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1.
Cecropins are a family of small, basic antibacterial polypeptides which can be isolated from pupae of immunized Lepidoptera. They are active against both gram-negative and gram-positive bacteria. We studied a mutant of Escherichia coli, strain SB1004, which is more sensitive to cecropin D than is the parental strain. The mutant was selected as resistant to a host range mutant of a Serratia marcescens phage. When the protein composition of the outer membrane was examined, strain SB1004 and some other phage-resistant mutants were found to be deficient in the OmpC protein. It was concluded that the OmpC protein is the receptor of the phage. Strain SB1004 was found to differ from other ompC mutants in being especially sensitive to hydrophobic antibiotics and to cecropin D. Furthermore, strain SB1004 has a tendency for spontaneous autolysis. A genetic analysis showed the mutations in strain SB1004 and a suppressor mutant to map in the ompC region. The activity of cecropin D against different strains of E. coli was specifically enhanced when divalent cations were absent. No such effect was found with cecropins A and B, which are less hydrophobic than the D form. 相似文献
2.
H Shirae K Kobayashi H Shiragami Y Irie N Yasuda K Yokozeki 《Applied and environmental microbiology》1989,55(2):419-424
A novel microbial method for the production of 2',3'-dideoxynucleosides by transdideoxyribosylation has been developed. By screening microorganisms producing 2',3'-dideoxyadenosine (DDA) from 2',3'-dideoxyuridine (DDU) and adenine, Escherichia coli AJ 2595 was selected as the best producer. Optimal pH and temperature for the DDA-producing reaction were ca. 6.5 and 50 degrees C, respectively. Pi seemed to be an essential factor for the reaction, and its optimal concentration was ca. 25 mM. Moreover, polyethylene glycol had a notable effect on DDA production. Under the best conditions established, 52 mM DDA was obtained from 100 mM DDU and 100 mM adenine after 48 h of incubation from resting cells of E. coli AJ 2595. This strain could also produce 2',3'-dideoxynucleosides, such as 2',3'-dideoxyinosine (DDI), 2',3'-dideoxyguanosine, and 2',3'-dideoxythymidine, from DDU and the corresponding bases. In particular, this strain could produce DDI in high yield (ca. 32 mM from 100 mM DDU and 100 mM hypoxanthine) after 24 h of incubation. However, 2',3'-dideoxycytidine was not produced from DDU and cytosine by resting cells of E. coli AJ 2595. 相似文献
3.
Production of 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine from 2',3'-dideoxyuridine and the corresponding purine bases by resting cells of Escherichia coli AJ 2595.
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H Shirae K Kobayashi H Shiragami Y Irie N Yasuda K Yokozeki 《Applied microbiology》1989,55(2):419-424
A novel microbial method for the production of 2',3'-dideoxynucleosides by transdideoxyribosylation has been developed. By screening microorganisms producing 2',3'-dideoxyadenosine (DDA) from 2',3'-dideoxyuridine (DDU) and adenine, Escherichia coli AJ 2595 was selected as the best producer. Optimal pH and temperature for the DDA-producing reaction were ca. 6.5 and 50 degrees C, respectively. Pi seemed to be an essential factor for the reaction, and its optimal concentration was ca. 25 mM. Moreover, polyethylene glycol had a notable effect on DDA production. Under the best conditions established, 52 mM DDA was obtained from 100 mM DDU and 100 mM adenine after 48 h of incubation from resting cells of E. coli AJ 2595. This strain could also produce 2',3'-dideoxynucleosides, such as 2',3'-dideoxyinosine (DDI), 2',3'-dideoxyguanosine, and 2',3'-dideoxythymidine, from DDU and the corresponding bases. In particular, this strain could produce DDI in high yield (ca. 32 mM from 100 mM DDU and 100 mM hypoxanthine) after 24 h of incubation. However, 2',3'-dideoxycytidine was not produced from DDU and cytosine by resting cells of E. coli AJ 2595. 相似文献
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6.
J. C. Cox 《Archives of microbiology》1989,152(4):397-400
Four classes of Escherichia coli mutants deficient in either or both of their anaerobic selenium-containing formate dehydrogenases (FDH) were isolated. A class I mutant devoid of FDHH activity specifically linked to benzyl viologen (BV) produced a small amount of the FDHH 80,000 dalton selenopeptide. Three class II mutants were deficient in FDHN activity specifically linked to phenazine methosulfate (PMS) and exhibited a selenopeptide doublet rather than the FDHN 110,000 dalton selenosubunit. Three class III mutants were selenium incorporation deficient and did not exhibit either FDH activity or 75Selabeled selenopolymers. A class IV mutant was devoid of PMS-linked FDHN activity; neither its FDHN 110,000 dalton selenosubunit nor its BV-linked FDHH activity was fully regulated by nitrate.Abbreviations FDH
formate dehydrogenase
- BV
benzyl viologen
- MV
methyl viologen
- PMS
phenazine methosulfate
- SDS-PAGE
sodium dodecyl sulfate-polyacrylamide gel electrophoresis 相似文献
7.
The effects of forskolin on the sensitivity of adenylate cyclase to 'P'-site-mediated inhibition were studied. Stimulation of crude and purified preparations of adenylate cyclase by forskolin led to decreased sensitivity to inhibition by 2',5' dideoxyadenosine with enzyme from rat and bovine brain. This is in contrast with the enhancement of P-site sensitivity induced by calmodulin, divalent cations, and stable GTP analogs and is in contrast with behavior seen with enzyme from liver and S49 cyc membranes. The effect of forskolin on P-site sensitivity of the brain adenylate cyclase was not dependent on the presence of G-proteins or calmodulin. It was not the consequence of proteolysis nor was it due to an obvious artifact in the assay procedures. This distinct behavior of the brain enzyme is most likely due to a structural difference in the catalytic subunit. 相似文献
8.
Takamatsu S Katayama S Naito M Yamashita K Ineyama T Izawa K 《Nucleosides, nucleotides & nucleic acids》2003,22(5-8):711-713
A facile method for the synthesis of 3'-alpha-fluoro-2',3'-dideoxyadenosine (5) has been developed using a novel rearrangement of 3'-beta-bromine to the 2'-beta position during 3'-alpha fluorination. 相似文献
9.
Katsumi Isono A. Garth Cumberlidge Setsuko Isono Yukinori Hirota 《Molecular & general genetics : MGG》1977,152(3):239-243
Summary Various alterations in ribosomal proteins were detected in forty-one mutants ofE. coli isolated as temperature-sensitive mutants. Out of these, six are new classes of mutants harboring mutations in proteins S3, L5, L7 (L12), L29, L30 and L33. One of them apparently lacks protein L7 of the large subunit. These mutants together with those reported previously (Isono et al., 1976) total one hundred and one ribosomal mutants in thirty different proteins. 相似文献
10.
C Meier T Knispel V E Marquez E De Clercq J Balzarini 《Nucleosides & nucleotides》1999,18(4-5):907-912
Novel lipophilic cycloSal-triesters 3 and 4 from the ara- and ribo-configurated 2'-fluorinated ddAs 1 and 2, respectively, were prepared. The title compounds 3 and 4 delivered the corresponding monophosphates and thus, increasing the bioactivity or convert a formerly inactive compound into a RT inhibitor. 相似文献
11.
Harris SL Spears PA Havell EA Hamrick TS Horton JR Orndorff PE 《Journal of bacteriology》2001,183(13):4099-4102
PCR mutagenesis and a unique enrichment scheme were used to obtain two mutants, each with a single lesion in fimH, the chromosomal gene that encodes the adhesin protein (FimH) of Escherichia coli type 1 pili. These mutants were noteworthy in part because both were altered in the normal range of cell types bound by FimH. One mutation altered an amino acid at a site previously shown to be involved in temperature-dependent binding, and the other altered an amino acid lining the predicted FimH binding pocket. 相似文献
12.
Two F- mutants deficient in conjugation with F-donors have been characterized. They map at about 83 minut position, show resistance to T3 and T7 bacteriophages, and form mating aggregates in the liquid medium with lowered efficiency. Mutants have no detectable alterations in their outer membrane protein composition. 相似文献
13.
Escherichia coli K12 arabinose-binding protein mutants with altered transport properties. 总被引:1,自引:1,他引:0
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D. G. Kehres R. W. Hogg 《Protein science : a publication of the Protein Society》1992,1(12):1652-1660
The arabinose-binding protein (ABP) of Escherichia coli binds L-arabinose in the periplasm and delivers it to a cytoplasmic membrane complex consisting of the AraG and AraH proteins, for uptake into the cell. To study the interaction between the soluble and membrane components of this periplasmic transport system, regions of the ABP surface containing the opening of the arabinose-binding cleft were subjected to site-directed mutagenesis. Thirty-eight ABP variants containing one to three amino acid substitutions were recovered. ABP variants were expressed with wild-type AraG and AraH from a plasmid, in a strain lacking the chromosomal araFGH operon, and the whole cell uptake parameters, Ven (maximum initial velocity of arabinose entry) and K(en) (concentration of arabinose yielding half-maximal entry) were determined. Twenty-four mutants had normal Ven values, 3 mutants had Ven and K(en) values twice wild type, and 11 mutants had Ven and K(en) values 20-50% of wild type. Binding proteins that had altered uptake properties were each expressed, processed, and localized to the periplasm at levels equivalent to wild type. The mutant binding proteins behaved the same as wild type during purification, and each had a Kd (dissociation constant for bound arabinose) comparable to that of wild-type ABP. Mutations that resulted in altered uptake identified nine amino acids surrounding the arabinose-binding cleft, all of which are charged in the wild-type protein, and all of whose side chains project outward from the cleft. The evidence suggests that this surface of the binding protein and these nine charged loci play a major role in ABP interactions with the membrane complex. 相似文献
14.
Escherichia coli mutants with altered cation recognition by the melibiose carrier. 总被引:4,自引:2,他引:2
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Revertants that showed normal cation recognition for melibiose transport were isolated from mutants with altered cation recognition (W3133-2S and W3133-2T) of Escherichia coli. Although the original two mutants possessed a second alteration, an increased activity of the Na+(Li+)/H+ antiporter, the revertants, which possessed the normal melibiose carrier, still showed altered properties of the Na+(Li+)/H+ antiporter. These results support the view that the alterations in the melibiose carrier and in the Na+(Li+)/H+ antiporter, observed in the mutants, are not genetically linked. 相似文献
15.
J Balzarini M J Robins R M Zou P Herdewijn E De Clercq 《Biochemical and biophysical research communications》1987,145(1):277-283
The 2',3'-dideoxyriboside of 2,6-diaminopurine (ddDAPR) and its 2',3'-didehydro derivative (ddeDAPR) are poor substrates for adenosine deaminase (ADA) but potent inhibitors of the enzyme. Their Km values for ADA are of the same order of magnitude as those of the natural adenosine (Ado) and 2'-deoxyadenosine (dAdo), but their Vmax values are 35-fold (ddDAPR) to 350-fold (ddeDAPR) lower than those of Ado and dAdo. The Ki/K values of ADA for ddeDAPR (as inhibitor) and Ado, 2',3'-dideoxyadenosine (ddAdo) and 9-beta-D-arabinofuranosyladenine (araA) as the substrates are 0.17, 0.05 and 0.06, respectively. ddDAPR is about 3-fold less potent as an inhibitor of ADA than ddeDAPR. The 2,6-diaminopurine derivatives ddeDAPR and ddDAPR [which is also a potent inhibitor of human immunodeficiency virus (HIV)], may hold great promise, from a chemotherapeutic viewpoint, in combination with other adenosine analogues such as ddAdo and araA, which have been recognized and/or being pursued as either anti-retrovirus or anti-herpesvirus agents. 相似文献
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17.
Escherichia coli mutants with altered control of alcohol dehydrogenase and nitrate reductase. 总被引:7,自引:16,他引:7
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Mutants of Escherichia coli which overproduce alcohol dehydrogenase were obtained by selection for the ability to use ethanol as an acetate source in a strain auxotrophic for acetate. A mutant having a 20-fold overproduction of alcohol dehydrogenase was able to use ethanol only to fulfill its acetate requirement, whereas two mutants with a 60-fold overproduction were able to use ethanol as a sole carbon source. The latter two mutants produced only 25% of the wild-type level of nitrate reductase, when grown under anaerobic conditions. Alcohol dehydrogenase production was largely unaffected by catabolite repression but was repressed by nitrate under both aerobic and anaerobic conditions. The genetic locus responsible for alcohol dehydrogenase overproduction was located at min 27 on the E. coli genetic map; the gene order, as determined by transduction, was trp tonB adh chlC hemA. The possible relationship of alcohol dehydrogenase to anaerobic redox systems such as formate-nitrate reductase is discussed. 相似文献
18.
Protein synthesis defects in temperature-sensitive mutants of Escherichia coli with altered ribosomal proteins 总被引:2,自引:0,他引:2
W S Champney 《Biochimica et biophysica acta》1980,609(3):464-474
The ribosomes from four temperature-sensitive mutants of Escherichia coli have been examined for defects in cell-free protein synthesis. The mutants examined had alterations in ribosomal proteins S10, S15, or L22 (two strains). Ribosomes from each mutant showed a reduced activity in the translation of phage MS2 RNA at 44 degrees C and were more rapidly inactivated by heating at this temperature compared to control ribosomes. Ribosomal subunits from three of the mutants demonstrated a partial or complete inability to reassociate at 44 degrees C. 70-S ribosomes from two strains showed a reducton in messenger RNA binding. tRNA binding to the 30 S subunit was reduced in the strains with altered 30-S proteins and binding to the 50 S subunit was affected in the mutants with a change in 50 S protein L22. The relation between ribosomal protein structure and function in protein synthesis in these mutants is discussed. 相似文献
19.
Isolation of Escherichia coli mutants (cpdB) deficient in periplasmic 2':3'-cyclic phosphodiesterase and genetic mapping of the cpdB locus 总被引:7,自引:0,他引:7
Mutants of Escherichia coli deficient in the periplasmic enzyme 2':3'-cyclic phosphodiesterase have been obtained. The gene, designated cpdB, was mapped by conjugation and transduction and found to be located about 0 . 11 min to the right of the cycA locus on the E. coli genetic map. 相似文献
20.
Metabolism and anti-human immunodeficiency virus-1 activity of 2-halo-2',3'-dideoxyadenosine derivatives 总被引:14,自引:0,他引:14
T Haertle C J Carrera D B Wasson L C Sowers D D Richman D A Carson 《The Journal of biological chemistry》1988,263(12):5870-5875
Both 2',3'-dideoxyadenosine and 2',3'-dideoxyinosine have been shown (Mitsuya, H., and Broder, S. (1987) Nature 325, 773-778) to have in vitro activity against the human immunodeficiency virus-1 (HIV). However, these dideoxynucleosides may be catabolized by human T cells, even when adenosine deaminase is inhibited by deoxycoformycin. To overcome this problem, we have synthesized the 2-fluoro-, 2-chloro-, and 2-bromo-derivatives of 2',3'-dideoxyadenosine. The metabolism and anti-HIV activity of the 2-halo-2',3'-dideoxyadenosine derivatives and of 2',3'-dideoxyadenosine were compared. The 2-halo-2',3'-dideoxyadenosine derivatives were not deaminated significantly by cultured CEM T lymphoblasts. Experiments with 2-chloro-2',3'-dideoxyadenosine showed that the T cells converted the dideoxynucleoside to the 5'-monophosphate, 5'-diphosphate, and 5'-triphosphate metabolites. At concentrations lower than those producing cytotoxicity in uninfected cells (3-10 microM), the 2-halo-2',3-dideoxyadenosine derivatives inhibited the cytopathic effects of HIV toward MT-2 T lymphoblasts, and retarded viral replication in CEM T lymphoblasts. Experiments with a deoxycytidine kinase-deficient mutant CEM T cell line showed that this enzyme was necessary for the phosphorylation and anti-HIV activity of the 2-chloro-2',3'-dideoxyadenosine. In contrast, 2',3'-dideoxyadenosine was phosphorylated by the deoxycytidine kinase-deficient mutant and retained anti-HIV activity in this cell line. Thus, the 2-halo derivatives of 2',3'-dideoxyadenosine, in contrast to 2',3'-dideoxyadenosine itself, are not catabolized by T cells. Their anti-HIV and anti-proliferative activities are manifest only in cells expressing deoxycytidine kinase. The in vivo implications of these results for anti-HIV chemotherapy are discussed. 相似文献