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121.
Nucleotide sequence and expression of the Pseudomonas aeruginosa algF gene controlling acetylation of alginate 总被引:9,自引:0,他引:9
Dean Shinabarger † Thomas B. May Aoife Boyd Mita Ghosh A. M. Chakrabarty 《Molecular microbiology》1993,9(5):1027-1035
Colonization of the cystic fibrosis lung by Pseudomonas aeruginosa is greatly facilitated by the production of an exopolysaccharide called alginate. In this study we determined the nucleotide sequence of an alginate modification gene, algF, which controls the addition of acetyl groups to alginate. Expression of algF using a T7 promoter-expression system showed that algF codes for a 24.5 kDa polypeptide (predicted size 22 832 Da) that is processed to 19.5 kDa. The N-terminus of the processed polypeptide matched the predicted amino acid sequence of AlgF starting at Asp-29. An algF mutant failed to produce alginate owing to a polar effect on the downstream algA gene. Although the algA gene, provided in trans, restored synthesis of alginate, the alginate was non-acetylated. We show that a plasmid containing both the algF and algA gene complements the alginate acetylation defect of the algF mutant strain. 相似文献
122.
Alginic acid synthesis in Pseudomonas aeruginosa mutants defective in carbohydrate metabolism. 总被引:16,自引:11,他引:5 下载免费PDF全文
Mutant cells of mucoid Pseudomonas aeruginosa isolated from cystic fibrosis patients were examined for their ability to synthesize alginic acid in resting cell suspensions. Unlike the wild-type strain which synthesizes alginic acid from glycerol, fructose, mannitol, glucose, gluconate, glutamate, or succinate, mutants lacking specific enzymes of carbohydrate metabolism are uniquely impaired. A phosphoglucose isomerase mutant did not synthesize the polysaccharide from mannitol, nor did a glucose 6-phosphate dehydrogenase mutant synthesize the polysaccharide from mannitol or glucose. Mutants lacking the Entner-Doudoroff pathway dehydrase or aldolase failed to produce alginate from mannitol, glucose, or gluconate, as a 3-phosphoglycerate kinase or glyceraldehyde 3-phosphate dehydrogenase mutant failed to produce from glutamate or succinate. These results demonstrate the primary role of the Entner-Doudoroff pathway enzymes in the synthesis of alginate from glucose, mannitol, or gluconate and the role of glyceraldehyde 3-phosphate dehydrogenase reaction for the synthesis from gluconeogenic precursors such as glutamate. The virtual absence of any activity of phosphomannose isomerase in cell extracts of several independent mucoid bacteria and the impairment of alginate synthesis from mannitol in mutants lacking phosphoglucose isomerase or glucose 6-phosphate dehydrogenase rule out free mannose 6-phosphate as an intermediate in alginate biosynthesis. 相似文献
123.
The mutagenic activities of five 1-alkyl derivatives of 3-nitro-l-nitrosoguanidine,viz. methyl, ethyl, propyl, isopentyl and octyl, were tested onStreptomyces indicus. All the compounds were found to be mutagenic. The methyl and ethyl derivatives were most effective regarding mutational frequency. The effectiveness decreased with increasing number of C-atoms in the alkyl group of the mutagens. The compounds tested had a much higher mutagenic efficiency than UV-rays. A marked variation existed in antibiotic activity among the biochemical mutants: increase, decrease or complete absence of such activity were seen. 相似文献
124.
125.
Soluble starch was hydrolysed to maltose byAspergillus wentii Wehmer (IMI 17295). Studies on nutritional requirements ofAspergillus wentii for production of amylase revealed that the optimum conditions were achieved in fermentation culture medium containing 1%
starch, and incubated at 20 °C for 3 days at pH 6.0. Tryptophan was the best nitrogen source. The amylase activity was completely
inhibited when 1 mm sodium iodoacetate was incorporated into the medium. With 10 mm sodium citrate the amylase activity was
increased from 3.51 to 6.0 mg/ml. 相似文献
126.
Sunil Chakrabarty Anuradha Nandi Chinmay K. Mukhopadhyay Indu B. Chatterjee 《Molecular and cellular biochemistry》1992,111(1-2):41-47
Ascorbic acid (AH2) is a potential scavenger of superoxide radical and singlet oxygen. In the guinea pig, marginal AH2 deficiency results in intracellular oxidative damage in the cardiac tissue as evidenced by lipid peroxidation, formation of fluorescent pigment and loss of structural integrity of the microsomal membranes. The oxidative damage does not occur due to lack of enzymatic scavengers of reactive oxygen species such as superoxide dismutase, catalase and glutathione peroxidase. Also, glutathione transferase activity is not decreased in AH2 deficiency. Lipid peroxidation, fluorescent pigment formation and protein modification disappear after AH2 therapy. These results, if extra-polated to human beings, would indicate that chronic subclinical AH2 deficiency may result in progressive oxidative damage which in the long run may lead to permanent degenerative diseases in the heart. 相似文献
127.
Acinetobacter sp. strain P6 has previously been reported to utilize biphenyl (BP) and chlorinated BPs, with accumulation of corresponding chlorobenzoic acids. Arthrobacter sp. strain M5 was isolated as a contaminant in the culture of Acinetobacter sp. strain P6 growing on 4-chlorobiphenyl and showed properties similar to P6 in the degradation of chlorinated BPs. Both strains harbored an identical plasmid of 53.7 megadaltons. These strains spontaneously lost the ability to utilize BP and 4-chlorobiphenyl with high frequency (4 to 8%) after overnight growth in nutrient broth. The BP- derivatives could not regain the BP-assimilating ability (reversion frequency, less than 10(-9) per cell per generation) but retained the plasmid with small, detectable deletions. BP+ P6 cells grown on BP or benzoate oxidized BP and 2,3-dihydroxybiphenyl and produced meta cleavage compounds from the latter compound (lambda max, 434 nm) and also from catechol (lambda max, 375 nm) through the meta pathway. On the other hand, benzoate-grown BP- segregants totally lost the BP-metabolizing activities and oxidized catechol through the ortho pathway. A combined culture of the chlorinated BP-dissimilating P6 or M5 strain (harboring the putative 53.7-megadalton plasmid specifying conversion of chlorobiphenyls to chlorobenzoic acids) and genetically constructed mono- or dichlorobenzoate-utilizing pseudomonads (harboring plasmids encoding complete utilization of mono- or dichlorobenzoates) allowed greater than 98% utilization of mono- and dichlorobiphenyls, with the liberation of equivalent amounts of chloride ions. 相似文献
128.
129.
Cloning, physical mapping and expression of chromosomal genes specifying degradation of the herbicide 2,4,5-T by Pseudomonas cepacia AC1100 总被引:14,自引:0,他引:14
A genomic library of total DNA of Pseudomonas cepacia AC1100 was constructed on a broad-host-range cosmid vector pCP13 in Escherichia coli AC80. A 25-kb segment was isolated from the library which complemented a Tn5-generated, 2,4,5-trichlorophenoxyacetic acid-negative (2,4,5-T-) mutant, P. cepacia PT88. This mutation was partially characterized and appeared to be lacking functional enzyme required for metabolism of an intermediate of the 2,4,5-T pathway, recently identified as 5-chloro-1,2,4-trihydroxybenzene [Chapman et al., Abstr. Soc. Environ. Toxicol. Chem. USA 8 (1987) 127]. A simple colorimetric assay was developed to detect the presence of this active enzyme in intact cells and was used to determine the expression of complementing genes. Subcloning experiments showed that a 4-kb BamHI-PstI fragment and a 290-bp PstI-EcoRI fragment, separated by 1.3-kb, were required for complementation. Both fragments are identified to be chromosomal in origin. Hybridization studies using the subcloned fragments revealed that in addition to a Tn5 insertion, mutant PT88 contained an extensive chromosomal deletion accounting for its 2,4,5-T- phenotype. The cloned fragments did not show homology to plasmid DNAs carrying degradative genes for toluene, naphthalene and 3-chlorobenzoate. 相似文献
130.
Subhendu Chakrabarty Debasish Nag Arnab Ganguli Amlan Das Debabrata Ghosh Dastidar Gopal Chakrabarti 《Journal of cellular biochemistry》2019,120(4):5987-6003
Theaflavin (TF) and epigallocatechin-3-gallate (EGCG) both have been reported previously as microtubule depolymerizing agents that also have anticancer effects on various cancer cell lines and in animal models. Here, we have applied TF and EGCG in combination on HeLa cells to investigate if they can potentiate each other to improve their anticancer effect in lower doses and the underlying mechanism. We found that TF and EGCG acted synergistically, in lower doses, to inhibit the growth of HeLa cells. We found the combination of 50 µg/mL TF and 20 µg/mL EGCG to be the most effective combination with a combination index of 0.28. The same combination caused larger accumulation of cells in the G 2/M phase of the cell cycle, potent mitochondrial membrane potential loss, and synergistic augmentation of apoptosis. We have shown that synergistic activity might be due to stronger microtubule depolymerization by simultaneous binding of TF and EGCG at different sites on tubulin: TF binds at vinblastine binding site on tubulin, and EGCG binds near colchicines binding site on tubulin. A detailed mechanistic analysis revealed that stronger microtubule depolymerization caused effective downregulation of PI3K/Akt signaling and potently induced mitochondrial apoptotic signals, which ultimately resulted in the apoptotic death of HeLa cells in a synergistic manner. 相似文献