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  • American Society for Microbiology  (8)
  • Biodiversity Research  (8)
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  • American Society for Microbiology  (8)
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  • Biodiversity Research  (8)
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  • 1
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 83, No. 20 ( 2017-10-15)
    Abstract: Putrescine is an important polyamine that participates in a variety of stress responses. Ornithine decarboxylase (ODC) is a key enzyme that catalyzes the biosynthesis of putrescine. A homolog of the gene encoding ODC was cloned from Ganoderma lucidum . In the ODC -silenced strains, the transcript levels of the ODC gene and the putrescine content were significantly decreased. The ODC -silenced strains were more sensitive to oxidative stress. The content of ganoderic acid was increased by approximately 43 to 46% in the ODC -silenced strains. The content of ganoderic acid could be recovered after the addition of exogenous putrescine. Additionally, the content of reactive oxygen species (ROS) was significantly increased by approximately 1.3-fold in the ODC -silenced strains. The ROS content was significantly reduced after the addition of exogenous putrescine. The gene transcript levels and the activities of four major antioxidant enzymes were measured to further explore the effect of putrescine on the intracellular ROS levels. Further studies showed that the effect of the ODC-mediated production of putrescine on ROS might be a factor influencing the biosynthesis of ganoderic acid. Our study reports the role of putrescine in large basidiomycetes, providing a basis for future studies of the physiological functions of putrescine in microbes. IMPORTANCE It is well known that ODC and the ODC-mediated production of putrescine play an important role in resisting various environmental stresses, but there are few reports regarding the mechanisms underlying the effect of putrescine on secondary metabolism in microorganisms, particularly in fungi. G. lucidum is gradually becoming a model organism for studying environmental regulation and metabolism. In this study, a homolog of the gene encoding ODC was cloned in Ganoderma lucidum . We found that the transcript level of the ODC gene and the content of putrescine were significantly decreased in the ODC -silenced strains. The content of ganoderic acid was significantly increased in the ODC -silenced strains. Further studies showed that the effect of the ODC-mediated production of putrescine on ROS might be a factor influencing the biosynthesis of ganoderic acid. Our study reports the role of putrescine in large basidiomycetes, providing a basis for future studies of the physiological functions of putrescine in microbes.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2017
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 2
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 83, No. 8 ( 2017-04-15)
    Abstract: Burkholderia contaminans MS14 was isolated from soil in Mississippi. When it is cultivated on nutrient broth-yeast extract agar, the colonies exhibit bactericidal activity against a wide range of plant-pathogenic bacteria. A bacteriostatic compound with siderophore activity was successfully purified and was determined by nuclear magnetic resonance spectroscopy to be ornibactin. Isolation of the bactericidal compound has not yet been achieved; therefore, the exact nature of the bactericidal compound is still unknown. During an attempt to isolate the bactericidal compound, an interesting relationship between the production of ornibactin and the bactericidal activity of MS14 was characterized. Transposon mutagenesis resulted in two strains that lost bactericidal activity, with insertional mutations in a nonribosomal peptide synthetase (NRPS) gene for ornibactin biosynthesis and a luxR family transcriptional regulatory gene. Coculture of these two mutant strains resulted in restoration of the bactericidal activity. Furthermore, the addition of ornibactin to the NRPS mutant restored the bactericidal phenotype. It has been demonstrated that, in MS14, ornibactin has an alternative function, aside from iron sequestration. Comparison of the ornibactin biosynthesis genes in Burkholderia species shows diversity among the regulatory elements, while the gene products for ornibactin synthesis are conserved. This is an interesting observation, given that ornibactin is thought to have the same defined function within Burkholderia species. Ornibactin is produced by most Burkholderia species, and its role in regulating the production of secondary metabolites should be investigated. IMPORTANCE Identification of the antibacterial product from strain MS14 is not the key feature of this study. We present a series of experiments that demonstrate that ornibactin is directly involved in the bactericidal phenotype of MS14. This observation provides evidence for an alternative function for ornibactin, aside from iron sequestration. Ornibactin should be further evaluated for its role in regulating the biosynthesis of secondary metabolites in other Burkholderia species.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2017
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 3
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 84, No. 18 ( 2018-09-15)
    Abstract: Soil fungi produce a wide range of chemical compounds and enzymes with potential for applications in medicine and biotechnology. Cellular processes in soil fungi are highly dependent on the regulation under environmentally induced stress, but most of the underlying mechanisms remain unclear. Previous work identified a key GATA-type transcription factor, Penicillium oxalicum NsdD (PoxNsdD; also called POX08415), that regulates the expression of cellulase and xylanase genes in P. oxalicum . PoxNsdD shares 57 to 64% identity with the key activator NsdD, involved in asexual development in Aspergillus . In the present study, the regulatory roles of PoxNsdD in P. oxalicum were further explored. Comparative transcriptomic profiling revealed that PoxNsdD regulates major genes involved in starch, cellulose, and hemicellulose degradation, as well as conidiation and pigment biosynthesis. Subsequent experiments confirmed that a Δ PoxNsdD strain lost 43.9 to 78.8% of starch-digesting enzyme activity when grown on soluble corn starch, and it produced 54.9 to 146.0% more conidia than the Δ PoxKu70 parental strain. During cultivation, Δ PoxNsdD cultures changed color, from pale orange to brick red, while the Δ PoxKu70 cultures remained bluish white. Real-time quantitative reverse transcription-PCR showed that PoxNsdD dynamically regulated the expression of a glucoamylase gene ( POX01356 / Amy15A ), an α-amylase gene ( POX09352 / Amy13A ), and a regulatory gene ( POX03890 / amyR ), as well as a polyketide synthase gene ( POX01430 / alb1 / wA ) for yellow pigment biosynthesis and a conidiation-regulated gene ( POX06534 / brlA ). Moreover, in vitro binding experiments showed that PoxNsdD bound the promoter regions of the above-described genes. This work provides novel insights into the regulatory mechanisms of fungal cellular processes and may assist in genetic engineering of P . oxalicum for potential industrial and medical applications. IMPORTANCE Most filamentous fungi produce a vast number of extracellular enzymes that are used commercially for biorefineries of plant biomass to produce biofuels and value-added chemicals, which might promote the transition to a more environmentally friendly economy. The expression of these extracellular enzyme genes is tightly controlled at the transcriptional level, which limits their yields. Hitherto our understanding of the regulation of expression of plant biomass-degrading enzyme genes in filamentous fungi has been rather limited. In the present study, regulatory roles of a key regulator, PoxNsdD, were further explored in the soil fungus Penicillium oxalicum , contributing to the understanding of gene regulation in filamentous fungi and revealing the biotechnological potential of P . oxalicum via genetic engineering.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2018
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
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  • 4
    Online Resource
    Online Resource
    American Society for Microbiology ; 2022
    In:  Applied and Environmental Microbiology Vol. 88, No. 9 ( 2022-05-10)
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 88, No. 9 ( 2022-05-10)
    Abstract: Isopropanol dehydrogenase (IPADH) is one of the most attractive options for nicotinamide cofactor regeneration due to its low cost and simple downstream processing. However, poor thermostability and strict cofactor dependency hinder its practical application for bioconversions. In this study, we simultaneously improved the thermostability (433-fold) and catalytic activity (3.3-fold) of IPADH from Brucella suis via a flexible segment engineering strategy. Meanwhile, the cofactor preference of IPADH was successfully switched from NAD(H) to NADP(H) by 1.23 × 10 6 -fold. When these variants were employed in three typical bioredox reactions to drive the synthesis of important chiral pharmaceutical building blocks, they outperformed the commonly used cofactor regeneration systems (glucose dehydrogenase [GDH], formate dehydrogenase [FDH] , and lactate dehydrogenase [LDH]) with respect to efficiency of cofactor regeneration. Overall, our study provides two promising IPADH variants with complementary cofactor specificities that have great potential for wide applications. IMPORTANCE Oxidoreductases represent one group of the most important biocatalysts for synthesis of various chiral synthons. However, their practical application was hindered by the expensive nicotinamide cofactors used. Isopropanol dehydrogenase (IPADH) is one of the most attractive biocatalysts for nicotinamide cofactor regeneration. However, poor thermostability and strict cofactor dependency hinder its practical application. In this work, the thermostability and catalytic activity of an IPADH were simultaneously improved via a flexible segment engineering strategy. Meanwhile, the cofactor preference of IPADH was successfully switched from NAD(H) to NADP(H). The resultant variants show great potential for regeneration of nicotinamide cofactors, and the engineering strategy might serve as a useful approach for future engineering of other oxidoreductases.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
    Location Call Number Limitation Availability
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  • 5
    Online Resource
    Online Resource
    American Society for Microbiology ; 2022
    In:  Applied and Environmental Microbiology Vol. 88, No. 9 ( 2022-05-10)
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 88, No. 9 ( 2022-05-10)
    Abstract: d -Alanine belongs to nonessential amino acids that have diverse applications in the fields of food and health care. ( R )-transaminase [( R )-TA]-catalyzed asymmetric amination of pyruvate is a feasible alternative for the synthesis of d -alanine, but low catalytic efficiency and thermostability limit enzymatic utilization. In this work, several potential ( R )-TAs were discovered using NCBI database mining synchronously with enzymatic structure-function analysis, among which Capronia epimyces TA (CeTA) showed the highest activity for amination of pyruvate using ( R )-α-methylbenzylamine as the donor. Furthermore, enzymatic residues surrounding a large catalysis pocket were subjected to saturation and combinatorial mutagenesis, and positive mutant F113T showed dramatic improvement in activity and thermostability. Molecular modeling indicated that the substitution of phenylalanine with threonine afforded alleviation of steric hindrance in the pocket and induced formation of additional hydrogen bonds with neighboring residues. Finally, using recombinant cells containing F113T as a biocatalyst, the conversion yield of amination of 100 mM pyruvate to d -alanine achieved up to 95.2%, which seemed to be the highest level in the literature regarding synthesis of d -alanine using TAs. The inherent characteristics rendered CeTA F113T a promising platform for efficient preparation of d -alanine operating with high productivity. IMPORTANCE d -Alanine is an important compound with many valuable applications. Its asymmetric synthesis employing ( R )-ω-TA is considered an attractive choice. According to the stereoselectivity, ω-TAs have either ( R )- or ( S )-enantiopreference. There has been a variety of literature regarding screening, characterizing, and molecular modification of ( S )-ω-TAs; in contrast, the research about ( R )-ω-TA has lagged behind. In this work, we identify several ( R )-ω-TAs and succeeded in creating mutant F113T, which showed not only better efficiency toward pyruvate but also higher thermostability compared with the original enzyme. The obtained original enzymes and positive mutants displayed important application value for pushing symmetric synthesis of d -alanine to a higher level.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2022
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
    Location Call Number Limitation Availability
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  • 6
    Online Resource
    Online Resource
    American Society for Microbiology ; 2015
    In:  Applied and Environmental Microbiology Vol. 81, No. 17 ( 2015-09), p. 6078-6087
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 81, No. 17 ( 2015-09), p. 6078-6087
    Abstract: The type III secretion system (T3SS) of Edwardsiella tarda plays an important role in infection by translocating effector proteins into host cells. EseB, a component required for effector translocation, is reported to mediate autoaggregation of E. tarda . In this study, we demonstrate that EseB forms filamentous appendages on the surface of E. tarda and is required for biofilm formation by E. tarda in Dulbecco's modified Eagle's medium (DMEM). Biofilm formation by E. tarda in DMEM does not require FlhB, an essential component for assembling flagella. Dynamic analysis of EseB filament formation, autoaggregation, and biofilm formation shows that the formation of EseB filaments occurs prior to autoaggregation and biofilm formation. The addition of an EseB antibody to E. tarda cultures before bacterial autoaggregation prevents autoaggregation and biofilm formation in a dose-dependent manner, whereas the addition of the EseB antibody to E. tarda cultures in which biofilm is already formed does not destroy the biofilm. Therefore, EseB filament-mediated bacterial cell-cell interaction is a prerequisite for autoaggregation and biofilm formation.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2015
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
    Location Call Number Limitation Availability
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  • 7
    In: Eukaryotic Cell, American Society for Microbiology, Vol. 4, No. 4 ( 2005-04), p. 799-813
    Abstract: Eukaryotic protein kinases are key molecules mediating signal transduction that play a pivotal role in the regulation of various biological processes, including cell cycle progression, cellular morphogenesis, development, and cellular response to environmental changes. A total of 106 eukaryotic protein kinase catalytic-domain-containing proteins have been found in the entire fission yeast genome, 44% (or 64%) of which possess orthologues (or nearest homologues) in humans, based on sequence similarity within catalytic domains. Systematic deletion analysis of all putative protein kinase-encoding genes have revealed that 17 out of 106 were essential for viability, including three previously uncharacterized putative protein kinases. Although the remaining 89 protein kinase mutants were able to form colonies under optimal growth conditions, 46% of the mutants exhibited hypersensitivity to at least 1 of the 17 different stress factors tested. Phenotypic assessment of these mutants allowed us to arrange kinases into functional groups. Based on the results of this assay, we propose also the existence of four major signaling pathways that are involved in the response to 17 stresses tested. Microarray analysis demonstrated a significant correlation between the expression signature and growth phenotype of kinase mutants tested. Our complete microarray data sets are available at http://giscompute.gis.a-star.edu.sg/∼gisljh/kinome .
    Type of Medium: Online Resource
    ISSN: 1535-9778 , 1535-9786
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2005
    detail.hit.zdb_id: 2071564-X
    SSG: 12
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  • 8
    In: Applied and Environmental Microbiology, American Society for Microbiology, Vol. 85, No. 24 ( 2019-12)
    Abstract: Transcriptional regulation of cellulolytic and xylolytic genes in ascomycete fungi is controlled by specific carbon sources in different external environments. Here, comparative transcriptomic analyses of Penicillium oxalicum grown on wheat bran (WB), WB plus rice straw (WR), or WB plus Avicel (WA) as the sole carbon source under solid-state fermentation (SSF) revealed that most of the differentially expressed genes (DEGs) were involved in metabolism, specifically, carbohydrate metabolism. Of the DEGs, the basic core carbohydrate-active enzyme-encoding genes which responded to the plant biomass resources were identified in P. oxalicum , and their transcriptional levels changed to various extents depending on the different carbon sources. Moreover, this study found that three deletion mutants of genes encoding putative transcription factors showed significant alterations in filter paper cellulase production compared with that of a parental P. oxalicum strain with a deletion of Ku70 (Δ PoxKu70 strain) when grown on WR under SSF. Importantly, the Δ PoxAtf1 mutant (with a deletion of P. oxalicum Atf1 , also called POX03016 ) displayed 46.1 to 183.2% more cellulase and xylanase production than a Δ PoxKu70 mutant after 2 days of growth on WR. RNA sequencing and quantitative reverse transcription-PCR revealed that PoxAtf1 dynamically regulated the expression of major cellulase and xylanase genes under SSF. PoxAtf1 bound to the promoter regions of the key cellulase and xylanase genes in vitro . This study provides novel insights into the regulatory mechanism of fungal cellulase and xylanase gene expression under SSF. IMPORTANCE The transition to a more environmentally friendly economy encourages studies involving the high-value-added utilization of lignocellulosic biomass. Solid-state fermentation (SSF), that simulates the natural habitat of soil microorganisms, is used for a variety of applications such as biomass biorefinery. Prior to the current study, our understanding of genome-wide gene expression and of the regulation of gene expression of lignocellulose-degrading enzymes in ascomycete fungi during SSF was limited. Here, we employed RNA sequencing and genetic analyses to investigate transcriptomes of Penicillium oxalicum strain EU2101 cultured on medium containing different carbon sources and to identify and characterize transcription factors for regulating the expression of cellulase and xylanase genes during SSF. The results generated will provide novel insights into genetic engineering of filamentous fungi to further increase enzyme production.
    Type of Medium: Online Resource
    ISSN: 0099-2240 , 1098-5336
    RVK:
    Language: English
    Publisher: American Society for Microbiology
    Publication Date: 2019
    detail.hit.zdb_id: 223011-2
    detail.hit.zdb_id: 1478346-0
    SSG: 12
    Location Call Number Limitation Availability
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