Engineering an efficient and tight D-amino acid-inducible gene expression system in Rhodosporidium/Rhodotorula species.


BACKGROUND:Rhodosporidium and Rhodotorula are two genera of oleaginous red yeast with great potential for industrial biotechnology. To date, there is no effective method for inducible expression of proteins and RNAs in these hosts. RESULTS:We have developed a luciferase gene reporter assay based on a new codon-optimized LUC2 reporter gene (RtLUC2), which is flanked with CAR2 homology arms and can be integrated into the CAR2 locus in the nuclear genome at >90 % efficiency. We characterized the upstream DNA sequence of a D-amino acid oxidase gene (DAO1) from R. toruloides ATCC 10657 by nested deletions. By comparing the upstream DNA sequences of several putative DAO1 homologs of Basidiomycetous fungi, we identified a conserved DNA motif with a consensus sequence of AGGXXGXAGX11GAXGAXGG within a 0.2 kb region from the mRNA translation initiation site. Deletion of this motif led to strong mRNA transcription under non-inducing conditions. Interestingly, DAO1 promoter activity was enhanced about fivefold when the 108 bp intron 1 was included in the reporter construct. We identified a conserved CT-rich motif in the intron with a consensus sequence of TYTCCCYCTCCYCCCCACWYCCGA, deletion or point mutations of which drastically reduced promoter strength under both inducing and non-inducing conditions. Additionally, we created a selection marker-free DAO1-null mutant (∆dao1e) which displayed greatly improved inducible gene expression, particularly when both glucose and nitrogen were present in high levels. To avoid adding unwanted peptide to proteins to be expressed, we converted the original translation initiation codon to ATC and re-created a translation initiation codon at the start of exon 2. This promoter, named P DAO1-in1m1 , showed very similar luciferase activity to the wild-type promoter upon induction with D-alanine. The inducible system was tunable by adjusting the levels of inducers, carbon source and nitrogen source. CONCLUSION:The intron 1-containing DAO1 promoters coupled with a DAO1 null mutant makes an efficient and tight D-amino acid-inducible gene expression system in Rhodosporidium and Rhodotorula genera. The system will be a valuable tool for metabolic engineering and enzyme expression in these yeast hosts.


Microb Cell Fact


Microbial cell factories


Liu Y,Koh CMJ,Ngoh ST,Ji L




Has Abstract


2015-10-26 00:00:00










  • Tailoring nutritional and process variables for hyperproduction of catalase from a novel isolated bacterium Geobacillus sp. BSS-7.

    abstract:BACKGROUND:Catalase (EC is one of the important industrial enzyme employed in diagnostic and analytical methods in the form of biomarkers and biosensors in addition to their enormous applications in textile, paper, food and pharmaceutical sectors. The present study demonstrates the utility of a newly isolated...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Kauldhar BS,Sooch BS

    更新日期:2016-01-14 00:00:00

  • Rewiring a secondary metabolite pathway towards itaconic acid production in Aspergillus niger.

    abstract:BACKGROUND:The industrially relevant filamentous fungus Aspergillus niger is widely used in industry for its secretion capabilities of enzymes and organic acids. Biotechnologically produced organic acids promise to be an attractive alternative for the chemical industry to replace petrochemicals. Itaconic acid (IA) has ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Hossain AH,Li A,Brickwedde A,Wilms L,Caspers M,Overkamp K,Punt PJ

    更新日期:2016-07-28 00:00:00

  • Protein folding and conformational stress in microbial cells producing recombinant proteins: a host comparative overview.

    abstract:: Different species of microorganisms including yeasts, filamentous fungi and bacteria have been used in the past 25 years for the controlled production of foreign proteins of scientific, pharmacological or industrial interest. A major obstacle for protein production processes and a limit to overall success has been th...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Gasser B,Saloheimo M,Rinas U,Dragosits M,Rodríguez-Carmona E,Baumann K,Giuliani M,Parrilli E,Branduardi P,Lang C,Porro D,Ferrer P,Tutino ML,Mattanovich D,Villaverde A

    更新日期:2008-04-04 00:00:00

  • Transcriptome-enabled discovery and functional characterization of enzymes related to (2S)-pinocembrin biosynthesis from Ornithogalum caudatum and their application for metabolic engineering.

    abstract:BACKGROUND:(2S)-Pinocembrin is a chiral flavanone with versatile pharmacological and biological activities. Its health-promoting effects have spurred on research effects on the microbial production of (2S)-pinocembrin. However, an often-overlooked salient feature in the analysis of microbial (2S)-pinocembrin is its chi...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Guo L,Chen X,Li LN,Tang W,Pan YT,Kong JQ

    更新日期:2016-02-04 00:00:00

  • Engineering Corynebacterium glutamicum for violacein hyper production.

    abstract:BACKGROUND:Corynebacterium glutamicum was used as a metabolic engineering chassis for production of crude violacein (mixture of violacein and deoxyviolacein) due to Corynebacterium's GRAS status and advantages in tryptophan fermentation. The violacein is a commercially potential compound with various applications deriv...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Sun H,Zhao D,Xiong B,Zhang C,Bi C

    更新日期:2016-08-24 00:00:00

  • Genetic manipulation of longevity-related genes as a tool to regulate yeast life span and metabolite production during winemaking.

    abstract:BACKGROUND:Yeast viability and vitality are essential for different industrial processes where the yeast Saccharomyces cerevisiae is used as a biotechnological tool. Therefore, the decline of yeast biological functions during aging may compromise their successful biotechnological use. Life span is controlled by a varie...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Orozco H,Matallana E,Aranda A

    更新日期:2013-01-02 00:00:00

  • Relative contributions of non-essential Sec pathway components and cell envelope-associated proteases to high-level enzyme secretion by Bacillus subtilis.

    abstract:BACKGROUND:Bacillus subtilis is an important industrial workhorse applied in the production of many different commercially relevant proteins, especially enzymes. Virtually all of these proteins are secreted via the general secretion (Sec) pathway. Studies from different laboratories have demonstrated essential or non-e...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Neef J,Bongiorni C,Schmidt B,Goosens VJ,van Dijl JM

    更新日期:2020-02-28 00:00:00

  • Improved triacylglycerol production in Acinetobacter baylyi ADP1 by metabolic engineering.

    abstract:BACKGROUND:Triacylglycerols are used in various purposes including food applications, cosmetics, oleochemicals and biofuels. Currently the main sources for triacylglycerol are vegetable oils, and microbial triacylglycerol has been suggested as an alternative for these. Due to the low production rates and yields of micr...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Santala S,Efimova E,Kivinen V,Larjo A,Aho T,Karp M,Santala V

    更新日期:2011-05-18 00:00:00

  • MAL62 overexpression enhances uridine diphosphoglucose-dependent trehalose synthesis and glycerol metabolism for cryoprotection of baker's yeast in lean dough.

    abstract:BACKGROUND:In Saccharomyces cerevisiae, alpha-glucosidase (maltase) is a key enzyme in maltose metabolism. In addition, the overexpression of the alpha-glucosidase-encoding gene MAL62 has been shown to increase the freezing tolerance of yeast in lean dough. However, its cryoprotection mechanism is still not clear. RES...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Sun X,Zhang J,Fan ZH,Xiao P,Li F,Liu HQ,Zhu WB

    更新日期:2020-10-19 00:00:00

  • Efficient ammonium uptake and mobilization of vacuolar arginine by Saccharomyces cerevisiae wine strains during wine fermentation.

    abstract:BACKGROUND:Under N-limiting conditions, Saccharomyces cerevisiae strains display a substantial variability in their biomass yield from consumed nitrogen -in particular wine yeasts exhibit high growth abilities- that is correlated with their capacity to complete alcoholic fermentation, a trait of interest for fermented ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Crépin L,Sanchez I,Nidelet T,Dequin S,Camarasa C

    更新日期:2014-08-19 00:00:00

  • A novel prokaryotic vector for identification and selection of recombinants: direct use of the vector for expression studies in E. coli.

    abstract:BACKGROUND:The selection of bacterial recombinants that harbour a desired insert, has been a key factor in molecular cloning and a series of screening procedures need to be performed for selection of clones carrying the genes of interest. The conventional cloning techniques are reported to have problems such as screeni...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Banerjee S,Kumar J,Apte-Deshpande A,Padmanabhan S

    更新日期:2010-05-11 00:00:00

  • Rational selection and engineering of exogenous principal sigma factor (σ(HrdB)) to increase teicoplanin production in an industrial strain of Actinoplanes teichomyceticus.

    abstract:BACKGROUND:Transcriptional engineering has presented a strong ability of phenotypic improvement in microorganisms. However, it could not be directly applied to Actinoplanes teichomyceticus L-27 because of the paucity of endogenous transcription factors in the strain. In this study, exogenous transcription factors were ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Wang H,Yang L,Wu K,Li G

    更新日期:2014-01-16 00:00:00

  • Fermentation of mixed glucose-xylose substrates by engineered strains of Saccharomyces cerevisiae: role of the coenzyme specificity of xylose reductase, and effect of glucose on xylose utilization.

    abstract:BACKGROUND:In spite of the substantial metabolic engineering effort previously devoted to the development of Saccharomyces cerevisiae strains capable of fermenting both the hexose and pentose sugars present in lignocellulose hydrolysates, the productivity of reported strains for conversion of the naturally most abundan...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Krahulec S,Petschacher B,Wallner M,Longus K,Klimacek M,Nidetzky B

    更新日期:2010-03-10 00:00:00

  • Enhanced production of taxadiene in Saccharomyces cerevisiae.

    abstract:BACKGROUND:Cost-effective production of the highly effective anti-cancer drug, paclitaxel (Taxol®), remains limited despite growing global demands. Low yields of the critical taxadiene precursor remains a key bottleneck in microbial production. In this study, the key challenge of poor taxadiene synthase (TASY) solubili...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Nowrouzi B,Li RA,Walls LE,d'Espaux L,Malcı K,Liang L,Jonguitud-Borrego N,Lerma-Escalera AI,Morones-Ramirez JR,Keasling JD,Rios-Solis L

    更新日期:2020-11-02 00:00:00

  • Surface components and metabolites of probiotics for regulation of intestinal epithelial barrier.

    abstract::The gut microbiota can significantly affect the function of the intestinal barrier. Some intestinal probiotics (such as Lactobacillus, Bifidobacteria, a few Escherichia coli strains, and a new generation of probiotics including Bacteroides thetaiotaomicron and Akkermansia muciniphila) can maintain intestinal epithelia...

    journal_title:Microbial cell factories

    pub_type: 杂志文章,评审


    authors: Liu Q,Yu Z,Tian F,Zhao J,Zhang H,Zhai Q,Chen W

    更新日期:2020-02-05 00:00:00

  • Development of a GIN11/FRT-based multiple-gene integration technique affording inhibitor-tolerant, hemicellulolytic, xylose-utilizing abilities to industrial Saccharomyces cerevisiae strains for ethanol production from undetoxified lignocellulosic hemicel

    abstract:BACKGROUND:Bioethanol produced by the yeast Saccharomyces cerevisiae is currently one of the most promising alternatives to conventional transport fuels. Lignocellulosic hemicelluloses obtained after hydrothermal pretreatment are important feedstock for bioethanol production. However, hemicellulosic materials cannot be...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Hasunuma T,Hori Y,Sakamoto T,Ochiai M,Hatanaka H,Kondo A

    更新日期:2014-10-12 00:00:00

  • Expression of the Aspergillus terreus itaconic acid biosynthesis cluster in Aspergillus niger.

    abstract:BACKGROUND:Aspergillus terreus is a natural producer of itaconic acid and is currently used to produce itaconic acid on an industrial scale. The metabolic process for itaconic acid biosynthesis is very similar to the production of citric acid in Aspergillus niger. However, a key enzyme in A. niger, cis-aconitate decarb...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: van der Straat L,Vernooij M,Lammers M,van den Berg W,Schonewille T,Cordewener J,van der Meer I,Koops A,de Graaff LH

    更新日期:2014-01-17 00:00:00

  • Yeast prions form infectious amyloid inclusion bodies in bacteria.

    abstract:BACKGROUND:Prions were first identified as infectious proteins associated with fatal brain diseases in mammals. However, fungal prions behave as epigenetic regulators that can alter a range of cellular processes. These proteins propagate as self-perpetuating amyloid aggregates being an example of structural inheritance...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Espargaró A,Villar-Piqué A,Sabaté R,Ventura S

    更新日期:2012-06-25 00:00:00

  • Use of chitin and chitosan to produce new chitooligosaccharides by chitinase Chit42: enzymatic activity and structural basis of protein specificity.

    abstract:BACKGROUND:Chitinases are ubiquitous enzymes that have gained a recent biotechnological attention due to their ability to transform biological waste from chitin into valued chito-oligomers with wide agricultural, industrial or medical applications. The biological activity of these molecules is related to their size and...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Kidibule PE,Santos-Moriano P,Jiménez-Ortega E,Ramírez-Escudero M,Limón MC,Remacha M,Plou FJ,Sanz-Aparicio J,Fernández-Lobato M

    更新日期:2018-03-22 00:00:00

  • A review on Lactococcus lactis: from food to factory.

    abstract::Lactococcus lactis has progressed a long way since its discovery and initial use in dairy product fermentation, to its present biotechnological applications in genetic engineering for the production of various recombinant proteins and metabolites that transcends the heterologous species barrier. Key desirable features...

    journal_title:Microbial cell factories

    pub_type: 杂志文章,评审


    authors: Song AA,In LLA,Lim SHE,Rahim RA

    更新日期:2017-04-04 00:00:00

  • Engineering E. coli-E. coli cocultures for production of muconic acid from glycerol.

    abstract:BACKGROUND:cis, cis-Muconic acid is an important chemical that can be biosynthesized from simple substrates in engineered microorganisms. Recently, it has been shown that engineering microbial cocultures is an emerging and promising approach for biochemical production. In this study, we aim to explore the potential of ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Zhang H,Li Z,Pereira B,Stephanopoulos G

    更新日期:2015-09-15 00:00:00

  • Identification, characterization of two NADPH-dependent erythrose reductases in the yeast Yarrowia lipolytica and improvement of erythritol productivity using metabolic engineering.

    abstract:BACKGROUND:Erythritol is a four-carbon sugar alcohol with sweetening properties that is used by the agro-food industry as a food additive. In the yeast Yarrowia lipolytica, the last step of erythritol synthesis involves the reduction of erythrose by specific erythrose reductase(s). In the earlier report, an erythrose r...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Cheng H,Wang S,Bilal M,Ge X,Zhang C,Fickers P,Cheng H

    更新日期:2018-08-29 00:00:00

  • Significantly improved solvent tolerance of Escherichia coli by global transcription machinery engineering.

    abstract:BACKGROUND:Escherichia coli has emerged as a promising platform microorganism to produce biofuels and fine chemicals of industrial interests. Certain obstacles however remain to be overcome, among which organic-solvent tolerance is a crucial one. RESULTS:We used global transcription machinery engineering (gTME) to imp...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Zhang F,Qian X,Si H,Xu G,Han R,Ni Y

    更新日期:2015-11-05 00:00:00

  • Cocktail delta-integration: a novel method to construct cellulolytic enzyme expression ratio-optimized yeast strains.

    abstract:BACKGROUND:The filamentous fungus T. reesei effectively degrades cellulose and is known to produce various cellulolytic enzymes such as beta-glucosidase, endoglucanase, and cellobiohydrolase. The expression levels of each cellulase are controlled simultaneously, and their ratios and synergetic effects are important for...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Yamada R,Taniguchi N,Tanaka T,Ogino C,Fukuda H,Kondo A

    更新日期:2010-05-14 00:00:00

  • Mevalonate production from ethanol by direct conversion through acetyl-CoA using recombinant Pseudomonas putida, a novel biocatalyst for terpenoid production.

    abstract:BACKGROUND:Bioethanol is one of the most representative eco-friendly fuels developed to replace the non-renewable fossil fuels and is the most successful commercially available bio-conversion technology till date. With the availability of inexpensive carbon sources, such as cellulosic biomass, bioethanol production has...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Yang J,Son JH,Kim H,Cho S,Na JG,Yeon YJ,Lee J

    更新日期:2019-10-10 00:00:00

  • Purification and characterization of a novel cold adapted fungal glucoamylase.

    abstract:BACKGROUND:Amylases are used in various industrial processes and a key requirement for the efficiency of these processes is the use of enzymes with high catalytic activity at ambient temperature. Unfortunately, most amylases isolated from bacteria and filamentous fungi have optimal activity above 45 °C and low pH. For ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Carrasco M,Alcaíno J,Cifuentes V,Baeza M

    更新日期:2017-05-02 00:00:00

  • Evaluation of GFP reporter utility for analysis of transcriptional slippage during gene expression.

    abstract:BACKGROUND:Epimutations arising from transcriptional slippage seem to have more important role in regulating gene expression than earlier though. Since the level and the fidelity of transcription primarily determine the overall efficiency of gene expression, all factors contributing to their decrease should be identifi...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Wons E,Koscielniak D,Szadkowska M,Sektas M

    更新日期:2018-09-21 00:00:00

  • Coiled-coil inspired functional inclusion bodies.

    abstract:BACKGROUND:Recombinant protein expression in bacteria often leads to the formation of intracellular insoluble protein deposits, a major bottleneck for the production of soluble and active products. However, in recent years, these bacterial protein aggregates, commonly known as inclusion bodies (IBs), have been shown to...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Gil-Garcia M,Navarro S,Ventura S

    更新日期:2020-06-01 00:00:00

  • Enzymatic improvement of mitochondrial thiol oxidase Erv1 for oxidized glutathione fermentation by Saccharomyces cerevisiae.

    abstract:BACKGROUND:Oxidized glutathione (GSSG) is the preferred form for industrial mass production of glutathione due to its high stability compared with reduced glutathione (GSH). In our previous study, over-expression of the mitochondrial thiol oxidase ERV1 gene was the most effective for high GSSG production in Saccharomyc...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Kobayashi J,Sasaki D,Hara KY,Hasunuma T,Kondo A

    更新日期:2017-03-15 00:00:00

  • A step ahead: combining protein purification and correct folding selection.

    abstract::The success of recombinant protein expression seems unpredictable and even good yields of soluble proteins do not guarantee the correct folding. The search for soluble constructs can be performed by exploiting libraries and speeded up by automation, but these approaches are money and time consuming and the tags used f...

    journal_title:Microbial cell factories

    pub_type: 社论


    authors: de Marco A

    更新日期:2004-10-07 00:00:00