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141.
Ming-Tsair Chan Hsin-Hsiung Chang Shin-Lon Ho Wu-Fu Tong Su-May Yu 《Plant molecular biology》1993,22(3):491-506
We have successfully transferred and expressed a reporter gene driven by an -amylase promoter in a japonica type of rice (Oryza sativa L. cv. Tainung 62) using the Agrobacterium-mediated gene transfer system. Immature rice embryos (10–12 days after anthesis) were infected with an Agrobacterium strain carrying a plasmid containing chimeric genes of -glucuronidase (uidA) and neomycin phosphotransferase (nptII). Co-incubation of potato suspension culture (PSC) with the Agrobacterium inoculum significantly improved the transformation efficiency of rice. The uidA and nptII genes, which are under the control of promoters of a rice -amylase gene (Amy8) and Agrobacterium nopaline synthase gene (nos), respectively, were both expressed in G418-resistant calli and transgenic plants. Integration of foreign genes into the genomes of transgenic plants was confirmed by Southern blot analysis. Histochemical localization of GUS activity in one transgenic plant (R0) revealed that the rice -amylase promoter functions in all cell types of the mature leaves, stems, sheaths and roots, but not in the very young leaves. This transgenic plant grew more slowly and produced less seeds than the wild-type plant, but its R1 and R2 progenies grew normally and produced as much seeds as the wild-type plant. Inheritance of foreign genes to the progenies was also confirmed by Southern blot analysis. These data demonstrate successful gene transfer and sexual inheritance of the chimeric genes. 相似文献
142.
Glucocorticoids regulate the expression of many liver-specific genes via glucocorticoid receptors. The presence of glucocorticoid receptors in liver has been reported in many mammalian species but not in nude mice. In the present study, we demonstrate the presence of specific glucocorticoid receptors in nude mouse liver. The binding of ligands to these receptors could be completely inhibited by RU486, and partially blocked by hydrocortisone and progesterone, whereas estrogen and testosterone had no effect. Hydrocortisone down-regulated the level of glucocorticoid receptors in livers of nude mice and correspondingly enhanced the activities of tyrosine aminotransferase and -glutamyltransferase. Our results indicate that glucocorticoid receptors in nude mouse liver are specific, fully functional, and present at levels 28.5-fold higher than in the liver of normal inbred mice. We suggest that the nude mouse is a valuable model for studies of hepatic glucocorticoid action and may provide a clue to a putative hepatic-thymic interaction. 相似文献
143.
Effect of amino acid changes in the V1/V2 region of the human immunodeficiency virus type 1 gp120 glycoprotein on subunit association, syncytium formation, and recognition by a neutralizing antibody. 总被引:42,自引:40,他引:2 下载免费PDF全文
The contributions of the first and second variable regions of the human immunodeficiency virus type 1 gp120 glycoprotein to envelope glycoprotein structure, function, and recognition by a neutralizing antibody were studied. Several mutants with substitutions in the V2 loop demonstrated complete dissociation of the gp120 and gp41 glycoproteins, suggesting that inappropriate changes in V2 conformation can affect subunit assembly. Some glycoproteins with changes in V1 or V2 were efficiently expressed on the cell surface and were able to bind CD4 but were deficient in syncytium formation and/or virus entry. Recognition of gp120 by the neutralizing monoclonal antibody G3-4 was affected by particular substitutions affecting residues 176 to 184 in the V2 loop. These results suggest that the V1/V2 variable regions of the human immunodeficiency virus type 1 gp120 glycoprotein play a role in postreceptor binding events in the membrane fusion process and can act as a target for neutralizing antibodies. 相似文献
144.
Characterization of conserved human immunodeficiency virus type 1 gp120 neutralization epitopes exposed upon gp120-CD4 binding. 总被引:33,自引:29,他引:4 下载免费PDF全文
M Thali J P Moore C Furman M Charles D D Ho J Robinson J Sodroski 《Journal of virology》1993,67(7):3978-3988
Interaction with the CD4 receptor enhances the exposure on the human immunodeficiency type 1 gp120 exterior envelope glycoprotein of conserved, conformation-dependent epitopes recognized by the 17b and 48d neutralizing monoclonal antibodies. The 17b and 48d antibodies compete with anti-CD4 binding antibodies such as 15e or 21h, which recognize discontinuous gp120 sequences near the CD4 binding region. To characterize the 17b and 48d epitopes, a panel of human immunodeficiency virus type 1 gp120 mutants was tested for recognition by these antibodies in the absence or presence of soluble CD4. Single amino acid changes in five discontinuous, conserved, and generally hydrophobic regions of the gp120 glycoprotein resulted in decreased recognition and neutralization by the 17b and 48d antibodies. Some of these regions overlap those previously shown to be important for binding of the 15e and 21h antibodies or for CD4 binding. These results suggest that discontinuous, conserved epitopes proximal to the binding sites for both CD4 and anti-CD4 binding antibodies become better exposed upon CD4 binding and can serve as targets for neutralizing antibodies. 相似文献
145.
Antibodies to discontinuous or conformationally sensitive epitopes on the gp120 glycoprotein of human immunodeficiency virus type 1 are highly prevalent in sera of infected humans. 总被引:45,自引:35,他引:10 下载免费PDF全文
We have used an indirect-capture enzyme-linked immunosorbent assay to quantitate the reactivity of sera from human immunodeficiency virus type 1 (HIV-1)-infected humans with native recombinant gp120 (HIV-1 IIIB or SF-2) or with the gp120 molecule (IIIB or SF-2) denatured by being boiled in the presence of dithiothreitol with or without sodium dodecyl sulfate. Denaturation of IIIB gp120 reduced the titers of sera from randomly selected donors by at least 100-fold, suggesting that the majority of cross-reactive anti-gp120 antibodies present are directed against discontinuous or otherwise conformationally sensitive epitopes. When SF-2 gp120 was used, four of eight serum samples reacted significantly with the denatured protein, albeit with ca. 3- to 50-fold reductions in titer. Only those sera reacting with denatured SF-2 gp120 bound significantly to solid-phase-adsorbed SF-2 V3 loop peptide, and none bound to IIIB V3 loop peptide. Almost all antibody binding to reduced SF-2 gp120 was blocked by preincubation with the SF-2 V3 loop peptide, as was about 50% of the binding to native SF-2 gp120. When sera from a laboratory worker or a chimpanzee infected with IIIB were tested, the pattern of reactivity was reversed, i.e., there was significant binding to reduced IIIB gp120, but not to reduced SF-2 gp120. Binding of these sera to reduced IIIB gp120 was 1 to 10% that to native IIIB gp120 and was substantially decreased by preincubation with IIIB (but not SF-2) V3 loop peptide. To analyze which discontinuous or conformational epitopes were predominant in HIV-1-positive sera, we prebound monoclonal antibodies (MAbs) to IIIB gp120 and then added alkaline phosphatase-labelled HIV-1-positive sera. MAbs (such as 15e) that recognize discontinuous epitopes and compete directly with CD4 reduced HIV-1-positive sera binding by about 50%, whereas neutralizing MAbs to the C4, V2, and V3 domains of gp120 were either not inhibitory or only weakly so. Thus, antibodies to the discontinuous CD4-binding site on gp120 are prevalent in HIV-1-positive sera, antibodies to linear epitopes are less common, most of the antibodies to linear epitopes are directed against the V3 region, and most cross-reactive antibodies are directed against discontinuous epitopes, including regions involved in CD4 binding. 相似文献
146.
Employing a photoaffinity labeling procedure with 8-azido-S-adenosyl-l-[methyl-3H]methionine (8-N3-Ado[methyl-3H]Met), the binding sites for S-adenosyl-l-methionine (AdoMet) of three protein N-methyltransferases [AdoMet:myelin basic protein-arginine N-methyltransferase (EC2.1.1.23); AdoMet:histone-arginin N-methyltransferase (EC2.1.1.23); and AdoMet:cytochromec-lysine N-methyltransferase (EC2.1.1.59)] have been investigated. The incorporation of the photoaffinity label into the enzymes upon UV irradiation was highly specific. In order to define the AdoMet binding sites, the photolabeled enzymes were sequentially digested with trypsin, chymotrypsin, and endoproteinase Glu-C. After each proteolytic digestion, radiolabeled peptide from each enzyme was resolved on HPLC first by gradient elution and further purified by an isocratic elution. Retention times of the purified radiolabeled peptides from the three enzymes from the corresponding proteolysis were significantly different, indicating that their sizes and compositions were different. Amino acid composition analysis of these peptides confirmed further that the AdoMet binding sites of these protein N-methyltransferases are quite different. 相似文献
147.
Zhi Jian Yu Changbae Jin Robin William Rockhold Beth Hoskins Ing Kang Ho 《Neurochemical research》1993,18(11):1203-1209
Wistar-Kyoto and spontaneously hypertensive rats received i.v. infusions of cocaine hydrochloride (60 mg/kg per day) for 3, 7, and 14 days, or saline for 7 days. Acute cocaine challenge (40 mg/kg, s.c.) was given to treated and control rats 24 hr after the termination of each infusion period. There were no strain differences in brain levels of cocaine during cocaine infusion, nor after cocaine challenges. There were no strain differences in resting levels of [3H]dopamine release. Release of [3H]dopamine decreased in nuclei accumbens of 7- and 14-day cocaine-infused animals. Release of [3H]dopamine was maximal in both brain regions 2 hr after acute cocaine challenge. After 14 days of cocaine infusion, cocaine challenge in both strains reduced [3H]dopamine release in the nucleus accumbens, but not in the striatum; the reduction being greater in Wistar-Kyoto rats. The behavioral tolerance which accompanies similar cocaine infusion regimens may be related to striatal tolerance to cocaine-induced dopamine release. 相似文献
148.
Jong An Lee Jung Ho Ahn Gi Bae Kim Sol Choi Ji Yeon Kim Sang Yup Lee 《Biotechnology and bioengineering》2023,120(1):203-215
Microbial production of various TCA intermediates and related chemicals through the reductive TCA cycle has been of great interest. However, rumen bacteria that naturally possess strong reductive TCA cycle have been rarely studied to produce these chemicals, except for succinic acid, due to their dependence on fumarate reduction to transport electrons for ATP synthesis. In this study, malic acid (MA), a dicarboxylic acid of industrial importance, was selected as a target chemical for mass production using Mannheimia succiniciproducens, a rumen bacterium possessing a strong reductive branch of the TCA cycle. The metabolic pathway was reconstructed by eliminating fumarase to prevent MA conversion to fumarate. The respiration system of M. succiniciproducens was reconstructed by introducing the Actinobacillus succinogenes dimethylsulfoxide (DMSO) reductase to improve cell growth using DMSO as an electron acceptor. Also, the cell membrane was engineered by employing Pseudomonas aeruginosa cis-trans isomerase to enhance MA tolerance. High inoculum fed-batch fermentation of the final engineered strain produced 61 g/L of MA with an overall productivity of 2.27 g/L/h, which is the highest MA productivity reported to date. The systems metabolic engineering strategies reported in this study will be useful for developing anaerobic bioprocesses for the production of various industrially important chemicals. 相似文献
149.
AxenicTrentepohlia odorata was cultured at three different NH4Cl levels (3.5 × 10–2, 3.5 × 10–3, 3.5 × 10–4 M) and three different light intensities (48, 76, 122 µmol m–2 s–1). Chloride had no effect on growth over this range of concentration. High light intensity and high NH4Cl concentration enhanced the specific growth rate. The carotenoid content increased under a combination of high light intensity and low N concentration. WhenD. bardawil was exposed to the same combination of growth conditions, there was an increase in its carotenoid content. The light saturation and the light inhibition constants (K
s andK
i, respectively) for growth, and the saturation constant (K
m) for NH4Cl were determined. TheK
s andK
i values were higher inT. odorata (66.7 and> 122 mol m–2 s–1, respectively) than inD. bardawil (5.1 and 14.7 µmol m–2 s–1, respectively). TheK
m value determined at 122 µmol m–2 s–1, however, was lower inT. odorata (0.048 µM) than inD. bardawil (0.062 µM).Author for correspondence 相似文献
150.
Bacteriophage P22 and λ are related bacteriophages with similar gene organizations. In λ the cII-dependent PI promoter is responsible for λint gene expression. The only apparent counterpart to PI in P22 is oriented in the opposite direction, and cannot transcribe the P22 int gene. We show that this promoter, called Pal, is active both in vivo and in vitro, and is dependent upon the P22 cII-like gene, called c1. We have also determined the DNA sequence of a 3.3 kb segment that closes the gap between previously reported sequences to give a continuous sequence between the P22 pL promoter and the int gene. The newly determined sequence is densely packed with genes from the pL direction, and the proteins predicted by the sequence show excellent correlation with the proteins mapped by Youderian and Susskind in 1980. However, the sequence contains no apparent genes in the opposite (pal) direction, and no additional binding motifs for the P22 c1 protein. We conclude that int gene expression in P22 is regulated by a different mechanism than in λ. 相似文献