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Volume 142,
Issue 10,
1996
Volume 142, Issue 10, 1996
- Biochemistry
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The O polysaccharide chain of the lipopolysaccharide from Vibrio cholerae 076 is a homopolymer of N-[(S)-(+)-2-hydroxypropionyl]-α-L-perosamine
More LessChemical and serological studies of LPS from Vibrio cholerae 076 (076) were performed. The LPS of 076 contained d-glucose, d-galactose, l-glycero-d-manno-heptose, d-fructose, d-glucosamine, d-quinovosamine (2-amino-2,6-dideoxy-d-glucose) and L-perosamine (4-amino-4,6-dideoxy-l-mannopyranose). The sugar composition of the LPS from 076 was quite similar to that of LPS from V. cholerae 01 with the exception of the presence of a small amount of d-galactose in the LPS of 076. However, perosamine, a major sugar component of the LPS from 0076, was in the l configuration in contrast to the d configuration of the perosamine in the LPS of V. cholerae 01. The l-perosamine was N-acylated with an (S)-(+)-2-hydroxypropionyl group in the LPS from 076. Structural analysis by NMR spectroscopy, as well as GC/MS, revealed that the O polysaccharide chain of the LPS from 076 was an α(1 → 2)-linked homopolymer of N-[(S)-(+)-2-hydroxypropionyl]-l-perosamine. The serological cross-reactivity between the LPS of 076 and the LPS from other strains, such as V. cholerae 01 (Ogawa and Inaba 0 forms), Vibrio bio-serogroup 1875 (Original and Variant strains), V. cholerae O140 (Hakata) and Yersinia enterocolitica O9, was examined in passive haemolysis tests with sheep red blood cells that had been sensitized with LPS and antisera raised against whole cells of these bacteria. The latter six strains have in common the O antigen that includes Inaba antigen factor C, in addition to their own O-antigenic factors. Thus, they cross-react serologically. The O polysaccharide chains of the LPS of these six strains are known to consist exclusively of α(1 → 2)-linked d-perosamine homopolymers and differences are found only among the N-acyl substituents. In passive haemolysis tests, the LPS of 076 did not cross-react serologically with any of the other LPS examined. Thus, the results obtained in this study support the hypothesis that Inaba antigen factor C, associated with the antigens of these six strains, which include V. cholerae O1, is related substantially and exclusively to their α(1 → 2)-linked homopolymers of N-acylated d-perosamine, and not to such homopolymers of N-acylated l-perosamine.
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Evidence for an arginine-specific mono(ADP-ribosyl)transferase in dormant spores of the fungus Phycomyces blakesleeanus
More LessA soluble mono(ADP-ribosyl)transferase was detected in dormant spores of Phycomyces blakesleeanus. Soluble proteins incubated with [32P]NAD revealed, after two-dimensional electrophoresis, three major ADP-ribosylated substrates with molecular masses of 38, 37 and 36 kDa and pl values of 6·9, 8·1 and 4·6, respectively. When these endogenous substrates were first ADP-ribosylated with [32P]NAD and then incubated for different times with either 3 M hydroxylamine (pH 7·0) or 1 mM HgCl2, only hydroxylamine released the incorporated radioactivity after 30 min incubation. Additionally, agmatine was used as a substrate for this enzyme. These data suggest that the mono(ADP-ribosyl)transferase is an arginine-specific enzyme. This enzymic activity was stimulated by 10 mM MgCl2 and by 250 μM of the nitric-oxide-releasing agent sodium nitroprusside, and inhibited by 8 mM benzamide, 0·4 mM m-iodobenzylguanidine and 0·5 mM novobiocin. The three ADP-ribosylation inhibitors affected the germination of P. blakesleeanus spores, leaving them as swollen cells. The effect of MgCl2, GTP and ATP on the ADP-ribosylation of the endogenous proteins was studied. The presence of two additional [32P]ADP-ribosylated proteins of 57 and 55 kDa was observed in the absence of MgCl2. An increase in incorporation of radioactivity into the 55 kDa band was observed when the assay was performed in the presence of GTP or ATP. The addition of Mg2+ together with either or both nucleotides eliminated the appearance of the 57 and 55 kDa bands, but intensified the 37 kDa band. Photoaffinity-labelling of the soluble fraction with [α-32P]GTP revealed a 55 kDa band together with other proteins of 32 and 17 kDa. These results suggest that among the five different endogenous substrates for the fungal mono(ADP-ribosyl)transferase, the 55 kDa protein may be a GTP-binding protein.
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Glycerol is not an inhibitor of mitochondrial citrate oxidation by Aspergillus niger
More LessWe have re-assessed the hypothesis that an accumulation of intracellular glycerol triggers the accumulation of citric acid by Aspergillus niger by inhibiting the activity of the mitochondrial NADP-dependent isocitrate dehydrogenase isoenzyme. To this end, we have incubated mycelia of A. niger with 0·5 M glycerol, which resulted in a maximal intracellular glycerol pool level of 0·92 M, comparable to that determined during the early phase of citrate accumulation. This addition affected neither the uptake of [1,5-14C]citrate from the medium nor the rate of the subsequent release of 14CO2 by the mycelia, indicating no effect on citrate oxidation. Mitochondria isolated from mycelia previously loaded with 0·5 M glycerol contained 8% of the total mycelial glycerol. They released 14CO2 from exogenously added [1,5-14C]citrate at the same rate as mitochondria isolated from mycelia not loaded with 0·5 M glycerol. The addition of glycerol had no effect on the activity of the purified NADP-specific isocitrate dehydrogenase, but appeared to inhibit the activity in crude cell-free extracts of A. niger. We conclude that the intracellular accumulation of glycerol does not affect the rate of mitochondrial citrate oxidation and is therefore, in contrast to previous claims, not a trigger of citrate accumulation.
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The urea cycle of Helicobacter pylori
More LessThe presence and activities of the enzymes of the urea cycle in the bacterium Helicobacter pylori were investigated employing one- and two-dimensional NMR spectroscopy and radioactive tracer analysis. Cell suspensions, lysates and membrane preparations generated l-ornithine and ammonium at high rates in incubations with l-arginine, indicating the presence of arginase activity. Anabolic ornithine transcarbamoylase (OTCase) activity was identified by the formation of heat-stable products in incubations of cell-free extracts with ornithine and radiolabelled carbamoyl phosphate. The heat-labile product that resulted from incubations of cell-free extracts with citrulline radiolabelled in the guanidino moiety revealed the presence of catabolic OTCase activity. Argininosuccinate formation and catalysis indicated the presence of argininosuccinate synthetase and argininosuccinase activities. The findings suggested that H. pylori has a urea cycle which acts as an effective mechanism to extrude excess nitrogen from cells.
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- Biotechnology
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Selection for carotenogenesis in the yeast Phaffia rhodozyma by dark-generated singlet oxygen
More LessSelection for carotenogenesis in Phaffia rhodozyma was achieved by exposure of yeast strains to dark chemical reactions that generate singlet oxygen. Incubation of a mixture of P. rhodozyma strains containing varying levels of carotenoids in hypochlorous acid or hydrogen peroxide resulted in weak selection for pigmented strains. However, the combination of hydrogen peroxide and hypochlorous acid was strongly selective for carotenogenesis and gave a monoculture of a carotenoid-hyperproducer. Exposure of the yeast to ozone for 10 to 20 min also selected for a hyperproducing strain. These selections were relieved by 1,4-diazabicyclo[2.2.2]-octane, a specific quencher of singlet oxygen or by l-ascorbic acid. Continuous growth of P. rhodozyma on agar plates in an ozone/air atmosphere for 5 d decreased astaxanthin and total carotenoid levels and increased the levels of carotenoid biosynthetic intermediates. Repeated rounds of random mutagenesis followed by ozone exposure yielded mutant strains with higher pigmentation than control cultures. Our results support the hypothesis that a primary function of carotenoids in P. rhodozyma is to protect against singlet oxygen generated in the natural environment of the yeast and that a practical method for preventing strain degeneration during industrial fermentations may be achieved by generation of singlet oxygen using simple chemical supplements or by bubbling ozone through P. rhodozyma cultures during fermentation.
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- Development And Structure
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The response of selected members of the archaea to the Gram stain
More LessArchaea possess a broader range of cell envelope structural formats than eubacteria and their cell walls do not contain peptidoglycan. Some archaea have only a single S-layer as their cell wall (e.g. Methanococcus jannaschii and Sulfolobus acidocaldarius), whereas others have multiple layers (e.g. Methanospirillum hungatei). Sometimes there can also be a high proportion of tetraether lipids in membranes to make the envelope more resilient to environmental stress (e.g. Methanococcus jannaschii and Sulfolobus acidocaldarius grown at 70 °C). Since the Gram reaction depends on both the structural format and the chemical composition of the cell envelope of eubacteria, it was important to determine if the same is true for archaea. Methanospirillum hungatei, Methanosarcina mazei, Methanobacterium formicicum, Methanococcus jannaschii and Sulfolobus acidocaldarius, chosen because of their different envelope formats and chemistries, were subjected to a Gram stain that can be used for transmission electron microscopy. In this staining regimen, the iodine is replaced by potassium trichloro(η2-ethylene)platinate(II) as the mordant, and the platinum of the new compound is the electron-scattering agent for electron microscopy. Of all these archaea, only Methanobacterium formicicum stained Gram-positive since its pseudomurein wall remained intact; the platinum compound formed large electron-dense aggregates with the crystal violet that were located in the vicinity of the cell wall and the plasma membrane. All but the terminal filament cells of Methanospirillum hungatei stained Gram-negative because the limiting porosity of its external sheath was so small that the Gram reagents could not enter the cells. The terminal cells of filaments stained Gram-positive because the staining reagents gained entry through the terminal plugs. All other archaea stained Gram-negative because their cell walls were so disrupted during staining that the crystal violet-platinum complex could not be retained by the cells. Methanococcus jannaschii was grown at both 50 °C and 70 °C so that the tetraether lipids in its plasma membrane could be increased from 20% (50 °C) to 45% (70 °C) of the total lipids; in both cases the cells stained Gram-negative.
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SPR28, a sixth member of the septin gene family in Saccharomyces cerevisiae that is expressed specifically in sporulating cells
More LessThe septins are a recently recognized family of proteins that are present in a wide variety of fungal and animal cells, where they are involved in cytokinesis and apparently in other processes involving the organization of the cell surface. Five previously described Saccharomyces cerevisiae septins are associated with the neck filaments of vegetative cells and/or with the developing prospore wall of sporulating cells. We report here the characterization of SPR28, a sixth member of the S. cerevisiae septin gene family whose existence was revealed by the yeast genome project. Analysis of mRNA levels showed that SPR28 is a new member of the group of ‘late genes’ that are expressed at high levels during the meiotic divisions and ascospore formation. The septin it encodes, Spr28p, exhibited specific two-hybrid interactions with itself and with three other septins that are expressed in sporulating cells. Consistent with these results, an Spr28p-green fluorescent protein fusion was induced during meiosis 1 and appeared to be associated with the developing prospore walls. Deletion of SPR28 in either a wild-type or an spr3∆ background produced no obvious abnormalities in vegetative cells and had little or no effect on sporulation, suggesting that the septins have redundant roles during spore formation.
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- Genetics And Molecular Biology
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Isolation of an IHF-deficient mutant of a Pseudomonas aeruginosa mucoid isolate and evaluation of the role of IHF in algD gene expression
More LessThe role of integration host factor (IHF) in the regulation of alginate synthesis was investigated in a mucoid strain of Pseudomonas aeruginosa (strain CHA) isolated from a cystic fibrosis patient. Escherichia coli strain BL21 (DE3) was made IHF-deficient by inactivation of its chromosomal IHF genes, himA and himD, then used as host strain to overproduce P. aeruginosa IHF. The purified recombinant IHF protein was used to determine the affinity of IHF for the two IHF binding sites in the algD promoter. The K d values were determined to be 130 nM for algD IHF site 2 and about 2μM for algD IHF site 1. Two IHF-deficient mutants of P. aeruginosa strain CHA were constructed by insertional inactivation of the himA gene, and the activity of the algD promoter was determined using transcriptional fusion with xylE as reporter gene. The expression of algD, the structural gene for GDP-mannose dehydrogenase, was decreased three- to fourfold in the himA mutants under conditions of high salinity and nitrogen limitation. Assays of alginate production by cultures grown on agar plates indicated that the IHF-deficient mutants synthesized 50% less polymer than the mucoid parental strain. These results demonstrate clearly that although IHF is dispensable for alginate production, himA expression is required for full activation of algD expression.
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Functional definition of regions necessary for replication and incompatibility in the Mycobacterium fortuitum plasmid pAL5000
More LessDifferent parts of the Mycobacterium fortuitum plasmid pAL5000 necessary for plasmid replication and incompatibility were defined and studied. Two ORFs, named repA and repB, were defined which are necessary for replication. A pAL5000 derivative deleted in these genes can be made to replicate by providing the gene products in trans. A 435 bp fragment was defined which was necessary in cis for replication and which had an influence on copy number. This region (inc), which contains several repeated motifs, was also able to confer a degree of incompatibility when cloned into an otherwise unrelated mycobacterial replicon. pAL5000-derived plasmids carrying two copies of the inc region had a lower copy number and were less stable than the wild-type. These effects were only observed when the two regions were in the same orientation. Plasmids carrying only the inc region and no other parts of pAL5000 could be made to replicate if repA and repB were supplied in trans from another plasmid. Based on these findings, systems for selectively curing cells of one plasmid of a pair were designed and shown to be functional in Mycobacterium smegmatis. These have potential as a simple delivery system for achieving transposon mutagenesis or gene replacement in mycobacteria.
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Mutants of Streptomyces roseosporus that express enhanced recombination within partially homologous genes
More LessStreptomyces roseosporus mutants that express enhanced recombination between partially homologous (homeologous) sequences were isolated by selection for recombination between the bacteriophage øC31 derivative KC570 containing the Streptomyces coelicolor glucose kinase (glk) gene and the S. roseosporus chromosome. The frequencies of homeologous recombination in the ehr mutants were determined by measuring the chromosomal insertion frequencies of plasmids containing S. coelicolor glnA or whiG genes. S. roseosporus ehr mutants showed 102- to 104-fold increases in homeologous recombination relative to Ehr+ strains, but no increase in homologous recombination. Southern hybridization analysis revealed single unique sites for the insertion of each of the plasmids, and the crossovers occurred in frame and in proper translational register, yielding functional chimeric glnA and whiG genes.
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An amplifiable and deletable locus of Streptomyces ambofaciens RP181110 contains a very large gene homologous to polyketide synthase genes
Streptomyces ambofaciens RP181110 produces the macrolide polyketide spiramycin. Like many other Streptomyces species, the RP181110 strain is prone to genetic instability involving genomic rearrangements (deletions and/or amplifications) in the large unstable region of the genome. It has previously been demonstrated that the amplification of a particular locus (AUD205) affects spiramycin biosynthesis and, conversely, the loss of this amplification is correlated with the restoration of antibiotic production. This report focuses on a 0·93 kb reiterated fragment specific for the AUD205 locus. Sequencing of 3596 bp including this reiteration revealed the presence of an ORF (orfPS) whose potential product was highly homologous to the EryA and Raps proteins, responsible for the biosynthesis of erythromycin in Saccharopolyspora erythraea and rapamycin in Streptomyces hygroscopicus, respectively. orfPS encodes a protein with at least four successive domains: ketoacyl synthase, acyltransferase, ketoreductase and acyl carrier protein. This organization is very similar to most eryA and rap modules. The reiterated sequence corresponds to the acyltransferase domain. orfPS was transcribed during rapid growth and stationary phase in RP181110 and overtranscribed in the amplified mutant. Both these results suggest that the gene encodes a type I polyketide synthase and its reorganization is responsible for the loss of spiramycin production in the amplified strains.
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Mutational analysis and chemical modification of Cys24 of lactococcin B, a bacteriocin produced by Lactococcus lactis
More LessUsing site-directed mutagenesis the single cysteine residue at position 24 of lactococcin B was replaced by all other possible amino acids. Most of these mutant molecules retained bacteriocin activity, with the exception of those in which cysteine was replaced by a positively charged amino acid. This would seem to be in agreement with the authors’ earlier observation that treatment of the wild-type molecule with HgCI2 resulted in its inactivation. The factor that causes inactivation of lactococcin B seems to be the introduction of a positive charge at position 24 by HgCI2 rather than oxidation of this residue, as treatment of the bacteriocin with other oxidative chemicals did not interfere with the ability of lactococcin B to dissipate the membrane potential of sensitive cells. Results are also reported which imply that inactive lactococcin B can still bind to its receptor. It can be replaced by an active bacteriocin molecule, resulting in dissipation of the membrane potential.
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Photosynthesis gene expression in Rhodobacter sphaeroides is regulated by redox changes which are linked to electron transport
More LessThe effect of the respiratory electron acceptor nitrous oxide on the synthesis of the photosynthetic apparatus of Rhodobacter sphaeroides was examined. In phototrophically grown cells, the addition of nitrous oxide caused a reduction in the level of light-harvesting complex I and light-harvesting complex II under conditions of high light intensity (200 W m−2) and low light intensity (16 W m−2). 5-Aminolaevulinate synthase activity was decreased during growth in the presence of nitrous oxide and this limited production of spectral complexes since addition of exogenous 5-aminolaevulinic acid partially suppressed the effect of nitrous oxide. The effect of nitrous oxide on the expression of the operons encoding the pigment-binding proteins of light-harvesting complex I (puf), light-harvesting complex II (puc) and the two isoenzymes of 5-aminolaevulinate synthase (hemA and hemT) were measured using transcriptional fusions to lacZ. Nitrous oxide caused a decrease in puf and hemA transcription. However, there was an apparent increase in the expression of puc and hemT transcriptional fusions. The level of light-harvesting complexes in cells grown in the dark with different electron acceptors was also examined. Cells grown anaerobically with DMSO had a higher level of light-harvesting complexes than those grown anaerobically with nitrous oxide as electron acceptor. Cells grown aerobically had the lowest level of light-harvesting complexes. It is proposed that FnrL-dependent and FnrL-independent expression of photosynthesis genes is modulated by redox changes elicited by nitrous oxide respiration.
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Uptake-sequence-independent DNA transformation exists in Neisseria gonorrhoeae
More LessA DNA transformation dose-response curve of piliated (P+) gonococci with the use of cloned DNA containing a pilE2-cat fusion showed saturation at high and low levels of transforming DNA. At low DNA concentrations, transformation of the P+ strain MS11-A was effectively inhibited by a 1000-fold molar excess of the gonococcal transformation uptake sequence (GCUS). The same molar excess of the GCUS did not inhibit transformation of MS11-A at high DNA concentrations. In MS11-B2, a nonpiliated (P−), pilin-nonproducing, isogenic variant of MS11-A, the GCUS did not inhibit transformation at any level of transforming DNA. These data suggest that two mechanisms of transformation exist in P+ cells: one which utilizes the GCUS and one which does not. In MS11-B2 P− cells, no evidence was found for the presence of the GCUS-dependent mechanism, suggesting that transformation in this background occurs solely by the GCUS-independent mechanism.
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The extracellular acid protease gene of Yarrowia lipolytica: sequence and pH-regulated transcription
The gene encoding an acid extracellular protease (AXP) from Yarrowia lipolytica (Candida olea) 148 was cloned and the complete nucleotide sequence was determined. The amino acid sequence deduced from the nucleotide sequence reveals that the mature AXP consists of 353 amino acids with an M r of 37427. The gene also encodes a putative 17 amino acid hydrophobic prepeptide and a 27 amino acid propeptide containing no potential N-glycosylation sites. The mature extracellular enzyme is produced by cleavage between Phe and Ala. AXP is a member of the aspartyl family of proteases. AXP shows homology to proteases of several fungal genera and to human progastricin. The coding sequence is preceded by a potential regulatory region of 1982 bp. Transcription of both AXP and alkaline extracellular protease genes of Y. lipolytica 148 is regulated by the pH of culture.
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Detection of genetic variation in Ustilago maydis strains by probes derived from telomeric sequences
More LessGenetic variation using probes derived from telomeric sequences was analysed among several Ustilago maydis strains in an attempt to identify discriminative fingerprint patterns. Three groups of wild isolates from different geographical areas and one group of standard laboratory strains were examined. Analysis of the endmost restriction fragments (EFs) and of the endmost-associated restriction fragments (EAFs) of the chromosomes revealed group differences. Most of the EFs in two groups of strains showed a similar length whereas in the other two groups they were distributed in classes of different lengths. Furthermore, analysis of the EAFs permitted possible fingerprint patterns to be predicted for each group of strains based on the occurrence of amplified bands as well as the presence or absence of distinct bands which were shown to be present in terminal as well as in interstitial sites of the chromosome. The approach evaluated in this work yielded highly polymorphic fingerprint patterns and could be used to distinguish groups of fungal isolates; this approach may also be effective for other fungal systems.
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Chorismate synthase from Staphylococcus aureus
More LessThe aroC gene encoding chorismate synthase and the ndk gene encoding nucleoside diphosphate kinase (Ndk) were cloned from Staphylococcus aureus. DNA sequencing suggests that aroC is located in an operon with aroB and aroA and encodes a protein of 388 amino acids with 61% identity to the aroF gene product of Bacillus subtilis. The ndk gene of S. aureus encodes a protein of 149 amino acids which exhibits a high degree of identity to other bacterial Ndk proteins. The 3′ end of the S. aureus gerCC gene was also identified by sequencing and was located immediately upstream of ndk. The gerCA and gerCB genes were found to be located upstream of gerCC by Southern hybridization analysis. This observed linkage of the gerC genes with the ndk, aroC and aroB genes has been similarly observed in B. subtilis. The S. aureus chorismate synthase was overexpressed to a high level in Escherichia coli using a T7 promoter plasmid construct, the enzyme was purified to near homogeneity in two steps and found to be a homotetramer with a subunit molecular mass, estimated by electrospray mass spectrometry, of 43024 Da. The properties of S. aureus chorismate synthase are compared with those of the B. subtilis and E. coli enzymes.
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Proline iminopeptidase gene from Xanthomonas campestris pv. citri
More LessThe pip gene coding for the proline iminopeptidase (Pip) of Xanthomonas campestris pv. citri was cloned in an Escherichia coli leuB strain using a selective medium containing the dipeptide d-Ala-l-Leu as the sole source of l-leucine. Nucleotide sequencing of this gene revealed a 939 bp open reading frame encoding a 312 amino acid protein (35 126 Da). The deduced amino acid sequence showed 47% identity with the Pip from Neisseria gonorrhoeae. A lacZ-pip fusion gene was overexpressed in E. coli under the control of the Plac promoter. The Pip of X. campestris hydrolysed l-prolyl-p-nitroanilide with the highest efficiency, but was also able to hydrolyse l-alanyl-p-nitroanilide and d-alanyl-p-nitroanilide. The molecular mass of Pip was found to be 35 kDa by SDS-PAGE and 120 kDa by gel filtration, suggesting that the active enzyme is a multimer.
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- Pathogenicity And Medical Microbiology
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Specific detection of Candida albicans and Candida tropicalis by fluorescent in situ hybridization with an 18S rRNA-targeted oligonucleotide probe
More LessIn situ hybridization of whole cells with rRNA-targeted, fluorescently labelled oligonucleotide probes is a powerful method to specifically detect micro-organisms in their natural habitat without cultivation and subsequent identification by phenotypic characterization. To examine the use of this method for the specific detection of pathogenic Candida species, we have designed an oligonucleotide probe which binds to the 18S rRNA of C. albicans and C. tropicalis, the two most important pathogenic Candida species, and differentiates them from other clinically relevant species. After establishing suitable hybridization conditions, we confirmed the specificity of our probe O20 in RNA dot blot hybridizations with a series of reference strains and clinical isolates of medically important Candida species. All C. albicans and C. tropicalis strains hybridized with the probe, whereas all strains of C. parapsilosis, C. glabrata, C. krusei, C. guilliermondii, C. kefyr and C. lusitaniae did not. When we used the fluorescently labelled probe O20 to specifically detect single cells of the two target species by in situ hybridization, both C. albicans and C. tropicalis reacted strongly with the probe and could be clearly differentiated from C. krusei and C. parapsilosis, although the latter organism contains only two nucleotide mismatches in the probe target region. This discrimination capacity was also seen when mixed suspensions of C. albicans and C. parapsilosis were hybridized with the probe. After infection of a human endothelial cell line with C. albicans and C. krusei, C. albicans cells adhering to the endothelial cells were easily distinguishable from the C. krusei cells by fluorescent in situ hybridization with probe O20. In addition, germ tubes and hyphae of C. albicans were also efficiently labelled. The application of fluorescently labelled rRNA-targeted oligonucleotide probes therefore appears to be a valuable tool for the specific detection and identification of different members of the genus Candida, which does not require any cultivation.
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Candida albicans adherence to a human oesophageal cell line
More LessThe oesophageal epithelium appears to be one of the primary cell targets of Candida albicans in AIDS patients. To study this interaction, we have established an in vitro adherence assay using a human epithelial oesophageal cell line (HET1-A). When yeast cells were grown in 500 mM d-galactose, adherence increased significantly over cultures prepared in 500 mM d-glucose. In addition to HET1-A cells, adherence of the organism grown in d-galactose to human buccal epithelial cells and a murine alveolar macrophage cell line was also higher. Adherence of yeast cells to HET1-A cells was partially inhibited in the presence of d-glucosamine or N-acetyl-d-glucosamine, but not with d-mannose, d-glucose, l-fucose or d-galactose. Attachment to HET1-A cells was studied using scanning and transmission electron microscopy. Partial phagocytosis of adhering yeast cells was observed occasionally within the first 90 min following infection, as evidenced by the formation of HET1-A pseudopodia in instances of close contact with yeast cells. The influence of d-galactose on cell surface proteins was studied by analysing β-mercaptoethanol-extracted proteins from yeast cells grown in either 500 mM d-galactose or d-glucose. From d-galactose-grown cells only, a glycoprotein of approximately 190 kDa was observed in Aurodye-stained SDS-PAGE gels and in Western blots using an immunoglobulin fraction (IgG) prepared from sera of rabbits infected with the organism. These studies demonstrate that C. albicans adheres to human oesophageal cells and may utilize cell surface proteins whose synthesis is nutritionally regulated.
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Volume 35 (1964)
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Volume 34 (1964)
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Volume 33 (1963)
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Volume 32 (1963)
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Volume 31 (1963)
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Volume 30 (1963)
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Volume 29 (1962)
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Volume 28 (1962)
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Volume 27 (1962)
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Volume 26 (1961)
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Volume 25 (1961)
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Volume 24 (1961)
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Volume 23 (1960)
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Volume 22 (1960)
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Volume 21 (1959)
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Volume 20 (1959)
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Volume 19 (1958)
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Volume 18 (1958)
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Volume 17 (1957)
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Volume 16 (1957)
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Volume 15 (1956)
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Volume 14 (1956)
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Volume 13 (1955)
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Volume 12 (1955)
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Volume 11 (1954)
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Volume 10 (1954)
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Volume 9 (1953)
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Volume 8 (1953)
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Volume 7 (1952)
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Volume 6 (1952)
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Volume 5 (1951)
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Volume 4 (1950)
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Volume 3 (1949)
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Volume 2 (1948)
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Volume 1 (1947)
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