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Characterization of recombinant respiratory syncytial viruses with the region responsible for type 2 T-cell responses and pulmonary eosinophilia deleted from the attachment (G) protein 总被引:1,自引:0,他引:1 下载免费PDF全文
Elliott MB Pryharski KS Yu Q Boutilier LA Campeol N Melville K Laughlin TS Gupta CK Lerch RA Randolph VB LaPierre NA Dack KM Hancock GE 《Journal of virology》2004,78(16):8446-8454
It is essential that preventative vaccines for respiratory syncytial virus (RSV) elicit balanced T-cell responses. Immune responses dominated by type 2 T cells against RSV antigens are believed to cause exaggerated respiratory tract disease and may also contribute to unwanted inflammation in the airways that predisposes infants to wheeze through adolescence. Here we report on the construction and characterization of recombinant RSV (rRSV) strains with amino acids 151 to 221 or 178 to 219 of the attachment (G) glycoprotein deleted (rA2cpDeltaG150-222 or rA2cpDeltaG177-220, respectively). The central ectodomain was chosen for modification because a peptide spanning amino acids 149 to 200 of G protein has recently been shown to prime several strains of na?ve inbred mice for polarized type 2 T-cell responses, and peripheral blood T cells from most human donors recognize epitopes within this region. Quantitative PCR demonstrated that synthesis of nascent rRSV genomes in human lung epithelial cell lines was similar to that for the parent virus (cp-RSV). Plaque assays further indicated that rRSV replication was not sensitive to 37 degrees C, but pinpoint morphology was observed at 39 degrees C. Both rRSV strains replicated in the respiratory tracts of BALB/c mice and elicited serum neutralization and anti-F-protein immunoglobulin G titers that were equivalent to those elicited by cp-RSV and contributed to a 3.9-log(10)-unit reduction in RSV A2 levels 4 days after challenge. Importantly, pulmonary eosinophilia was significantly diminished in BALB/c mice primed with native G protein and challenged with either rA2cpDeltaG150-222 or rA2cpDeltaG177-220. These findings are important for the development of attenuated RSV vaccines. 相似文献
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Belánová M Dianisková P Brennan PJ Completo GC Rose NL Lowary TL Mikusová K 《Journal of bacteriology》2008,190(3):1141-1145
Two galactosyl transferases can apparently account for the full biosynthesis of the cell wall galactan of mycobacteria. Evidence is presented based on enzymatic incubations with purified natural and synthetic galactofuranose (Galf) acceptors that the recombinant galactofuranosyl transferase, GlfT1, from Mycobacterium smegmatis, the Mycobacterium tuberculosis Rv3782 ortholog known to be involved in the initial steps of galactan formation, harbors dual β-(1→4) and β-(1→5) Galf transferase activities and that the product of the enzyme, decaprenyl-P-P-GlcNAc-Rha-Galf-Galf, serves as a direct substrate for full polymerization catalyzed by another bifunctional Galf transferase, GlfT2, the Rv3808c enzyme. 相似文献
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Tizazu H. Mekonnen Paolo G. Mussone Natisha Stashko Phillip Y. Choi David C. Bressler 《Process Biochemistry》2013,48(5-6):885-892
The purpose of this study was to characterize proteinacious material recovered from cattle tissues known as specified risk materials (SRM) hydrolyzed according to two protocols approved by the Canadian Food Inspection Agency and Food and Drug Administration and to evaluate its potential for non-food/feed value-added applications. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), size exclusion high performance liquid chromatography (SEC-HPLC), and free amino acid determination methods were adopted to measure the extent of protein cleavage and the average molecular weight of hydrolyzates, and to identify the reactive functionalities available. Molecular weight analysis revealed that the alkaline hydrolysis protocol cleaved the proteins more severely than thermal hydrolysis. Both protocols narrowed the molecular weight distribution of the proteins and increased their solubility in water. Several active functional groups amenable to further chemical processing were identified in the protein hydrolyzates. In conclusion, this work demonstrated that both hydrolytic protocols can be adopted as pre-processing steps to produce safe and renewable proteinacious feedstock for industrial applications. 相似文献
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Backes BJ Longenecker K Hamilton GL Stewart K Lai C Kopecka H von Geldern TW Madar DJ Pei Z Lubben TH Zinker BA Tian Z Ballaron SJ Stashko MA Mika AK Beno DW Kempf-Grote AJ Black-Schaefer C Sham HL Trevillyan JM 《Bioorganic & medicinal chemistry letters》2007,17(7):2005-2012
A novel series of pyrrolidine-constrained phenethylamines were developed as dipeptidyl peptidase IV (DPP4) inhibitors for the treatment of type 2 diabetes. The cyclohexene ring of lead-like screening hit 5 was replaced with a pyrrolidine to enable parallel chemistry, and protein co-crystal structural data guided the optimization of N-substituents. Employing this strategy, a >400x improvement in potency over the initial hit was realized in rapid fashion. Optimized compounds are potent and selective inhibitors with excellent pharmacokinetic profiles. Compound 30 was efficacious in vivo, lowering blood glucose in ZDF rats that were allowed to feed freely on a mixed meal. 相似文献
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Detection of Bacteriocins by Matrix-Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry 总被引:1,自引:1,他引:0 下载免费PDF全文
The use of matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for the detection of bacteriocins was investigated. A 30-s water wash of the sample on the MALDI-TOF MS probe was effective in removing contaminants of the analyte. This method was used for rapid detection of nisin, pediocin, brochocin A and B, and enterocin A and B from culture supernatants and for detection of enterocin B throughout its purification. 相似文献
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Longenecker KL Stewart KD Madar DJ Jakob CG Fry EH Wilk S Lin CW Ballaron SJ Stashko MA Lubben TH Yong H Pireh D Pei Z Basha F Wiedeman PE von Geldern TW Trevillyan JM Stoll VS 《Biochemistry》2006,45(24):7474-7482
Dipeptidyl peptidase IV (DPP-IV) belongs to a family of serine peptidases, and due to its indirect regulatory role in plasma glucose modulation, DPP-IV has become an attractive pharmaceutical target for diabetes therapy. DPP-IV inactivates the glucagon-like peptide (GLP-1) and several other naturally produced bioactive peptides that contain preferentially a proline or alanine residue in the second amino acid sequence position by cleaving the N-terminal dipeptide. To elucidate the details of the active site for structure-based drug design, we crystallized a natural source preparation of DPP-IV isolated from rat kidney and determined its three-dimensional structure using X-ray diffraction techniques. With a high degree of similarity to structures of human DPP-IV, the active site architecture provides important details for the design of inhibitory compounds, and structures of inhibitor-protein complexes offer detailed insight into three-dimensional structure-activity relationships that include a conformational change of Tyr548. Such accommodation is exemplified by the response to chemical substitution on 2-cyanopyrrolidine inhibitors at the 5 position, which conveys inhibitory selectivity for DPP-IV over closely related homologues. A similar conformational change is also observed in the complex with an unrelated synthetic inhibitor containing a xanthine core that is also selective for DPP-IV. These results suggest the conformational flexibility of Tyr548 is unique among protein family members and may be utilized in drug design to achieve peptidase selectivity. 相似文献