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541.
We report the preparation of the 7alpha-cyano derivative of the isomeric (17alpha,20E/Z)-[125I]iodovinyl-19-nortestosterones (IVNT) together with their binding affinity for the androgen receptor (AR) and their biodistribution in two different animal models. The cyano group was introduced at the 7alpha-position by hydrocyanation of 4,6-estradien-17beta-ol-3-one with diethylaluminum cyanide. Selective protection of the A-ring enone system as the dienol ether followed by ethynylation and deprotection under base and acid hydrolysis condition gave 7alpha-cyano-17alpha-ethynyl-19-nortestosterone. The stannyl derivatives were prepared by addition of tri-n-butylstannyl hydride and converted stereospecifically to the corresponding [125I]iodovinyl analog using [125I]NaI and H2O2. The [125I]iodovinylsteroids were intravenously administered to male rats and estrogen-primed immature female rats and tissue uptake was measured up to 6h post-injection. Co-administration of NLP-004 or ORG-2058, highly selective ligands for the progesterone receptor, to the female rats did not affect uterus uptake of the 125I-ligands. However co-injection of testosterone to DES-primed male rats induced a marked increase in prostate uptake of the 20Z-isomer of 7alpha-cyano-[125I]-IVNT. The relative binding affinity (RBA) of either 7alpha-cyano-(17alpha,20E/Z)-IVNT isomer for the AR is low (RBA=4 and 3, respectively, versus 100 for 5alpha-dihydrotestosterone (DHT)), suggesting the absence of a possible role of the AR in the localization process. These findings contrast previously reported data for the analogous 7alpha-methyl-[125I]-IVNT where co-administration of testosterone was shown to result in a 50% drop in prostate uptake. These data indicate that the addition of an electron withdrawing 7alpha-cyano group to 123I-labeled nortestosterone derivatives does not improve their potential to serve as SPECT agents for the imaging of AR densities in the prostate. 相似文献
542.
543.
Sequence identification and characterization of human carnosinase and a closely related non-specific dipeptidase 总被引:6,自引:0,他引:6
Teufel M Saudek V Ledig JP Bernhardt A Boularand S Carreau A Cairns NJ Carter C Cowley DJ Duverger D Ganzhorn AJ Guenet C Heintzelmann B Laucher V Sauvage C Smirnova T 《The Journal of biological chemistry》2003,278(8):6521-6531
Carnosine (beta-alanyl-L-histidine) and homocarnosine (gamma-aminobutyric acid-L-histidine) are two naturally occurring dipeptides with potential neuroprotective and neurotransmitter functions in the brain. Peptidase activities degrading both carnosine and homocarnosine have been described previously, but the genes linked to these activities were unknown. Here we present the identification of two novel cDNAs named CN1 and CN2 coding for two proteins of 56.8 and 52.7 kDa and their classification as members of the M20 metalloprotease family. Whereas human CN1 mRNA and protein are brain-specific, CN2 codes for a ubiquitous protein. In contrast, expression of the mouse and rat CN1 orthologues was detectable only in kidney. The recombinant CN1 and CN2 proteins were expressed in Chinese hamster ovary cells and purified to homogeneity. CN1 was identified as a homodimeric dipeptidase with a narrow substrate specificity for Xaa-His dipeptides including those with Xaa = beta Ala (carnosine, K(m) 1.2 mM), N-methyl beta Ala, Ala, Gly, and gamma-aminobutyric acid (homocarnosine, K(m) 200 microM), an isoelectric point of pH 4.5, and maximal activity at pH 8.5. CN2 protein is a dipeptidase not limited to Xaa-His dipeptides, requires Mn(2+) for full activity, and is sensitive to inhibition by bestatin (IC(50) 7 nM). This enzyme does not degrade homocarnosine and hydrolyzes carnosine only at alkaline pH with an optimum at pH 9.5. Based on their substrate specificity and biophysical and biochemical properties CN1 was identified as human carnosinase (EC ), whereas CN2 corresponds to the cytosolic nonspecific dipeptidase (EC ). 相似文献
544.
Access of antibody molecules to the conserved coreceptor binding site on glycoprotein gp120 is sterically restricted on primary human immunodeficiency virus type 1 总被引:1,自引:0,他引:1 下载免费PDF全文
Labrijn AF Poignard P Raja A Zwick MB Delgado K Franti M Binley J Vivona V Grundner C Huang CC Venturi M Petropoulos CJ Wrin T Dimitrov DS Robinson J Kwong PD Wyatt RT Sodroski J Burton DR 《Journal of virology》2003,77(19):10557-10565
Anti-human immunodeficiency virus type 1 (HIV-1) antibodies whose binding to gp120 is enhanced by CD4 binding (CD4i antibodies) are generally considered nonneutralizing for primary HIV-1 isolates. However, a novel CD4i-specific Fab fragment, X5, has recently been found to neutralize a wide range of primary isolates. To investigate the precise nature of the extraordinary neutralizing ability of Fab X5, we evaluated the abilities of different forms (immunoglobulin G [IgG], Fab, and single-chain Fv) of X5 and other CD4i monoclonal antibodies to neutralize a range of primary HIV-1 isolates. Our results show that, for a number of isolates, the size of the neutralizing agent is inversely correlated with its ability to neutralize. Thus, the poor ability of CD4i-specific antibodies to neutralize primary isolates is due, at least in part, to steric factors that limit antibody access to the gp120 epitopes. Studies of temperature-regulated neutralization or fusion-arrested intermediates suggest that the steric effects are important in limiting the binding of IgG to the viral envelope glycoproteins after HIV-1 has engaged CD4 on the target cell membrane. The results identify hurdles in using CD4i epitopes as targets for antibody-mediated neutralization in vaccine design but also indicate that the CD4i regions could be efficiently targeted by small molecule entry inhibitors. 相似文献
545.
Philippe C Morand Veronique Veuillez Claire Poyart Eric Abachin Gilles Quesne Bertrand Dupont Patrick Berche Jean-Paul Viard 《Annals of clinical microbiology and antimicrobials》2003,2(1):1-4
Background
Hepatitis B virus infection although preventable by vaccination remains an important health issue throughout the world due to its morbidity, mortality and economical losses. Early seroprotection is desirable for people at high risk of exposure. The aim of this study was to determine whether three-week hepatitis B vaccination (on days 0, 10 and 21) provide seroprotection or not.Methods
The 120 subjects enrolled into the study were divided into two groups and vaccinated by the classic (months 0, 1, and 2) or the accelerated (days 0, 10, and 21) schedules and antibody response determined on days 30, 60, and 90 and, if below 10 mIU/ml-1, again on day 180. For each individual in the classic group (B) three subjects were enrolled in the accelerated group (A). Recombinant hepatitis B vaccine (Gen-Hevac B, Pasteur) was given as 20 micrograms intramuscular injections via the deltoid muscle. A booster dose on day 365 was administered for each group. Family members of hepatitis B carriers and volunteer health personnel were enrolled into group A. To the B group only volunteers who wanted vaccination against hepatitis B were included.Results
After three doses of vaccine, Anti-HBs titers reached protective levels in both groups. The number of vaccinees with seroprotective levels of Anti-HBs (≥10 mIU/ml-1) on day 30 was 53 (58.9%) in group A and 9 (30.0%) in group B (p < 0.05). On day 60, there was no difference between group A and B, with response rates of 84.4% (n = 76) and 80.0% (n = 24) respectively (p > 0.05). On day 90 there was no difference between group B and group A; with 26 (86.7%) and 79 (87.7%) responders respectively. In both groups those with Anti-HBs levels <10 mIU/ml-1 attained protective levels by day 180.Conclusion
In this study, the three-week vaccination provided protective antibody titers within a shorter time compared to the classic schedule. Therefore, in order to provide rapid antibody production against hepatitis B virus, the accelerated vaccination schedule seems to be a good preference. 相似文献546.
Fission yeast Sap1 protein is essential for chromosome stability 总被引:2,自引:0,他引:2
Sap1 is a dimeric sequence-specific DNA binding-protein, initially identified for its role in mating-type switching of the fission yeast Schizosaccharomyces pombe. The protein is relatively abundant, around 10,000 dimers/cell, and is localized in the nucleus. sap1+ is essential for viability, and transient overexpression is accompanied by rapid cell death, without an apparent checkpoint response and independently of mating-type switching. Time lapse video microscopy of living cells revealed that the loss of viability is accompanied by abnormal mitosis and chromosome fragmentation. Overexpression of the C terminus of Sap1 induces minichromosome loss associated with the “cut” phenotype (uncoupling mitosis and cytokinesis). These phenotypes are favored when the C terminus of Sap1 is overexpressed during DNA replication. Fluorescence in situ hybridization experiments demonstrated that the cut phenotype is related to precocious centromere separation, a typical marker for loss of cohesion. We propose that Sap1 is an architectural chromatin-associated protein, required for chromosome organization. 相似文献
547.
Stimulation of neutrophils with chemotactic peptide induces actin reorganization, formation of actin-rich protrusions, and development of polarity. Shape changes and actin polymerization can also be induced by phorbol ester-mediated direct activation of protein kinase C (PKC). We have investigated the role of cholesterol in stimulus-dependent motile events and in activation of signaling pathways in neutrophil-like differentiated HL-60 cells. Depletion of plasma membrane cholesterol using methyl-beta-cyclodextrin (MbetaCD) prevented chemotactic peptide and phorbol ester-induced shape changes and increases in cytoskeletal actin. Cholesterol depletion almost completely suppressed chemotactic peptide-mediated activation of p42/44 mitogen-activated protein kinase (MAPK). Phosphorylation of protein kinase B on Thr-308, which is indicative of activation of phosphatidylinositol 3-kinase, was in contrast only partially inhibited. Stimulus-mediated membrane recruitment of different PKC isoforms was differentially affected by treatment of cells with MbetaCD. Membrane recruitment of PKCalpha induced by chemotactic peptide or phorbol ester was suppressed, whereas that of PKCbetaII was only partially affected. Membrane association of PKCdelta was almost insensitive to cholesterol depletion. In summary, our results implicate an important role of cholesterol-containing lipid microdomains (rafts) especially in chemotactic peptide-induced activation of MAPK pathways and in chemotactic peptide- and phorbol ester-mediated activation of PKCalpha. 相似文献
548.
Gutierrez MC Brisse S Brosch R Fabre M Omaïs B Marmiesse M Supply P Vincent V 《PLoS pathogens》2005,1(1):e5
The highly successful human pathogen Mycobacterium tuberculosis has an extremely low level of genetic variation, which suggests that the entire population resulted from clonal expansion following an evolutionary bottleneck around 35,000 y ago. Here, we show that this population constitutes just the visible tip of a much broader progenitor species, whose extant representatives are human isolates of tubercle bacilli from East Africa. In these isolates, we detected incongruence among gene phylogenies as well as mosaic gene sequences, whose individual elements are retrieved in classical M. tuberculosis. Therefore, despite its apparent homogeneity, the M. tuberculosis genome appears to be a composite assembly resulting from horizontal gene transfer events predating clonal expansion. The amount of synonymous nucleotide variation in housekeeping genes suggests that tubercle bacilli were contemporaneous with early hominids in East Africa, and have thus been coevolving with their human host much longer than previously thought. These results open novel perspectives for unraveling the molecular bases of M. tuberculosis evolutionary success. 相似文献
549.
Delivery of short hairpin RNA sequences by using a replication-competent avian retroviral vector 下载免费PDF全文
Bromberg-White JL Webb CP Patacsil VS Miranti CK Williams BO Holmen SL 《Journal of virology》2004,78(9):4914-4916
While recent studies have demonstrated that retroviral vectors can be used to stably express short hairpin RNA (shRNA) to inhibit gene expression, these studies have utilized replication-defective retroviruses. We describe the creation of a replication-competent, Gateway-compatible retroviral vector capable of expressing shRNA that inhibits the expression of specific genes. 相似文献
550.
Anderson JV Delseny M Fregene MA Jorge V Mba C Lopez C Restrepo S Soto M Piegu B Verdier V Cooke R Tohme J Horvath DP 《Plant molecular biology》2004,56(4):527-539
Cassava (Manihot esculenta) is a major food staple for nearly 600 million people in Africa, Asia, and Latin America. Major losses in yield result from biotic and abiotic stresses that include diseases such as Cassava Mosaic Disease (CMD) and Cassava Bacterial Blight (CBB), drought, and acid soils. Additional losses also occur from deterioration during the post-harvest storage of roots. To help cassava breeders overcome these obstacles, the scientific community has turned to modern genomics approaches to identify key genetic characteristics associated with resistance to these yield-limiting factors. One approach for developing a genomics program requires the development of ESTs (expressed sequence tags). To date, nearly 23000 ESTs have been developed from various cassava tissues, and genotypes. Preliminary analysis indicates existing EST resources contain at least 6000–7000 unigenes. Data presented in this report indicate that the cassava ESTs will be a valuable resource for the study of genetic diversity, stress resistance, and growth and development, not only in cassava, but also other members of the Euphorbiaceae family. 相似文献