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1.
The structural gene for sphingomyelinase (SMase) from Streptomyces
griseocarneus, was introduced into Streptomyces lividans using a shuttle vector, pUC702, for Escherichia coli/S. lividans. High-level secretory production of SMase was achieved using the promoter, signal sequence and terminator regions of phospholipase
D from Streptoverticillium cinnamoneum. The transformant constitutively expressed a high specific activity of SMase extracellularly during batch culture. Maximum
SMase activity (555 ± 114 U/mg protein) was with 1.75 M MgCl2 which was about 50-fold more than that with 10 mM MgCl2. 相似文献
2.
Díaz M Adham SA Ramón D Gil JA Santamaría RI 《Applied microbiology and biotechnology》2004,65(4):401-406
The Aspergillus nidulans gene xlnA coding for the fungal xylanase X22 has been cloned and expressed in two heterologous bacterial hosts: Streptomyces lividans and Brevibacterium lactofermentum. Streptomyces strains yielded 10 units/ml of xylanase when the protein was produced with its own signal peptide, and 19 units/ml when its signal peptide was replaced by the one for xylanase Xys1 from Streptomyces halstedii. B. lactofermentum was also able to produce xylanase X22, affording 6 units/ml upon using either the Aspergillus xlnA signal peptide or Streptomyces xysA. These production values are higher than those previously reported for the heterologous expression of the A. nidulans xlnA gene in Saccharomyces cerevisiae (1 unit/ml). Moreover, the X22 enzyme produced by Streptomyces lividans showed oenological properties, indicating that this Streptomyces recombinant strain is a good candidate for the production of this enzyme at the industrial scale. 相似文献
3.
Mohamed Belal Hamed Spyridoula Karamanou Solveig Ólafsdottir Joana Sofia Martins Basílio Kenneth Simoens Kostantinos C. Tsolis Lieve Van Mellaert Eik Elísabet Guðmundsdóttir Gudmundur Oli Hreggvidsson Jozef Anné Kristel Bernaerts Olafur H. Fridjonsson Anastassios Economou 《Microbial cell factories》2017,16(1):232
Background
The gene encoding a thermostable cellulase of family 12 was previously isolated from a Rhodothermus marinus through functional screening. CelA is a protein of 260 aminoacyl residues with a 28-residue amino-terminal signal peptide. Mature CelA was poorly synthesized in some Escherichia coli strains and not at all in others. Here we present an alternative approach for its heterologous production as a secreted polypeptide in Streptomyces.Results
CelA was successfully over-expressed as a secreted polypeptide in Streptomyces lividans TK24. To this end, CelA was fused C-terminally to the secretory signal peptide of the subtilisin inhibitor protein (Sianidis et al. in J Biotechnol. 121: 498–507, 2006) from Streptomyces venezuelae and a new cloning strategy developed. Optimal growth media and conditions that stall biomass production promote excessive CelA secretion. Under optimal growth conditions in nutrient broth medium, significant amounts of mature CelA (50–90 mg/L or 100–120 mg/g of dry cell weight) are secreted in the spent growth media after 7 days. A protocol to rapidly purify CelA to homogeneity from culture supernatants was developed and specific anti-sera raised against it. Biophysical, biochemical and immmuno-detection analyses indicate that the enzyme is intact, stable and fully functional. CelA is the most thermostable heterologous polypeptide shown to be secreted from S. lividans.Conclusion
This study further validates and extends the use of the S. lividans platform for production of heterologous enzymes of industrial importance and extends it to active thermostable enzymes. This study contributes to developing a platform for poly-omics analysis of protein secretion in S. lividans.4.
The protein glutaminase (PG) secreted by the Gram-negative bacterium Chryseobacterium proteolyticum can deamidate glutaminyl residues in several substrate proteins, including insoluble wheat glutens. This enzyme therefore
has potential application in the food industry. We assessed the possibility to produce PG containing a pro-domain in Corynebacterium glutamicum which we have successfully used for production of several kinds of proteins at industrial-scale. When it was targeted to
the general protein secretion pathway (Sec) via its own signal sequence, the protein glutaminase was not secreted in this
strain. In contrast, we showed that pro-PG could be efficiently produced using the recently discovered twin-arginine translocation
(Tat) pathway when the typical Sec-dependent signal peptide was replaced by a Tat-dependent signal sequence from various bacteria.
The accumulation of pro-PG in C. glutamicum ATCC13869 reached 183 mg/l, and the pro-PG was converted to an active form as the native one by SAM-P45, a subtilisin-like
serine protease derived from Streptomyces albogriseolus. The successful secretion of PG via this approach confirms that the Tat pathway of C. glutamicum is an efficient alternative for the industrial-scale production of proteins that are not efficiently secreted by other systems. 相似文献
5.
Thuy ML Kharel MK Lamichhane R Lee HC Suh JW Liou K Sohng JK 《Biotechnology letters》2005,27(7):465-470
An actinomycetes expression vector (pIBR25) was constructed and applied to express a gene from the kanamycin biosynthetic gene cluster encoding 2-deoxy-scyllo-inosose synthase (kanA) in Streptomyces lividans TK24. The expression of kanA in pIBR25 transformants reached a maximum after 72 h of culture. The plasmid pIBR25 showed better expression than pSET152, and resulted in the formation of insoluble KanA when it was expressed in Escherichia coli. This strategy thus provides a valuable tool for expressing aminoglycoside-aminocyclitols (AmAcs) biosynthetic genes in Streptomyces spp. 相似文献
6.
Background
Wolbachia are endosymbiotic bacteria that commonly infect numerous arthropods. Despite their broad taxonomic distribution, the transmission patterns of these bacteria within and among host species are not well understood. We sequenced a portion of the wsp gene from the Wolbachia genome infecting 138 individuals from eleven geographically distributed native populations of the fire ant Solenopsis invicta. We then compared these wsp sequence data to patterns of mitochondrial DNA (mtDNA) variation of both infected and uninfected host individuals to infer the transmission patterns of Wolbachia in S. invicta. 相似文献7.
8.
Rosa Garcia Sanchez Bärbel Hahn-Hägerdal Marie F Gorwa-Grauslund 《Microbial cell factories》2010,9(1):40
Background
In Saccharomyces cerevisiae galactose is initially metabolized through the Leloir pathway after which glucose 6-phosphate enters glycolysis. Galactose is controlled both by glucose repression and by galactose induction. The gene PGM2 encodes the last enzyme of the Leloir pathway, phosphoglucomutase 2 (Pgm2p), which catalyses the reversible conversion of glucose 1-phosphate to glucose 6-phosphate. Overexpression of PGM2 has previously been shown to enhance aerobic growth of S. cerevisiae in galactose medium. 相似文献9.
10.
Kazuyoshi Ohta Hidenori Tanaka Daisuke Yamakawa Hironori Hamasuna Hirohisa Fujimoto 《Journal of industrial microbiology & biotechnology》2011,38(8):967-973
An extracellular xylanase XynI of glycoside hydrolase family 11 from the dimorphic fungus Aureobasidium pullulans ATCC 20524 possesses an N-terminal extension of 34 amino acids (Ohta et al., J. Biosci. Bioeng. 92:262–270, 2001). The N-terminal
extension includes three sites (Ala-X-Ala-X-Ala-X-Ala) that are potentially cleavable by signal peptidase I of Escherichia
coli. The A. pullulans
xynI signal sequence was fused in frame to the mature protein region of the equivalent xylanase gene xynA from the filamentous fungus Penicillium citrinum. The gene fusion xynI::A was inserted into the plasmid pET-26b(+) to yield pEXP401. An E. coli BL21(DE3) transformant harboring the pEXP401 exhibited xylanase activity (per ml of the culture) of 16.8 U in the fraction
of culture supernatant as well as 4.29 U in the fraction of cell-free extract after 12 h of growth with isopropyl-β-d-thiogalactopyranoside at 30°C. N-terminal amino acid sequence analysis of the secreted recombinant proteins revealed cleavage
at four distinct sites within the N-terminal extension of XynI, two of which conformed to the Ala-X-Ala motif prior to the
cleavage site. The XynA proteins secreted into the culture medium showed high specific activities from 506 to 651 U/mg, which
were twofold higher than that of the native enzyme. 相似文献
11.
Background
Sperm cells must regulate the timing and location of activation to maximize the likelihood of fertilization. Sperm from most species, including the nematode Caenorhabditis elegans, activate upon encountering an external signal. Activation for C. elegans sperm occurs as spermatids undergo spermiogenesis, a profound cellular reorganization that produces a pseudopod. Spermiogenesis is initiated by an activation signal that is transduced through a series of gene products. It is now clear that an inhibitory pathway also operates in spermatids, preventing their premature progression to spermatozoa and resulting in fine-scale control over the timing of activation. Here, we describe the involvement of a newly assigned member of the inhibitory pathway: spe-4, a homolog of the human presenilin gene PS1. The spe-4(hc196) allele investigated here was isolated as a suppressor of sterility of mutations in the spermiogenesis signal transduction gene spe-27. 相似文献12.
Noda S Miyazaki T Miyoshi T Miyake M Okai N Tanaka T Ogino C Kondo A 《Journal of industrial microbiology & biotechnology》2011,38(5):643-648
Cinnamic acid production was demonstrated using Streptomyces as a host. A gene encoding phenylalanine ammonia lyase (PAL) from Streptomyces maritimus was introduced into Streptomyces lividans, and its expression was confirmed by Western blot analysis. After 4 days cultivation using glucose as carbon source, the
maximal level of cinnamic acid reached 210 mg/L. When glycerol (30 g/L) was used as carbon source, the maximal level of produced
cinnamic acid reached 450 mg/L. In addition, using raw starch, xylose or xylan as carbon source, the maximal level of cinnamic
acid reached 460, 300, and 130 mg/L, respectively. We demonstrated that S. lividans has great potential to produce cinnamic acid as well as other aromatic compounds. 相似文献
13.
Background
Hormones frequently guide animal development via the induction of cascades of gene activities, whose products further amplify an initial hormonal stimulus. In Drosophila the transformation of the larva into the pupa and the subsequent metamorphosis to the adult stage is triggered by changes in the titer of the steroid hormone 20-hydroxyecdysone. singed wings (swi) is the only gene known in Drosophila melanogaster for which mutations specifically interrupt the transmission of the regulatory signal from early to late ecdysone inducible genes. 相似文献14.
Background
Campylobacter jejuni has been divided into two subspecies: C. jejuni subsp. jejuni (Cjj) and C. jejuni subsp. doylei (Cjd). Nearly all of the C. jejuni strains isolated are Cjj; nevertheless, although Cjd strains are isolated infrequently, they differ from Cjj in two key aspects: they are obtained primarily from human clinical samples and are associated often with bacteremia, in addition to gastroenteritis. In this study, we utilized multilocus sequence typing (MLST) and a DNA microarray-based comparative genomic indexing (CGI) approach to examine the genomic diversity and gene content of Cjd strains. 相似文献15.
Background
Phospholipid biosynthesis commences with the acylation of glycerol-3-phosphate (G3P) to form 1-acyl-G3P. This step is catalyzed by the PlsB protein in Escherichia coli. The gene encoding this protein has not been identified, however, in the majority of bacterial genome sequences, including that of Bacillus subtilis. Recently, a new two-step pathway catalyzed by PlsX and PlsY proteins for the initiation of phospholipid formation in Streptococcus pneumoniae has been reported. 相似文献16.
Svetlana?E?Moskalenko Svetlana?V?Chabelskaya Sergei?G?Inge-Vechtomov Michel?Philippe Galina?A?Zhouravleva
Background
Termination of protein synthesis in eukaryotes involves at least two polypeptide release factors (eRFs) – eRF1 and eRF3. The highly conserved translation termination factor eRF1 in Saccharomyces cerevisiae is encoded by the essential gene SUP45. 相似文献17.
Ying Huang Xin Wang Zhigang Cui Yuhuan Yang Yuchun Xiao Liuying Tang Biao Kan Jianguo Xu Huaiqi Jing 《BMC microbiology》2010,10(1):211
Background
Yersinia enterocolitica is an enteric pathogen that invades the intestinal mucosa and proliferates within the lymphoid follicles (Peyer's patches). The attachment invasion locus (ail) mediates invasion by Y. enterocolitica and confers an invasive phenotype upon non-invasive E. coli; ail is the primary virulence factor of Y. enterocolitica. The ferrioxamine receptor (foxA) located on the Y. enterocolitica chromosome, together with its transport protein, transports a siderophore specific for ferric ion. Currently, ail is the primary target gene for nucleic acid detection of pathogenic Y. enterocolitica. 相似文献18.
19.
Purpose of work
Undecylprodigiosin has antimicrobial, immunosuppressive and anticancer properties. Pure cultures of Streptomyces coelicolor produce only low concentrations of undecylprodigiosin. We have therefore sought to increase this by interacting the cells with a bacterium. 相似文献20.
A novel lipase gene, lipJ08, was cloned from Candida rugosa ATCC14830, along with the already reported five lipase genes (lip1–lip5). Nucleotide sequencing indicated that the lipJ08 gene contains a 1650 bp open reading frame (ORF) without introns. The deduced amino acid sequence corresponds to 534 amino
acid residues, including a putative signal sequence of 15 amino acid residues. Seventeen of the non-universal serine codons
(CTG) of lipJ08 were converted into universal serine codons (TCT) by PCR-based mutagenesis. The native and codon-optimized lipJ08 genes were expressed in Pichia
pastoris. The hydrolytic activity of the recombinant LIPJ08 was 4.7 U/ml, whereas the activity of the recombinant wild-type lipase
could not be detected. 相似文献