The main objective of this study was using matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) for assembling of DSM (German Collection of Microorganisms) Streptomyces spectral database and identification of wild Streptomyces cultures, which were clustered by MALDI-TOF Biotyper OC software as well as for teracycline detection by observing of obtained spectra using flexAnalysis software. Production of tetracycline was confirmed by thin-layer chromatography. Presence of tetracycline mass spectrum was verified by several tetracycline producers (Streptomyces aureofaciens LMG 5968, S. aureofaciens 84/25, and S. aureofaciens BMK) and by pure tetracycline mass. Our results showed that it is possible to use MALDI-TOF MS for identification of tetracycline producers within Streptomyces genera by several easy steps. The purpose of this study was to establish cheap and quick detection of tetracycline producers.
- MeSH
- databáze faktografické MeSH
- lidé MeSH
- rychlé screeningové testy metody MeSH
- software MeSH
- spektrometrie hmotnostní - ionizace laserem za účasti matrice * MeSH
- Streptomyces izolace a purifikace metabolismus MeSH
- tetracyklin chemie izolace a purifikace metabolismus MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- MeSH
- alkaloidy biosyntéza MeSH
- Bacteria genetika metabolismus MeSH
- biosyntéza peptidů nezávislá na nukleových kyselinách MeSH
- databáze genetické MeSH
- genetické markery MeSH
- houby genetika metabolismus MeSH
- metagenom MeSH
- mezinárodní spolupráce MeSH
- multigenová rodina * MeSH
- peptidy metabolismus MeSH
- polyketidy metabolismus MeSH
- polysacharidy biosyntéza MeSH
- proteosyntéza * MeSH
- rostliny genetika metabolismus MeSH
- terminologie jako téma MeSH
- terpeny metabolismus MeSH
- výpočetní biologie normy MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
The polyketide gene cluster aur1 is responsible for the production of the antibiotic auricin in Streptomyces aureofaciens CCM 3239. Auricin production is low and strictly regulated by two regulators, Aur1P and Aur1R. To improve auricin yield, we genetically manipulated S. aureofaciens CCM 3239 strain to overcome this strict regulation. A regulatory region including aur1R, aur1P, aur1O and the target biosynthetic aur1Ap promoter were replaced by the strong constitutive ermEp* promoter. However, auricin production was decreased in such a genetically manipulated strain. In the second strategy we placed the aur1P gene for auricin pathway-specific activator under the control of the ermEp* promoter. The resulting strain has been shown to produce 2.8-fold higher amount of auricin compared with the WT strain.
- MeSH
- antibakteriální látky biosyntéza MeSH
- DNA bakterií genetika metabolismus MeSH
- makrolidy metabolismus MeSH
- multigenová rodina MeSH
- plazmidy genetika MeSH
- promotorové oblasti (genetika) MeSH
- regulace genové exprese u bakterií MeSH
- restrikční mapování MeSH
- Streptomyces aureofaciens genetika metabolismus MeSH
- vysokoúčinná kapalinová chromatografie MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
The role of MicA (repressing small regulatory non-coding RNAs of two Salmonella porins) was determined in virulence of Salmonella enterica serovar Typhimurium. Transcriptional analysis revealed that the expression of the micA gene is driven by a single σ(E)-dependent promoter, micAp. Its activity increased towards stationary phase; in exponential phase, the activity was induced by several stresses by a DegS-dependent mechanism. Although phenotypic analysis revealed no significant differences between wild-type and the micA mutant strains, in vivo studies showed that this mutant is more virulent in the mouse model.
- MeSH
- bakteriální proteiny genetika metabolismus MeSH
- bakteriální RNA genetika metabolismus MeSH
- lidé MeSH
- molekulární sekvence - údaje MeSH
- myši inbrední BALB C MeSH
- myši MeSH
- nekódující RNA genetika metabolismus MeSH
- regulace genové exprese u bakterií MeSH
- Salmonella typhimurium genetika metabolismus patogenita MeSH
- salmonelóza mikrobiologie MeSH
- sekvence nukleotidů MeSH
- sigma faktor genetika metabolismus MeSH
- virulence MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- ženské pohlaví MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
An incomplete oligoketide (PK; 'polyketide') gene cluster, aur1, responsible for the production of an angucycline-like antibiotic auricin was identified in Streptomyces aureofaciens CCM 3239. A region downstream of the aur1 was cloned and sequenced, revealing 28 new genes encoding putative protein products involved in deoxysugar biosynthesis and other putative PK-related biosynthetic functions. In addition, a gene, bpsA, encoding a protein similar to non-ribosomal peptide synthetases (NRPSs) was identified in this region. A deduced protein product of the gene showed the highest similarity to NRPSs IndC from Erwinia chrysanthemi and BpsA from Streptomyces lavendulae, both involved in the biosynthesis of a blue pigment indigoidine. S. aureofaciens CCM 3239 was found to produce an extracellular blue pigment with identical properties as indigoidine. A deletion mutant of bpsA in S. aureofaciens CCM 3239 failed to produce the blue pigment. In addition, the deletion of bpsA had a positive effect on auricin production. The results indicate the involvement of the bpsA gene in biosynthesis of the indigoidine blue pigment in S. aureofaciens CCM 3239.
The transcription start points of the penicillin biosynthesis genes from Penicillium chrysogenum were mapped using the primer extension method. For each of the three genes consensus sequences of the core promoter elements were identified, supporting the notion that the basal transcription of these genes is mediated separately. Interestingly, transcription start of the pcbC gene is located within the potential Inr element with no TATA box-like sequence being found at expected position. This is in contrast to pcbAB and penDE genes with proposed TATA boxes or even to Aspergillus nidulans ipnA (pcbC) gene indicating possible differences in basal transcription regulation. Using the quantitative RT-PCR analysis the expression of all three biosynthesis genes was monitored in both the high and low production strain of P. chrysogenum during a 3-d cultivation under production conditions. The differences were found between the strains in time regulation and transcript levels of the biosynthesis genes. Furthermore, we showed that the effect of higher gene dosage on productivity in the production strain is amplified by more efficient transcription of the biosynthesis genes with the RNA levels approximately 37- and 12-times higher, respectively, than in a low production strain.
- MeSH
- financování organizované MeSH
- fungální proteiny genetika metabolismus MeSH
- molekulární sekvence - údaje MeSH
- peniciliny biosyntéza MeSH
- Penicillium chrysogenum genetika metabolismus MeSH
- polymerázová řetězová reakce s reverzní transkripcí MeSH
- promotorové oblasti (genetika) MeSH
- regulace genové exprese u hub MeSH
- sekvence aminokyselin MeSH
- sekvence nukleotidů MeSH
A DNA fragment containing part of the salinomycin biosynthetic gene cluster from industrial strain Streptomyces albus CCM 4719 was cloned. Sequence analysis of the 25.809-kbp fragment revealed the presence of 8 open reading frames (ORFs), including two large ORFs encoding three modular sets of oligoketide synthase, followed by three genes (salRI, salRII, salRIII) encoding transcriptional regulators. The first two regulators, SalRI and SalRII, belonged to the novel LAL family of large transcriptional regulators. SalRIII was highly similar to the NysRIV, AmphRIV, and FscRI transcriptional regulators from the oligoene macrolides nystatin, amphotericin, and R008/candicidin clusters, respectively.
- MeSH
- finanční podpora výzkumu jako téma MeSH
- genomová knihovna MeSH
- klonování DNA metody MeSH
- otevřené čtecí rámce genetika MeSH
- polymerázová řetězová reakce metody využití MeSH
- pyrany izolace a purifikace MeSH
- regulační geny genetika MeSH
- sekvenční analýza DNA metody využití MeSH
- Streptomyces genetika izolace a purifikace růst a vývoj MeSH
