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1.
The Streptomyces coelicolor genome contains 17 TerD domain-encoding genes (tdd genes) of unknown function. The proteins encoded by these genes have been presumed to be involved in tellurite resistance
on the basis of their homology with the protein TerD of Serratia marcescens. To elucidate the role of a Tdd protein (Tdd8), both a deletion mutant for the corresponding gene tdd8 (SCO2368) and a recombinant strain over-expressing tdd8 were produced in S. coelicolor M145. The deletion mutant (Δtdd8), like the wild strain, was not resistant to potassium tellurite. The deletion was not lethal but had a marked effect on
differentiation. The deletion strain showed more rapid growth in liquid medium and produced long chains of short spores with
a dense and non-spherical spore wall on agar plates. The strain over-expressing tdd8 had a growth delay in liquid medium and produced very few spores of irregular shapes and sizes on solid medium. The results
of this study demonstrated that Tdd proteins might have a function other than tellurite resistance and this function seems
to be of crucial importance for the proper development of the actinomycete S. coelicolor. 相似文献
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Tetiana Gren Bohdan Ostash Volodymyr Babiy Ihor Rokytskyy Victor Fedorenko 《Folia microbiologica》2018,63(2):197-201
Streptomyces coelicolor genome carries two apparently paralogous genes, SCO4164 and SCO5854, that encode putative thiosulfate sulfurtransferases (rhodaneses). These genes (and their presumed translation products) are highly conserved and widely distributed across actinobacterial genomes. The SCO4164 knockout strain was unable to grow on minimal media with either sulfate or sulfite as the sole sulfur source. The SCO5854 mutant had no growth defects in the presence of various sulfur sources; however, it produced significantly less amounts of actinorhodin. Furthermore, we discuss possible links between basic interconversions of inorganic sulfur species and secondary metabolism in S. coelicolor. 相似文献
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Lee HN Kim HJ Kim P Lee HS Kim ES 《Journal of industrial microbiology & biotechnology》2012,39(5):805-811
Along with traditional random mutagenesis-driven strain improvement, cloning and heterologous expression of Streptomyces secondary metabolite gene clusters have become an attractive complementary approach to increase its production titer, of
which regulation is typically under tight control via complex multiple regulatory networks present in a metabolite low-producing
wild-type strain. In this study, we generated a polyketide non-producing strain by deleting the entire actinorhodin cluster
from the chromosome of a previously generated S. coelicolor mutant strain, which was shown to stimulate actinorhodin biosynthesis through deletion of two antibiotic downregulators as
well as a polyketide precursor flux downregulator (Kim et al. in Appl Environ Microbiol 77:1872–1877, 2011). Using this engineered S. coelicolor mutant strain as a surrogate host, a model minimal polyketide pathway for aloesaponarin II, an actinorhodin shunt product,
was cloned in a high-copy conjugative plasmid, followed by functional pathway expression and quantitative metabolite analysis.
Aloesaponarin II production was detected only in the presence of a pathway-specific regulatory gene, actII-ORF4, and its production level was the highest in the actinorhodin cluster-deleted and downregulator-deleted mutant strain, implying
that this engineered polyketide pathway-free and regulation-optimized S. coelicolor mutant strain could be used as a general surrogate host for efficient expression of indigenous or foreign polyketide pathways
derived from diverse actinomycetes in nature. 相似文献
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Lingjun Yu Shuxian Li Wenyan Gao Yuanyuan Pan Huarong Tan Gang Liu 《Applied microbiology and biotechnology》2015,99(7):3141-3153
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Previously it was shown that the Arabidopsis apyrase genes AtAPY1 and AtAPY2 are crucial for male fertility because mutant pollen (apy1-1; apy2-1) with T-DNA insertions in both genes could not germinate (Steinebrunner et al. (2003) Plant Physiol. 131: 1638–1647). In this study, pollen germination was restored and apyrase T-DNA double knockouts (DKO)
apy1-1/apy1-1; apy2-1/apy2-1 were generated by complementation with AtAPY2 under the control of a pollen-specific promoter. The DKO phenotype displayed developmental defects including the lack of
functional root and shoot meristems. In cotyledons, morphogenetic and patterning abnormalities were apparent, e.g., unlobed
pavement cells and stomatal clusters. Another set of lines was created which carried either AtAPY1 or AtAPY2 under a dexamethasone-(DEX)-inducible promoter as an additional transgene to the pollen-specific gene construct. Application
of DEX did not reverse the DKO phenotype to wild-type, but some inducible lines exhibited less severe defects even in the
absence of the inducer, probably due to some background expression. However, even these DKO mutants were seedling-lethal and
shared other defects regarding cell division, cell expansion and stomatal patterning. Taken together, the defects in the DKO
mutants demonstrate that AtAPY1 and AtAPY2 are essential for normal plant development. 相似文献
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We isolated several mutants of Arabidopsis thaliana (L.) Heynh. that accumulated less anthocyanin in the plant tissues, but had seeds with a brown color similar to the wild-type.
These mutants were allelic with the anthocyaninless1 (anl1) mutant that has been mapped at 15.0 cM of chromosome 5. We performed fine mapping of the anl1 locus and determined that ANL1 is located between the nga106 marker and a marker corresponding to the MKP11 clone. About 70 genes are located between these
two markers, including three UDP-glucose:flavonoid-3-O-glucosyltransferase-like genes and a glutathione transferase gene (TT19). A mutant of one of the glucosyltransferase genes (At5g17050) was unable to complement the anl1 phenotype, showing that the ANL1 gene encodes UDP-glucose:flavonoid-3-O-glucosyltransferase. ANL1 was expressed in all tissues examined, including rosette leaves, stems, flower buds and roots. ANL1 was not regulated by TTG1. 相似文献
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A simple and random genome deletion method combining insertion sequence (IS) element IS31831 and the Cre/loxP excision system generated 42 Corynebacterium glutamicum mutants (0.2–186 kb). A total of 393.6 kb (11.9% of C. glutamicum R genome) coding for 331 genes was confirmed to be nonessential under standard laboratory conditions. The deletion strains,
generated using only two vectors, varied not only in their lengths but also the location of the deletion along the C. glutamicum R genome. By comparing and analyzing the generated deletion strains, identification of nonessential genes, the roles of genes
of hitherto unknown function, and gene–gene interactions can be easily and efficiently determined.
Electronic supplementary material Supplementary material is available in the online version of this article at and is accessible for authorized users. 相似文献
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Zummo FP Marineo S Pace A Civiletti F Giardina A Puglia AM 《Applied microbiology and biotechnology》2012,94(3):719-728
Most enzymes involved in tryptophan catabolism via kynurenine formation are highly conserved in Prokaryotes and Eukaryotes.
In humans, alterations of this pathway have been related to different pathologies mainly involving the central nervous system.
In Bacteria, tryptophan and some of its derivates are important antibiotic precursors. Tryptophan degradation via kynurenine
formation involves two different pathways: the eukaryotic kynurenine pathway, also recently found in some bacteria, and the
tryptophan-to-anthranilate pathway, which is widespread in microorganisms. The latter produces anthranilate using three enzymes
also involved in the kynurenine pathway: tryptophan 2,3-dioxygenase (TDO), kynureninase (KYN), and kynurenine formamidase
(KFA). In Streptomyces coelicolor, where it had not been demonstrated which genes code for these enzymes, tryptophan seems to be important for the calcium-
dependent antibiotic (CDA) production. In this study, we describe three adjacent genes of S. coelicolor (SCO3644, SCO3645, and SCO3646), demonstrating their involvement in the tryptophan-to-anthranilate pathway: SCO3644 codes for a KFA, SCO3645 for a KYN and SCO3646 for a TDO. Therefore, these genes can be considered as homologous respectively to kynB, kynU, and kynA of other microorganisms and belong to a constitutive catabolic pathway in S. coelicolor, which expression increases during the stationary phase of a culture grown in the presence of tryptophan. Moreover, the S. coelicolor ΔkynU strain, in which SCO3645 gene is deleted, produces higher amounts of CDA compared to the wild-type strain. Overall, these results describe a pathway,
which is used by S. coelicolor to catabolize tryptophan and that could be inactivated to increase antibiotic production. 相似文献
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Lisette Maria Catharina Nitsch Carla Oplaat Richard Feron Qian Ma Mieke Wolters-Arts Peter Hedden Celestina Mariani Wim Hendrik Vriezen 《Planta》2009,229(6):1335-1346
Although the hormones, gibberellin and auxin, are known to play a role in the initiation of fruits, no such function has yet
been demonstrated for abscisic acid (ABA). However, ABA signaling and ABA responses are high in tomato (Solanum lycopersicum L.) ovaries before pollination and decrease thereafter (Vriezen et al. in New Phytol 177:60–76, 2008). As a first step to understanding the role of ABA in ovary development and fruit set in tomato, we analyzed ABA content
and the expression of genes involved in its metabolism in relation to pollination. We show that ABA levels are relatively
high in mature ovaries and decrease directly after pollination, while an increase in the ABA metabolite dihydrophaseic acid
was measured. An important regulator of ABA biosynthesis in tomato is 9-cis-epoxy-carotenoid dioxygenase (LeNCED1), whose mRNA level in ovaries is reduced after pollination. The increased catabolism
is likely caused by strong induction of one of four newly identified putative (+)ABA 8′-hydroxylase genes. This gene was named
SlCYP707A1 and is expressed specifically in ovules and placenta. Transgenic plants, overexpressing SlCYP707A1, have reduced ABA levels and exhibit ABA-deficient phenotypes suggesting that this gene encodes a functional ABA 8′-hydroxylase.
Gibberellin and auxin application have different effects on the LeNCED1 and SlCYP707A1 gene expression. The crosstalk between auxins, gibberellins and ABA during fruit set is discussed. 相似文献
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Bijay Singh Tae-Jin Oh Jae Kyung Sohng 《Journal of industrial microbiology & biotechnology》2009,36(10):1257-1265
Thorough investigation of Streptomyces peucetius ATCC 27952 genome revealed a sesquiterpene synthase, named spterp13, which encodes a putative protein of 732 amino acids with significant similarity to S. avermitilis MA-4680 (SAV2163, GeoA) and S. coelicolor A3(2) (SCO6073). The proteins encoded by SAV2163 and SCO6073 produce geosmin in the respective strains. However, the spterp13 gene seemed to be silent in S. peucetius. Deletion of the doxorubicin gene cluster from S. peucetius resulted in increased cell growth rate along with detectable production of geosmin. When we over expressed the spterp13 gene in S. peucetius DM07 under the control of an ermE* promoter, 2.4 ± 0.4-fold enhanced production of geosmin was observed. 相似文献
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The nifA gene is an important regulatory gene and its product, NifA protein, regulates the expression of many nif genes involved in the nitrogen fixation process. We introduced multiple copies of the constitutively expressed Sinorhizobium meliloti (Sm) or Enterobacter cloacae (Ec) nifA gene into both the nifA mutant strain SmY and the wild-type strain Sm1021. Root nodules produced by SmY containing a constitutively expressed Sm nifA gene were capable of fixing nitrogen, while nodules produced by SmY containing the Ec nifA gene remained unable to fix nitrogen, as is the case for SmY itself. However, transfer of an additional Sm nifA gene into Sm1021 improved the nitrogen-fixing efficiency of root nodules to a greater extent than that observed upon transfer
of the Ec nifA gene into Sm1021. Comparative analysis of amino acid sequences between Sm NifA and Ec NifA showed that the N-terminal domain was the least similar, but this domain is indispensable for complementation of the
Fix- phenotype of SmY by Sm NifA. We conclude that more than one domain is involved in determining functional differences between Sm NifA and Ec NifA. 相似文献
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Genome predictions based on selected genes would be a very welcome approach for taxonomic studies. We analyzed three genes,
recN, flaA, and ftsY, for determining if these genes are useful tools for systematic analyses in the genus Anoxybacillus. The genes encoding a DNA repair and genetic recombination protein (recN), the flagellin protein (flaA), and GTPase signal docking protein (ftsY) were sequenced for ten Anoxybacillus species. The sequence comparisons revealed that recN sequence similarities range between 61% and 99% in the genus Anoxybacillus. Comparisons to other bacterial recN genes indicated that levels of similarity did not differ from the levels within genus Anoxybacillus. These data showed that recN is not a useful marker for the genus Anoxybacillus. A 550–600-bp region of the flagellin gene was amplified for all Anoxybacillus strains except for Anoxybacillus contaminans. The sequence similarity of flaA gene varies between 61% and 76%. Comparisons to other bacterial flagellin genes obtained from GenBank (Bacillus, Pectinatus, Proteus, and Vibrio) indicated that the levels of similarity were lower (3–42%). Based on these data, we concluded that the variability in this
single gene makes it a particularly useful marker. Another housekeeping gene ftsY suggested to reflect the G+C (mol/mol) content of whole genome was analyzed for Anoxybacillus strains. A mean difference of 1.4% was observed between the G+C content of the gene ftsY and the G+C content of the whole genome. These results showed that the gene ftsY can be used to represent whole G+C content of the Anoxybacillus species. 相似文献
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The aim of this study was to evaluate the MPK1 (SLT2) gene deletion upon filamentous growth induced by isoamyl alcohol (IAA) in two haploid industrial strains of Saccharomyces cerevisiae using oligonucleotides especially designed for a laboratory S. cerevisiae strain. The gene deletion was performed by replacing part of the open reading frames from the target gene with the KanMX gene. The recombinant strains were selected by their resistance to G418, and after deletion confirmation by polymerase chain
reaction, they were cultivated in a yeast extract peptone dextrose medium + 0.5% IAA to evaluate the filamentous growth in
comparison to wild strains. Mpk1 derivatives were obtained for both industrial yeasts showing the feasibility of the oligonucleotides especially designed
for a laboratory strain (Σ1278b) by Martinez-Anaya et al. (In yeast, the pseudohyphal phenotype induced by isoamyl alcohol
results from the operation of the morphogenesis checkpoint. J Cell Sci 116:3423–3431, 2003). The filamentation rate in these
derivatives was significantly lower for both strains, as induced by IAA. This drastic reduction in the filamentation ability
in the deleted strains suggests that the gene MPK1 is required for IAA-induced filamentation response. The growth curves of wild and derivative strains did not differ substantially.
It is not known yet whether the switch to filamentous growth affects the fermentative characteristics of the yeast or other
physiological traits. A genetically modified strain for nonfilamentous growth would be useful for these studies, and the gene
MPK1 could be a target gene. The feasibility of designed oligonucleotides for this deletion in industrial yeast strains is shown. 相似文献
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Mono-ADP-ribosylation is the enzymatic transfer of ADP-ribose from NAD+ to acceptor proteins catalyzed by ADP-ribosyltransferases. Using m-aminophenylboronate affinity chromatography, 2D-gel electrophoresis,
in-gel digestion and MALDI-TOF analysis we have identified eight in vitro ADP-ribosylated proteins in Streptomyces coelicolor, which can be classified into three categories: (i) secreted proteins; (ii) metabolic enzymes using NAD+/NADH or NADP+/NADPH as coenzymes; and (iii) other proteins. The secreted proteins could be classified into two functional categories: SCO2008
and SC05477 encode members of the family of periplasmic extracellular solute-binding proteins, and SCO6108 and SC01968 are
secreted hydrolases. Dehydrogenases are encoded by SC04824 and SC04771. The other targets are GlnA (glutamine synthetase I.,
SC02198) and SpaA (starvation-sensing protein encoded by SC07629). SCO2008 protein and GlnA had been identified as ADP-ribosylated
proteins in previous studies. With these results we provided experimental support for a previous suggestion that ADP-ribosylation
may regulate membrane transport and localization of periplasmic proteins. Since ADP-ribosylation results in inactivation of
the target protein, ADP-ribosylation of dehydrogenases might modulate crucial primary metabolic pathways in Streptomyces. Several of the proteins identified here could provide a strong connection between protein ADP-ribosylation and the regulation
of morphological differentiation in S. coelicolor. 相似文献