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










  • 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

  • A novel fungal beta-propeller phytase from nematophagous Arthrobotrys oligospora: characterization and potential application in phosphorus and mineral release for feed processing.

    abstract::Phytases are widely utilized in feed industry to increase the utilization of phosphorus, minerals, and amino acids for improvement of animal and human nutrition. At present, all known β-propeller phytases (BPP) have been generated from bacteria, particularly Bacillus. In this work we report for the first time a new fu...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Hou X,Shen Z,Li N,Kong X,Sheng K,Wang J,Wang Y

    更新日期:2020-04-06 00:00:00

  • Characterization, yield optimization, scale up and biopreservative potential of fermencin SA715, a novel bacteriocin from Lactobacillus fermentum GA715 of goat milk origin.

    abstract:BACKGROUND:Emergence of antibiotic resistance and growing consumer trend towards foods containing biopreservatives stimulated the search for alternative antimicrobials. This research is aimed at characterizing, investigating the mechanism of action, scale up optimization and evaluating the biopreservative potential of ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Wayah SB,Philip K

    更新日期:2018-08-13 00:00:00

  • The important ergot alkaloid intermediate chanoclavine-I produced in the yeast Saccharomyces cerevisiae by the combined action of EasC and EasE from Aspergillus japonicus.

    abstract:BACKGROUND:Ergot alkaloids are a group of highly bioactive molecules produced by a number of filamentous fungi. These compounds have been intensely studied for decades, mainly due to their deleterious effects in contaminated food and feeds, but also for their beneficial pharmaceutical and agricultural applications. Bio...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Nielsen CA,Folly C,Hatsch A,Molt A,Schröder H,O'Connor SE,Naesby M

    更新日期:2014-08-12 00:00:00

  • Improving yeast strains using recyclable integration cassettes, for the production of plant terpenoids.

    abstract:BACKGROUND:Terpenoids constitute a large family of natural products, attracting commercial interest for a variety of uses as flavours, fragrances, drugs and alternative fuels. Saccharomyces cerevisiae offers a versatile cell factory, as the precursors of terpenoid biosynthesis are naturally synthesized by the sterol bi...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Ignea C,Cvetkovic I,Loupassaki S,Kefalas P,Johnson CB,Kampranis SC,Makris AM

    更新日期:2011-01-28 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

  • Lipoteichoic acid is an important microbe-associated molecular pattern of Lactobacillus rhamnosus GG.

    abstract:BACKGROUND:Probiotic bacteria are increasingly used as immunomodulatory agents. Yet detailed molecular knowledge on the immunomodulatory molecules of these bacteria is lagging behind. Lipoteichoic acid (LTA) is considered a major microbe-associated molecular pattern (MAMP) of Gram-positive bacteria. However, many detai...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Claes IJ,Segers ME,Verhoeven TL,Dusselier M,Sels BF,De Keersmaecker SC,Vanderleyden J,Lebeer S

    更新日期:2012-12-15 00:00:00

  • Production of Nα-acetylated thymosin α1 in Escherichia coli.

    abstract:BACKGROUND:Thymosin α1 (Tα1), a 28-amino acid Nα-acetylated peptide, has a powerful general immunostimulating activity. Although biosynthesis is an attractive means of large-scale manufacture, to date, Tα1 can only be chemosynthesized because of two obstacles to its biosynthesis: the difficulties in expressing small pe...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Ren Y,Yao X,Dai H,Li S,Fang H,Chen H,Zhou C

    更新日期:2011-04-22 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

  • 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

  • Modeling and measuring intracellular fluxes of secreted recombinant protein in Pichia pastoris with a novel 34S labeling procedure.

    abstract:BACKGROUND:The budding yeast Pichia pastoris is widely used for protein production. To determine the best suitable strategy for strain improvement, especially for high secretion, quantitative data of intracellular fluxes of recombinant protein are very important. Especially the balance between intracellular protein for...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Pfeffer M,Maurer M,Köllensperger G,Hann S,Graf AB,Mattanovich D

    更新日期:2011-06-26 00:00:00

  • A cell factory of Bacillus subtilis engineered for the simple bioconversion of myo-inositol to scyllo-inositol, a potential therapeutic agent for Alzheimer's disease.

    abstract:BACKGROUND:A stereoisomer of inositol, scyllo-inositol, is known as a promising therapeutic agent for Alzheimer's disease, since it prevents the accumulation of beta-amyloid deposits, a hallmark of the disease. However, this compound is relatively rare in nature, whereas another stereoisomer of inositol, myo-inositol, ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Yamaoka M,Osawa S,Morinaga T,Takenaka S,Yoshida K

    更新日期:2011-09-07 00:00:00

  • Heterologous production of raspberry ketone in the wine yeast Saccharomyces cerevisiae via pathway engineering and synthetic enzyme fusion.

    abstract:BACKGROUND:Raspberry ketone is the primary aroma compound found in raspberries and naturally derived raspberry ketone is a valuable flavoring agent. The economic incentives for the production of raspberry ketone, combined with the very poor yields from plant tissue, therefore make this compound an excellent target for ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Lee D,Lloyd ND,Pretorius IS,Borneman AR

    更新日期:2016-03-04 00:00:00

  • Genome-wide identification of Saccharomyces cerevisiae genes required for tolerance to acetic acid.

    abstract:BACKGROUND:Acetic acid is a byproduct of Saccharomyces cerevisiae alcoholic fermentation. Together with high concentrations of ethanol and other toxic metabolites, acetic acid may contribute to fermentation arrest and reduced ethanol productivity. This weak acid is also a present in lignocellulosic hydrolysates, a high...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Mira NP,Palma M,Guerreiro JF,Sá-Correia I

    更新日期:2010-10-25 00:00:00

  • Development of novel surface display platforms for anchoring heterologous proteins in Saccharomyces cerevisiae.

    abstract:BACKGROUND:Cell surface display of recombinant proteins has become a powerful tool for biotechnology and biomedical applications. As a model eukaryotic microorganism, Saccharomyces cerevisiae is an ideal candidate for surface display of heterologous proteins. However, the frequently used commercial yeast surface displa...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Yang X,Tang H,Song M,Shen Y,Hou J,Bao X

    更新日期:2019-05-18 00:00:00

  • A new-generation of Bacillus subtilis cell factory for further elevated scyllo-inositol production.

    abstract:BACKGROUND:A stereoisomer of inositol, scyllo-inositol (SI), has been regarded as a promising therapeutic agent for Alzheimer's disease. However, this compound is relatively rare, whereas another stereoisomer of inositol, myo-inositol (MI) is abundant in nature. Bacillus subtilis 168 has the ability to metabolize inosi...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Tanaka K,Natsume A,Ishikawa S,Takenaka S,Yoshida KI

    更新日期:2017-04-21 00:00:00

  • Metabolic engineering of Escherichia coli for efficient production of L-alanyl-L-glutamine.

    abstract:BACKGROUND:L-Alanyl-L-glutamine (AQ) is a functional dipeptide with high water solubility, good thermal stability and high bioavailability. It is widely used in clinical treatment, post-operative rehabilitation, sports health care and other fields. AQ is mainly produced via chemical synthesis which is complicated, time...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Zhu J,Yang W,Wang B,Liu Q,Zhong X,Gao Q,Liu J,Huang J,Lin B,Tao Y

    更新日期:2020-06-11 00:00:00

  • Increased expression of the yeast multidrug resistance ABC transporter Pdr18 leads to increased ethanol tolerance and ethanol production in high gravity alcoholic fermentation.

    abstract:BACKGROUND:The understanding of the molecular basis of yeast tolerance to ethanol may guide the design of rational strategies to increase process performance in industrial alcoholic fermentations. A set of 21 genes encoding multidrug transporters from the ATP-Binding Cassette (ABC) Superfamily and Major Facilitator Sup...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Teixeira MC,Godinho CP,Cabrito TR,Mira NP,Sá-Correia I

    更新日期:2012-07-27 00:00:00

  • Altering the ribosomal subunit ratio in yeast maximizes recombinant protein yield.

    abstract:BACKGROUND:The production of high yields of recombinant proteins is an enduring bottleneck in the post-genomic sciences that has yet to be addressed in a truly rational manner. Typically eukaryotic protein production experiments have relied on varying expression construct cassettes such as promoters and tags, or cultur...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Bonander N,Darby RA,Grgic L,Bora N,Wen J,Brogna S,Poyner DR,O'Neill MA,Bill RM

    更新日期:2009-01-29 00:00:00

  • Invertase Suc2-mediated inulin catabolism is regulated at the transcript level in Saccharomyces cerevisiae.

    abstract:BACKGROUND:Invertase Suc2 was recently identified as a key hydrolase for inulin catabolism in Saccharomyces cerevisiae, whereas the Suc2 activity degrading inulin varies greatly in different S. cerevisiae strains. The molecular mechanism causing such variation remained obscure. The aim of this study is to investigate h...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Yang F,Liu ZC,Wang X,Li LL,Yang L,Tang WZ,Yu ZM,Li X

    更新日期:2015-04-17 00:00:00

  • One-step of tryptophan attenuator inactivation and promoter swapping to improve the production of L-tryptophan in Escherichia coli.

    abstract:BACKGROUND:L-tryptophan is an aromatic amino acid widely used in the food, chemical and pharmaceutical industries. In Escherichia coli, L-tryptophan is synthesized from phosphoenolpyruvate and erythrose 4-phosphate by enzymes in the shikimate pathway and L-tryptophan branch pathway, while L-serine and phosphoribosylpyr...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Gu P,Yang F,Kang J,Wang Q,Qi Q

    更新日期:2012-03-02 00:00:00

  • Development of a mixed feed strategy for a recombinant Pichia pastoris strain producing with a de-repression promoter.

    abstract:BACKGROUND:Recombinant protein production in the yeast Pichia pastoris is usually based on the alcohol oxidase promoters pAOX1 and pAOX2, which are regulated by methanol and strongly repressed by other C-sources, like glycerol and glucose. However, the use of methanol brings several disadvantages, which is why current ...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Capone S,Horvat J,Herwig C,Spadiut O

    更新日期:2015-07-10 00:00:00

  • Identification and utilization of two important transporters: SgvT1 and SgvT2, for griseoviridin and viridogrisein biosynthesis in Streptomyces griseoviridis.

    abstract:BACKGROUND:Griseoviridin (GV) and viridogrisein (VG, also referred as etamycin), both biosynthesized by a distinct 105 kb biosynthetic gene cluster (BGC) in Streptomyces griseoviridis NRRL 2427, are a pair of synergistic streptogramin antibiotics and very important in treating infections of many multi-drug resistant mi...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Xie Y,Ma J,Qin X,Li Q,Ju J

    更新日期:2017-10-25 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

  • Redirecting photosynthetic electron flux in the cyanobacterium Synechocystis sp. PCC 6803 by the deletion of flavodiiron protein Flv3.

    abstract:BACKGROUND:Oxygen-evolving photoautotrophic organisms, like cyanobacteria, protect their photosynthetic machinery by a number of regulatory mechanisms, including alternative electron transfer pathways. Despite the importance in modulating the electron flux distribution between the photosystems, alternative electron tra...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Thiel K,Patrikainen P,Nagy C,Fitzpatrick D,Pope N,Aro EM,Kallio P

    更新日期:2019-11-05 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

  • Overexpression of a C4-dicarboxylate transporter is the key for rerouting citric acid to C4-dicarboxylic acid production in Aspergillus carbonarius.

    abstract:BACKGROUND:C4-dicarboxylic acids, including malic acid, fumaric acid and succinic acid, are valuable organic acids that can be produced and secreted by a number of microorganisms. Previous studies on organic acid production by Aspergillus carbonarius, which is capable of producing high amounts of citric acid from varie...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Yang L,Christakou E,Vang J,Lübeck M,Lübeck PS

    更新日期:2017-03-14 00:00:00

  • Easy to use and reliable technique for online dissolved oxygen tension measurement in shake flasks using infrared fluorescent oxygen-sensitive nanoparticles.

    abstract:BACKGROUND:Despite the progressive miniaturization of bioreactors for screening purposes, shake flasks are still widespread in biotechnological laboratories and industry as cultivation vessels. Shake flasks are often applied as the first or second step in applications such as strain screening or media optimization. Thu...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Flitsch D,Ladner T,Lukacs M,Büchs J

    更新日期:2016-02-24 00:00:00

  • Engineering cofactor metabolism for improved protein and glucoamylase production in Aspergillus niger.

    abstract:BACKGROUND:Nicotinamide adenine dinucleotide phosphate (NADPH) is an important cofactor ensuring intracellular redox balance, anabolism and cell growth in all living systems. Our recent multi-omics analyses of glucoamylase (GlaA) biosynthesis in the filamentous fungal cell factory Aspergillus niger indicated that low a...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Sui YF,Schütze T,Ouyang LM,Lu H,Liu P,Xiao X,Qi J,Zhuang YP,Meyer V

    更新日期:2020-10-23 00:00:00

  • Metabolic regulation of Escherichia coli and its gdhA, glnL, gltB, D mutants under different carbon and nitrogen limitations in the continuous culture.

    abstract:BACKGROUND:It is quite important to understand how the central metabolism is regulated under nitrogen (N)- limitation as well as carbon (C)- limitation. In particular, the effect of C/N ratio on the metabolism is of practical interest for the heterologous protein production, PHB production, etc. Although the carbon and...

    journal_title:Microbial cell factories

    pub_type: 杂志文章


    authors: Kumar R,Shimizu K

    更新日期:2010-01-27 00:00:00