allows quantification of carnitine, butyrobetaine, acetylcarnitine and propionylcarnitine, and semiquantitative analysis of medium‐ and long‐chain acylcarnitines. In contrast with other methods, no derivatization step is needed. Copyright © 2004 John Wiley & Sons, Ltd.
Stable carbon isotope analysis of different tissues of beef animals in relation to their diet
Authors: De Smet Stefaan , Balcaen An , Claeys Erik , Boeckx Pascal , Van Cleemput Oswald 
Published: April 2004

AbstractAs part of a larger experiment, 31 young bulls, divided into three groups, were given different diets containing either C3 plants or a combination of C3 and C4 plant‐based feeds in three feeding periods before slaughter. Variation in the proportion of C4 plant material in the diets was made by including or not maize or maize‐derived ingredients, whereas the other dietary constituents were from C3 plants. Analysis of stable carbon isotope ratios (δ13C value) was performed on different tissues taken at slaughter: blood, plasma, liver, kidney fat, hair, muscle and ruminal contents. Blood and plasma samples were also taken at the beginning of each period. A highly significant difference was found in the δ13C values of blood and plasma samples taken from animals that had received a diet of only C3 plants or with 59% C4 material for 70 days. The δ13C values of all different samples taken at slaughter were highly significantly different between the three feeding groups that had received diets with 0, 13.5 or 35% C4 material for on average 137, 139 and 83 days, respectively. For the three groups, samples of hair, muscle, plasma, whole blood and liver were significantly enriched in 13C compared with the diet (except for liver in one group), whereas kidney fat was significantly depleted. The proportion of C4 plant material could be accurately estimated from the δ13C values of different tissue samples. Stable carbon analysis of different tissues from beef animals can be used to trace back diets containing variable proportions of C3 and C4 plant material. Copyright © 2004 John Wiley & Sons, Ltd.
“You can exercise your way out of HIV” and other stories: The role of biological knowledge in adolescents' evaluation of myths
Authors: Keselman Alla , Kaufman David R. , Patel Vimla L. 
Published: April 2004

AbstractA primary objective for science education is to impart robust knowledge that has applicability to real‐world problems. This article presents research investigating the relationship between adolescents' conceptual understanding of the biological basis of HIV and critical reasoning. Middle and high school students were interviewed about their understanding of HIV and were subsequently asked to evaluate scenarios that contained myths about HIV. On the basis of their responses to the interview questions, students' understanding of HIV was categorized into three models, naïve, intermediate, and advanced. The results indicate that knowledge mediated students' responses in specific ways. Students at different levels of HIV knowledge reasoned in qualitatively different ways about the myths. A significant relationship was found between students' understanding of HIV biology and the level of biological reasoning. We found that students who employed cellular‐level biological reasoning were more likely to reject the myths than students who employed just system‐level reasoning or nonspecific biological reasoning. The findings emphasize the importance of conceptual understanding in the critical evaluation of information that may serve as a basis for making decisions about HIV. We conclude with discussing the implications of the findings for science and health education. © 2004 Wiley Periodicals, Inc. Sci Ed 88:548–573, 2004
Sexual experience alters D1 receptor–mediated cyclic AMP production in the nucleus accumbens of female Syrian hamsters
Authors: Bradley Katherine C. , Mullins Amanda J. , Meisel Robert L. , Watts Val J. 
Published: April 2004

AbstractDrugs of abuse produce long‐term changes in dopamine neurotransmission and receptor‐effected intracellular signaling. Similar changes in neuronal activity are mediated by motivated behaviors. To explore cellular mechanisms underlying these neuroadaptations following sexual experience, cyclic AMP accumulation following stimulation of D1 dopamine receptors, G‐proteins, and adenylate cyclase was compared in the nucleus accumbens and caudate nucleus of sexually naive and experienced female hamsters following sexual behavior. Direct stimulation of adenylate cyclase with forskolin or indirectly by activation of G‐proteins with Gpp(NH)p produced dose‐dependent increases in the formation of cyclic AMP in the nucleus accumbens and caudate nucleus, with no effects of sexual experience on these measures. Specific D1 receptor stimulation increased Gpp(NH)p‐induced adenylate cyclase activity in the nucleus accumbens and caudate nucleus of all animals. Interestingly, this stimulation was further enhanced only in membranes from the nucleus accumbens, but not from the caudate nucleus, of sexually experienced hamsters compared to the response of naive females. These results demonstrate that sexual behavior experience can sensitize mesolimbic dopamine pathways and that this sensitization occurs through an increase in D1 receptor–mediated signaling. Synapse 53:20–27, 2004. © 2004 Wiley‐Liss, Inc.
Osteoplastic Flap for Obliteration of the Frontal Sinus: Five years’ Experience
Authors: Correa Alex J. , Duncavage James A. , Fortune D. Scott , Reinisch Lou 
Published: April 2004

The objective of this retrospective study was to evaluate the osteoplastic flap (OPF) for the obliteration of the frontal sinus in this current era of endoscopic management of frontal sinus disease. A review of consecutive OPF procedures (n = 43) performed by the senior author (J.A.D.) from 1992 to 1997 was carried out. Data were gathered regarding chief symptom, medical history, previous sinus surgery, endoscopic findings in the office and at surgery, CT scan findings, and follow‐up results (mean 19.4 months). Previous endoscopic management of frontal sinus disease had failed in 24% of patients; 97% had eventual resolution of frontal sinusitis with OPF. After OPF, 63% also had improvement or resolution of disease in other paranasal sinuses. Statistically significant, Positive correlations (P< 0.05) were noted between the resolution of frontal sinusitis and improved or resolved pain, as well as the resolution of frontal sinusitis and improved or resolved infections in other paranasal sinuses. In 1998 OPF remains the standard for treating frontal sinus disease refractory to other methods. OPF can decrease the pain associated with frontal sinus infections and has a Positive impact on inflammatory disease in other paranasal sinuses.
Changes of the Secondary Structure of the 5′ End of the Sindbis Virus Genome Inhibit Virus Growth in Mosquito Cells and Lead to Accumulation of Adaptive Mutations
Authors: Fayzulin Rafik , Frolov Ilya 
Published: April 2004

ABSTRACT

Both the 5′ end of the Sindbis virus (SIN) genome and its complement in the 3′ end of the minus-strand RNA synthesized during virus replication serve as parts of the promoters recognized by the enzymes that comprise the replication complex (RdRp). In addition to the 5′ untranslated region (UTR), which was shown to be critical for the initiation of replication, another 5′ sequence element, the 51-nucleotide (nt) conserved sequence element (CSE), was postulated to be important for virus replication. It is located in the nsP1-encoding sequence and is highly conserved among all members of the
Alphavirus
genus. Studies with viruses containing clustered mutations in this sequence demonstrated that this RNA element is dispensable for SIN replication in cells of vertebrate origin, but its integrity can enhance the replication of SIN-specific RNAs. However, we showed that the same mutations had a deleterious effect on virus replication in mosquito cells. SIN with a mutated 51-nt CSE rapidly accumulated adaptive mutations in the nonstructural proteins nsP2 and nsP3 and the 5′ UTR. These mutations functioned synergistically in a cell-specific manner and had a stimulatory effect only on the replication of viruses with a mutated 51-nt CSE. Taken together, the results suggest the complex nature of interactions between nsP2, nsP3, the 5′ UTR, and host-specific protein factors binding to the 51-nt CSE and involved in RdRp formation. The data also demonstrate an outstanding potential of alphaviruses for adaptation. Within one passage, SIN can adapt to replication in cells of a vertebrate or invertebrate origin.
Treatment of Transmissible Spongiform Encephalopathy by Intraventricular Drug Infusion in Animal Models
Authors: Doh-ura Katsumi , Ishikawa Kensuke , Murakami-Kubo Ikuko , Sasaki Kensuke , Mohri Shirou , Race Richard , Iwaki Toru 
Published: April 2004

ABSTRACT
The therapeutic efficacy of direct drug infusion into the brain, the target organ of transmissible spongiform encephalopathies, was assessed in transgenic mice intracerebrally infected with 263K scrapie agent. Pentosan polysulfate (PPS) gave the most dramatic prolongation of the incubation period, and amphotericin B had intermediate effects, but antimalarial drugs such as quinacrine gave no significant prolongation. Treatment with the highest dose of PPS at an early or late stage of the infection prolonged the incubation time by 2.4 or 1.7 times that of the control mice, respectively. PPS infusion decreased not only abnormal prion protein deposition but also neurodegenerative changes and infectivity. These alterations were observed within the brain hemisphere fitted with an intraventricular infusion cannula but not within the contralateral hemisphere, even at the terminal disease stage long after the infusion had ended. Therapeutic effects of PPS were also demonstrated in mice infected with either RML agent or Fukuoka-1 agent. However, at doses higher than that providing the maximal effects, intraventricular PPS infusion caused adverse effects such as hematoma formation in the experimental animals. These findings indicate that intraventricular PPS infusion might be useful for the treatment of transmissible spongiform encephalopathies in humans, providing that the therapeutic dosage is carefully evaluated.
Prediction and Identification of a Permissive Epitope Insertion Site in the Vesicular Stomatitis Virus Glycoprotein
Authors: Schlehuber Lisa D. , Rose John K. 
Published: April 2004

ABSTRACT
We developed a rational approach to identify a site in the vesicular stomatitis virus (VSV) glycoprotein (G) that is exposed on the protein surface and tolerant of foreign epitope insertion. The foreign epitope inserted was the six-amino-acid sequence ELDKWA, a sequence in a neutralizing epitope from human immunodeficiency virus type 1. This sequence was inserted into six sites within the VSV G protein (Indiana serotype). Four sites were selected based on hydrophilicity and high sequence variability identified by sequence comparison with other vesiculovirus G proteins. The site showing the highest variability was fully tolerant of the foreign peptide insertion. G protein containing the insertion at this site folded correctly, was transported normally to the cell surface, had normal membrane fusion activity, and could reconstitute fully infectious VSV. The virus was neutralized by the human 2F5 monoclonal antibody that binds the ELDKWA epitope. Additional studies showed that this site in G protein tolerated insertion of at least 16 amino acids while retaining full infectivity. The three other insertions in somewhat less variable sequences interfered with VSV G folding and transport to the cell surface. Two additional insertions were made in a conserved sequence adjacent to a glycosylation site and near the transmembrane domain. The former blocked G-protein transport, while the latter allowed transport to the cell surface but blocked membrane fusion activity of G protein. Identification of an insertion-tolerant site in VSV G could be important in future vaccine and targeting studies, and the general principle might also be useful in other systems.
Murine Gammaherpesvirus 68 Lacking gp150 Shows Defective Virion Release but Establishes Normal Latency In Vivo
Authors: de Lima Brigitte D. , May Janet S. , Stevenson Philip G. 
Published: April 2004

ABSTRACT
All gammaherpesviruses encode a virion glycoprotein positionally homologous to Epstein-Barr virus gp350. These glycoproteins are thought to be involved in cell binding, but little is known of the roles they might play in the whole viral replication cycle. We have analyzed the contribution of murine gammaherpesvirus 68 (MHV-68) gp150 to viral propagation in vitro and host colonization in vivo. MHV-68 lacking gp150 was viable and showed normal binding to fibroblasts and normal single-cycle lytic replication. Its capacity to infect glycosaminoglycan (GAG)-deficient CHO-K1 cells and NS0 and RAW264.7 cells, which express only low levels of GAGs, was paradoxically increased. However, gp150-deficient MHV-68 spread poorly through fibroblast monolayers, with reduced cell-free infectivity, consistent with a deficit in virus release. Electron microscopy showed gp150-deficient virions clustered on infected-cell plasma membranes. MHV-68-infected cells showed reduced surface GAG expression, suggesting that gp150 prevented virions from rebinding to infected cells after release by making MHV-68 infection GAG dependent. Surprisingly, gp150-deficient viruses showed only a transient lag in lytic replication in vivo and established normal levels of latency. Cell-to-cell virus spread and the proliferation of latently infected cells, for which gp150 was dispensable, therefore appeared to be the major route of virus propagation in an infected host.
Rapid and Sensitive Detection of Retrovirus Entry by Using a Novel Luciferase-Based Content-Mixing Assay
Authors: Kolokoltsov Andrey A. , Davey Robert A. 
Published: April 2004

ABSTRACT
We describe a novel assay that permits measurement of entry of murine leukemia virus and pseudotypes with greater sensitivity and more rapidly than previously possible. To achieve this, we encapsulated a sensitive reporter enzyme, luciferase, directly into fully infectious, intact viral particles. The enzyme is specifically targeted to the viral lumen, as a C-terminal fusion on the viral envelope protein. Only when the incorporated luciferase is released from the viral lumen and gains access to its substrates is light emitted and readily detected. When cells are perfused with luciferin, quantitative measurements of entry can be made in real time on live cells. Uniquely, the amount of cell-bound virus can be determined in the same assay by addition of detergent to expose the luciferase. We demonstrate that virus carrying a mutation in the fusion peptide binds normally to cells but is unable to infect them and gives no entry signal. Using this assay, we show that inhibitors of endosomal acidification inhibit signal from vesicular stomatitis virus pseudotypes but not murine leukemia virus, consistent with a pH-independent mode of entry for the latter virus. Additionally, the fusion kinetics are rapid, with a half-life of 25 min after a delay of 10 to 15 min. The future use of this assay will permit a detailed examination of the entry mechanism of viruses and provide a convenient platform to discover novel entry inhibitors. The design also permits packaging of potential therapeutic protein cargoes into functional virus particles and their specific delivery to cellular targets.
Accumulation of a 3′-Terminal Genome Fragment in Japanese Encephalitis Virus-Infected Mammalian and Mosquito Cells
Authors: Lin Kuo-Chih , Chang Huei-Lan , Chang Ruey-Yi 
Published: April 2004

ABSTRACT
Japanese encephalitis virus (JEV) contains a single positive-strand RNA genome nearly 11 kb in length and is not formally thought to generate subgenomic RNA molecules during replication. Here, we report the abundant accumulation of a 3′-terminal 521- to 523-nucleotide (nt) genome fragment, representing a major portion of the 585-nt 3′ untranslated region, in both mammalian (BHK-21) and mosquito (C6/36) cells infected with any of nine strains of JEV. In BHK-21 cells, the viral genome was detected as early as 24 h postinfection, the small RNA was detected as early as 28 h postinfection, and the small RNA was 0.25 to 1.5 times as abundant as the genome on a molar basis between 28 and 48 h postinfection. In C6/36 cells, the genome and small RNA were present 5 days postinfection and the small RNA was 1.25 to 5.14 times as abundant as the genome. The 3′-terminal 523-nt small RNA contains a 5′-proximal stable hairpin (nt 6 to 56) that may play a role in its formation and the conserved flavivirus 3′-cyclization motif (nt 413 to 420) and the 3′-terminal long stable hairpin structure (nt 440 to 523) that have postulated roles in genome replication. Abundant accumulation of the small RNA during viral replication in both mammalian and mosquito cells suggests that it may play a biological role, perhaps as a regulator of RNA synthesis.