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1.
Importance of cysteines in the LDLR-related domain of the subgroup A avian leukosis and sarcoma virus receptor for viral entry. 总被引:6,自引:6,他引:0
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The extracellular domain of the subgroup A avian sarcoma and leukemia virus (ALSV-A) receptor contains a region that is related in sequence to the ligand-binding motifs of the low-density lipoprotein receptor (LDLR). This domain contains six cysteines that are highly conserved between different members of the LDLR protein superfamily, and these residues are presumed to participate in intrachain disulfide bonds. To assess the importance of each cysteine in the ALSV-A receptor, individual or multiple cysteines were mutated to alanines and the altered receptors were tested for the ability to confer susceptibility to viral infection. Receptors bearing single mutations allowed subgroup A viral entry, albeit at less than wild-type levels. Receptors containing two or three substitutions were completely inactive if one of the changed residues was Cys-35 or Cys-50. Of the altered receptors tested, the only exception to this rule was a functional receptor which lacked both Cys-35 and Cys-50, an activity that was dependent on the presence of other cysteines in this protein. Most interestingly, a receptor containing both Cys-35 and Cys-50 but lacking the other four cysteines was completely functional. These results demonstrate the importance of Cys-35 and Cys-50 for viral entry mediated by the ALSV-A receptor and show that in the presence of these two residues, all of the other cysteines in this protein can be removed without loss of this function. 相似文献
2.
Repanas K Zingler N Layer LE Schumann GG Perrakis A Weichenrieder O 《Nucleic acids research》2007,35(14):4914-4926
The human LINE-1 endonuclease (L1-EN) is the targeting endonuclease encoded by the human LINE-1 (L1) retrotransposon. L1-EN guides the genomic integration of new L1 and Alu elements that presently account for ~28% of the human genome. L1-EN bears considerable technological interest, because its target selectivity may ultimately be engineered to allow the site-specific integration of DNA into defined genomic locations. Based on the crystal structure, we generated L1-EN mutants to analyze and manipulate DNA target site recognition. Crystal structures and their dynamic and functional analysis show entire loop grafts to be feasible, resulting in altered specificity, while individual point mutations do not change the nicking pattern of L1-EN. Structural parameters of the DNA target seem more important for recognition than the nucleotide sequence, and nicking profiles on DNA oligonucleotides in vitro are less well defined than the respective integration site consensus in vivo. This suggests that additional factors other than the DNA nicking specificity of L1-EN contribute to the targeted integration of non-LTR retrotransposons. 相似文献
3.
Leslie Edwards Danielle Turner Cody Champion Megha Khandelwal Kailee Zingler Cassidy Stone Ruwini D. Rajapaksha Jing Yang Mahinda I. Ranasinghe Alexander Kornienko Liliya V. Frolova Snezna Rogelj 《Bioorganic & medicinal chemistry letters》2018,28(10):1879-1886
Compounds based on the 2,3-distyrylindole scaffold were found to exhibit bactericidal properties upon irradiation with white light. At the concentration of 1?μM, the lead compound 1 completely (ca. 109?CFU/mL) eradicated such Gram-positive organisms as S. aureus (MRSA, MSSA), E. faecalis (VRE), S. pyogenes and S. mutans when irradiated with white light for 2?min. At the concentration of 5?μM and in the presence of polymyxin E at non-bactericidal 1.25?μg/mL concentration, 1 also showed a 7-log to 9-log reductions in bacterial counts of such Gram-negative organisms as multi-drug resistant (MDR) A. baumannii, MDR P. aeruginosa, E. coli and Klebsiella pneumoniae (CRE: KPC and NDM-1), also when irradiated with white light for 2?min. The structure-activity relationship studies revealed that unsubstituted at benzene rings 2,3-distyrylindole 2 was most potent and gave a 5-order of magnitude eradication of a MRSA strain at the concentration of 30?nM upon irradiation with white light. Initial mechanistic experiments revealed the disruption of bacterial cell membrane, but indicated that singlet oxygen production, which is commonly associated with photodynamic therapy, may not play a role in the bactericidal effects of the 2,3-distyrylindoles. 相似文献
4.
S Eberhardt N Zingler K Kemter G Richter M Cushman A Bacher 《European journal of biochemistry》2001,268(15):4315-4323
Riboflavin synthase of Escherichia coli is a homotrimer of 23.4 kDa subunits catalyzing the formation of the carbocyclic ring of the vitamin, riboflavin, by dismutation of 6,7-dimethyl-8-ribityllumazine. Intramolecular sequence similarity suggested that each subunit folds into two topologically similar domains. In order to test this hypothesis, sequence segments comprising amino-acid residues 1-97 or 101-213 were expressed in recombinant E. coli strains. The recombinant N-terminal domain forms a homodimer that can bind riboflavin, 6,7-dimethyl-8-ribityllumazine and trifluoromethyl-substituted 8-ribityllumazine derivatives as shown by absorbance, circular dichroism, and NMR spectroscopy. Most notably, the recombinant domain dimer displays the same diastereoselectivity for ligands as the full length protein. The minimum N-terminal peptide segment required for ligand binding comprises amino-acid residues 1-87. The recombinant C-terminal domain comprising amino-acid residues 101-213 is relatively unstable and was shown not to bind riboflavin but to differentiate between certain diastereomeric trifluoromethyl-8-ribityllumazine derivatives. The data show that a single domain comprises the intact binding site for one substrate molecule. The enzyme-catalyzed dismutation requires two substrate molecules to be bound in close proximity, and each active site of the enzyme appears to be located at the interface of an N-terminal and C-terminal domain. 相似文献
5.
Intracardiac injection of erythropoietin induces stem cell recruitment and improves cardiac functions in a rat myocardial infarction model 总被引:1,自引:0,他引:1
Christian Klopsch Dario Furlani Ralf Gäbel Wenzhong Li Erik Pittermann Murat Ugurlucan Guenther Kundt Christiana Zingler Ulf Titze Weiwei Wang Lee-Lee Ong Klaus Wagner Ren-Ke Li Nan Ma Gustav Steinhoff 《Journal of cellular and molecular medicine》2009,13(4):664-679
Erythropoietin (EPO) protects the myocardium from ischaemic injury and promotes beneficial remodelling. We assessed the therapeutic efficacy of intracardiac EPO injection and EPO-mediated stem cell homing in a rat myocardial infarction (MI) model. Following MI, EPO (3000 U/kg) or saline was delivered by intracardiac injection. Compared to myocardial infarction control group (MIC), EPO significantly improved left ventricular function ( n = 11–14, P < 0.05) and decreased right ventricular wall stress ( n = 8, P < 0.05) assessed by pressure-volume loops after 6 weeks. MI-EPO hearts exhibited smaller infarction size (20.1 ± 1.1% versus 27.8 ± 1.2%; n = 6–8, P < 0.001) and greater capillary density (338.5 ± 14.7 versus 259.8 ± 9.2 vessels per mm; n = 6–8, P < 0.001) than MIC hearts. Direct EPO injection reduced post-MI myocardial apoptosis by approximately 41% (0.27 ± 0.03% versus 0.42 ± 0.03%; n = 6, P = 0.005). The chemoattractant SDF-1 was up-regulated significantly assessed by quantitative realtime PCR and immunohistology. c-Kit+ and CD34+ stem cells were significantly more numerous in MI-EPO than in MIC at 24 hrs in peripheral blood ( n = 7, P < 0.05) and 48 hrs in the infarcted hearts ( n = 6, P < 0.001). Further, the mRNAs of Akt, eNOS and EPO receptor were significantly enhanced in MI-EPO hearts ( n = 7, P < 0.05). Intracardiac EPO injection restores myocardial functions following MI, which may attribute to the improved early recruitment of c-Kit+ and CD34+ stem cells via the enhanced expression of chemoattractant SDF-1. 相似文献
6.
The group II intron ai5gamma from S. cerevisiae requires high temperature and salt to self-splice in vitro, but it is assisted by the protein Mss116 in vivo. Here we show that Mss116 can stimulate splicing of ai5gamma under near-physiological conditions in vitro, which represents one of the first cases in which a DExH/D protein is shown to act on its natural target. Importantly, we demonstrate that a small subset of DEAD-box proteins can also stimulate ai5gamma splicing in vitro and may represent a distinct subfamily of DEAD-box proteins that functions in RNA tertiary structure assembly. Mutational analysis shows that while ATPase activity is required for stimulation of splicing by Mss116, helicase activity is not. This finding indicates that Mss116 is unlikely to promote intron splicing through the unwinding of kinetic traps. Rather, we propose that Mss116 promotes the ordered assembly of large RNA molecules through stabilization of on-pathway intermediates. 相似文献
7.
Truncation of the cytoplasmic domain of the simian immunodeficiency virus envelope glycoprotein increases env incorporation into particles and fusogenicity and infectivity. 总被引:26,自引:24,他引:2
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Growth of macaque simian immunodeficiency virus (SIVmac) in certain cloned human T-cell lines, such as HUT.78, selects for isolates containing a premature stop codon within the cytoplasmic domain of the transmembrane envelope glycoprotein. In contrast, propagation of virus in macaques or in their cultured T cells favors replication of virus containing the full-length envelope glycoprotein. To elucidate the causes of this phenomenon, we used a human immunodeficiency virus pseudotyping system to assess the effects on infectivity of the cytoplasmic domains of envelope glycoproteins obtained from SIVmac1A11 and SIVmac239. These envelopes contain truncated and full-length cytoplasmic domains, respectively. By analyzing human immunodeficiency virus particles containing selectable genes pseudotyped with each glycoprotein or with chimeric derivatives, we found that truncation of the cytoplasmic domain resulted in a significant advantage in viral entry into HUT.78 T cells and CD4+ U87.MG glial cells. Truncation of the cytoplasmic domain significantly enhanced both envelope density on particles and envelope-mediated cell-to-cell fusion. It is likely that one or both of these effects contribute to the observed differences in infectivity and to the selection of virions with short cytoplasmic tails in human T cells. 相似文献
8.
Wang Q Malherbe L Zhang D Zingler K Glaichenhaus N Killeen N 《Journal of immunology (Baltimore, Md. : 1950)》2001,167(8):4311-4320
A diverse population of MHC class II-restricted CD4 lineage T cells develops in mice that lack expression of the CD4 molecule. In this study, we show that the TCR repertoire selected in the absence of CD4 is distinct, but still overlapping in its properties with that selected in the presence of CD4. Immunization of mice lacking CD4 caused the clonal expansion of T cells that showed less breadth in the range of Ag-binding properties exhibited by their TCRs. Specifically, the CD4-deficient Ag-specific TCR repertoire was depleted of TCRs that demonstrated low-affinity binding to their ligands. The data thus suggest a key role for CD4 in broadening the TCR repertoire by potentiating productive TCR signaling and clonal expansion in response to the engagement of low-affinity antigenic ligands. 相似文献
9.
Kailee Schulz Philip W. Stevens Jeffrey E. Hill Alexis A. Trotter Jared L. Ritch Kyle L. Williams Joshua T. Patterson Quenton M. Tuckett 《Restoration Ecology》2020,28(5):1283-1295
Increasing human populations and urban development have led to losses of estuarine habitats for fish and wildlife. Where resource managers are restoring coastal wetlands, in addition to meeting goals related to hydrologic connectivity, biodiversity, and recreational opportunities, efforts are being made to provide habitat that is suitable for juvenile sportfish. An 18‐month study was conducted to compare juvenile sportfish use of natural, restored, and impacted sites along Tampa Bay, Florida, shorelines. Juvenile sportfish densities at restored sites were broadly comparable to natural sites and greater than at impacted sites. However, site‐specific differences in sportfish use did occur within site types. For example, one restored site had significantly higher densities of red drum Sciaenops ocellatus than any other site, while black drum Pogonias cromis were found exclusively at another restored site. To evaluate whether the restored sites are providing suitable habitat for juvenile fish, we assessed growth (estimated from counts of daily rings on otoliths) and condition (determined by lipid analyses) of juvenile common snook Centropomus undecimalis, an archetypal coastal wetland‐dependent species. Growth (0.43–0.56 mm SL/day) and condition (4.6–6.1% lipid of dry weight) exhibited only site‐specific differences and did not vary among natural, restored, and impacted site types. Although mortality rates of juvenile sportfish were not determined, use of a 40‐m seine found that densities of potential piscine predators in these coastal wetlands were relatively low compared to published studies of open estuarine shorelines. The restoration and creation of coastal wetlands in Tampa Bay provides improved habitat for juvenile sportfish. 相似文献
10.
Mariani Carla Prudente Batista Eliane de Fatima Duarte Michele Delarmelina dos Reis Borba Emilie Zingler João Mangussi-Gomes Beatriz Taynara Araújo dos Santos Olivia Laquis de Moraes Sylvia Asaka Yamashita Hayashida Edmund C. Baracat Francisco de Assis da Rocha Neves Gustavo Arantes Rosa Maciel Tania Aparecida Sartori Sanchez Bachega Gustavo Barcelos Barra Adriana Lofrano-Porto 《Gene》2014