Abstract:
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.
journal_name
Microb Cell Factjournal_title
Microbial cell factoriesauthors
Liu Y,Koh CMJ,Ngoh ST,Ji Ldoi
10.1186/s12934-015-0357-7subject
Has Abstractpub_date
2015-10-26 00:00:00pages
170issn
1475-2859pii
10.1186/s12934-015-0357-7journal_volume
14pub_type
杂志文章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
doi:10.1186/1475-2859-7-11
更新日期:2008-04-04 00:00:00
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...
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更新日期:2014-08-12 00:00:00
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
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doi:10.1186/1475-2859-10-4
更新日期:2011-01-28 00:00:00
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...
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更新日期:2015-11-05 00:00:00
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
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doi:10.1186/1475-2859-11-161
更新日期:2012-12-15 00:00:00
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
doi:10.1186/1475-2859-10-26
更新日期:2011-04-22 00:00:00
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
doi:10.1186/1475-2859-11-89
更新日期:2012-06-25 00:00:00
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journal_title:Microbial cell factories
pub_type: 杂志文章
doi:10.1186/s12934-016-0527-2
更新日期:2016-07-28 00:00:00
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
doi:10.1186/1475-2859-10-47
更新日期:2011-06-26 00:00:00
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
doi:10.1186/1475-2859-10-69
更新日期:2011-09-07 00:00:00
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
doi:10.1186/s12934-016-0446-2
更新日期:2016-03-04 00:00:00
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
doi:10.1186/1475-2859-9-79
更新日期:2010-10-25 00:00:00
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
doi:10.1186/s12934-019-1133-x
更新日期:2019-05-18 00:00:00
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
doi:10.1186/s12934-017-0682-0
更新日期:2017-04-21 00:00:00
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
doi:10.1186/s12934-020-01369-2
更新日期:2020-06-11 00:00:00
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
doi:10.1186/1475-2859-11-98
更新日期:2012-07-27 00:00:00
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...
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doi:10.1186/1475-2859-8-10
更新日期:2009-01-29 00:00:00
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journal_title:Microbial cell factories
pub_type: 杂志文章
doi:10.1186/s12934-015-0243-3
更新日期:2015-04-17 00:00:00
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
doi:10.1186/1475-2859-11-30
更新日期:2012-03-02 00:00:00
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journal_title:Microbial cell factories
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doi:10.1186/s12934-015-0292-7
更新日期:2015-07-10 00:00:00
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
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更新日期:2017-10-25 00:00:00
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
doi:10.1186/1475-2859-10-36
更新日期:2011-05-18 00:00:00
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
doi:10.1186/s12934-019-1238-2
更新日期:2019-11-05 00:00:00
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
doi:10.1186/s12934-020-01454-6
更新日期:2020-10-19 00:00:00
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
doi:10.1186/s12934-017-0660-6
更新日期:2017-03-14 00:00:00
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
doi:10.1186/s12934-016-0444-4
更新日期:2016-02-24 00:00:00
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
doi:10.1186/s12934-020-01450-w
更新日期:2020-10-23 00:00:00
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
doi:10.1186/1475-2859-9-8
更新日期:2010-01-27 00:00:00