|
|
|
|
|
|
Chemical Diversity of Polyene Macrolides Produced by Streptomyces noursei ATCC 11455 and Recombinant Strain ERD44 with Genetically Altered Polyketide Synthase NysC. Per Bruheim, 2004.The gram-positive bacterium Streptomyces noursei ATCC 11455 produces a complex mixture of polyene macrolides generally termed nystatins . Although the structures for nystatins A1 and A3 have been reported, the identities of other components of the nystatin complex remain obscure . Analyses of the culture extract from the S . noursei wild type revealed the presence of several nystatin-related compounds for which chemical structures could be suggested on the basis of their molecular weights, their UV spectra, and knowledge of the nystatin biosynthetic pathway . Nuclear magnetic resonance (NMR) studies with one of these polyene macrolides identified it as a nystatin analogue containing a mycarose moiety at C-35 . A similar investigation was performed with the culture extract of the ERD44 mutant, which has a genetically altered polyketide synthase (PKS) NysC and which was previously shown to produce a heptaene nystatin analogue . The latter compound, tentatively named S44HP, and its derivative, which contains two deoxysugar moieties, were purified; and their structures were confirmed by NMR analysis . Nystatin analogues with an expanded macrolactone ring were also observed in the extract of the ERD44 mutant, suggesting that the altered PKS can "stutter" during the polyketide chain assembly . These data provide new insights into the biosynthesis of polyene macrolide antibiotics and the functionalities of PKSs and post-PKS modification enzymes . Emergence of Macrolide-Resistant Streptococcus pyogenes Strains in French Children. Edouard Bingen, 2004.We studied the antimicrobial susceptibility of 322 Streptococcus pyogenes throat isolates from French children and their serotype and genomic diversity . A total of 22.4% were erythromycin resistant, and 69.4, 4.2, and 26.4% of these isolates harbored ermB, ermA, and mefA, respectively . Increasing resistance in France is mainly associated with a few emm type 28 clones . Interplay of the Czc System and Two P-Type ATPases in Conferring Metal Resistance to Ralstonia metallidurans. Antje Legatzki, 2003.Cadmium and zinc are removed from cells of Ralstonia metallidurans by the CzcCBA efflux pump and by two soft-metal-transporting P-type ATPases, CadA and ZntA . The czcCBA genes are located on plasmid pMOL30, and the cadA and zntA genes are on the bacterial chromosome . Expression of zntA from R . metallidurans in Escherichia coli predominantly mediated resistance to zinc, and expression of cadA predominantly mediated resistance to cadmium . Both transporters decreased the cellular content of zinc or cadmium in this host . In the plasmid-free R . metallidurans strain AE104, single gene deletions of cadA or zntA had only a moderate effect on cadmium and zinc resistance, but zinc resistance decreased 6-fold and cadmium resistance decreased 350-fold in double deletion strains . Neither single nor double gene deletions affected zinc resistance in the presence of czcCBA . In contrast, cadmium resistance of the cadA zntA double mutant could be elevated only partially by the presence of CzcCBA . lacZ reporter gene fusions indicated that expression of cadA was induced by cadmium but not by zinc in R . metallidurans strain AE104 . In the absence of the zntA gene, expression of cadA occurred at lower cadmium concentrations and zinc now served as an inducer . In contrast, expression of zntA was induced by both zinc and cadmium, and the induction pattern did not change in the presence or absence of CadA . However, expression of both genes, zntA and cadA, was diminished in the presence of CzcCBA . This indicated that CzcCBA efficiently decreased cytoplasmic cadmium and zinc concentrations . It is discussed whether these data favor a model in which the cations are removed either from the cytoplasm or the periplasm by CzcCBA . Monitoring the Bacterial Population Dynamics in Sourdough Fermentation Processes by Using PCR-Denaturing Gradient Gel Electrophoresis. Christiane B. Meroth, 2003.Four sourdoughs (A to D) were produced under practical conditions by using a starter mixture of three commercially available sourdough starters and a baker's yeast constitutively containing various species of lactic acid bacteria (LAB) . The sourdoughs were continuously propagated until the composition of the LAB flora remained stable . Two LAB-specific PCR-denaturing gradient gel electrophoresis (DGGE) systems were established and used to monitor the development of the microflora . Depending on the prevailing ecological conditions in the different sourdough fermentations, only a few Lactobacillus species were found to be competitive and became dominant . In sourdough A (traditional process with rye flour), Lactobacillus sanfranciscensis and a new species, L . mindensis, were detected . In rye flour sourdoughs B and C, which differed in the process temperature, exclusively L . crispatus and L . pontis became the predominant species in sourdough B and L . crispatus, L . panis, and L . frumenti became the predominant species in sourdough C . On the other hand, in sourdough D (corresponding to sourdough C but produced with rye bran), L . johnsonii and L . reuteri were found . The results of PCR-DGGE were consistent with those obtained by culturing, except for sourdough B, in which L . fermentum was also detected . Isolates of the species L . sanfranciscensis and L . fermentum were shown by randomly amplified polymorphic DNA-PCR analysis to originate from the commercial starters and the baker's yeast, respectively . The Bacterium Thermus thermophilus, Like Hyperthermophilic Archaea, Uses a Two-Step Pathway for the Synthesis of Mannosylglycerate. Nuno Empadinhas, 2003.The biosynthetic pathway for the synthesis of the compatible solute  -mannosylglycerate (MG) in the thermophilic bacterium Thermus thermophilus HB27 was identified based on the activities of recombinant mannosyl-3-phosphoglycerate synthase (MPGS) (EC 2.4.1.217) and mannosyl-3-phosphoglycerate phosphatase (MPGP) (EC 3.1.3.70) . The sequences of homologous genes from the archaeon Pyrococcus horikoshii were used to identify MPGS and MPGP genes in T . thermophilus HB27 genome . Both genes were separately cloned and overexpressed in Escherichia coli, yielding 3 to 4 mg of pure recombinant protein per liter of culture . The molecular masses were 43.6 and 28.1 kDa for MPGS and MPGP, respectively . The recombinant MPGS catalyzed the synthesis of
-mannosyl-3-phosphoglycerate (MPG) from GDP-mannose and D-3-phosphoglycerate, while the recombinant MPGP catalyzed the dephosphorylation of MPG to MG . The recombinant MPGS had optimal activity at 80 to 90°C and a pH optimum near 7.0; MPGP had maximal activity between 90 and 95°C and at pH 6.0 . The activities of both enzymes were strictly dependent on divalent cations; Mn2+ was most effective for MPGS, while Mn2+, Co2+, Mg2+, and to a lesser extent Ni2+ activated MPGP . The organization of MG biosynthetic genes in T . thermophilus HB27 is different from the P . horikoshii operon-like structure, since the genes involved in the conversion of fructose-6-phosphate to GDP-mannose are not found immediately downstream of the contiguous MPGS and MPGP genes . The biosynthesis of MG in the thermophilic bacterium T . thermophilus HB27, proceeding through a phosphorylated intermediate, is similar to the system found in hyperthermophilic archaea .
Marker Rescue Studies of the Transfer of Recombinant DNA to Streptococcus gordonii In Vitro, in Foods and Gnotobiotic Rats. Mitra Kharazmi, 2003.A plasmid marker rescue system based on restoration of the nptII gene was established in Streptococcus gordonii to study the transfer of bacterial and transgenic plant DNA by transformation . In vitro studies revealed that the marker rescue efficiency depends on the type of donor DNA . Plasmid and chromosomal DNA of bacteria as well as DNA of transgenic potatoes were transferred with efficiencies ranging from 8.1 x 10-6 to 5.8 x 10-7 transformants per nptII gene . Using a 792-bp amplification product of nptII the efficiency was strongly decreased (9.8 x 10-9) . In blood sausage, marker rescue using plasmid DNA was detectable (7.9 x 10-10), whereas in milk heat-inactivated horse serum (HHS) had to be added to obtain an efficiency of 2.7 x 10-11 . No marker rescue was detected in extracts of transgenic potatoes despite addition of HHS . In vivo transformation of S . gordonii LTH 5597 was studied in monoassociated rats by using plasmid DNA . No marker rescue could be detected in vivo, although transformation was detected in the presence of saliva and fecal samples supplemented with HHS . It was also shown that plasmid DNA persists in rat saliva permitting transformation for up to 6 h of incubation . It is suggested that the lack of marker rescue is due to the absence of competence-stimulating factors such as serum proteins in rat saliva .
|
© 2005
Transgalactic Ltd (manufacturer of Bioscreen C software) |
Privacy Statement | P.O. Box
1393, 00101 Helsinki, Finland,
Last modified: May 25, 2005
| ||||||
