41 results about "Geranyl pyrophosphate" patented technology

The invention provides a method for biosynthesizing menadione by using yarrowia lipolytica. The method is realized by constructing recombinant yarrowia lipolytica, and the construction method of the recombinant yarrowia lipolytica for synthesizing menadione comprises the following steps: intensively expressing a mevalonic acid pathway in the yarrowia lipolytica, and synthesizing menadione by introducing a synthesis pathway of heterologous geranyl and geranyl pyrophosphate. A sufficient isoprene side chain donor is provided for biosynthesis of menadione; on the basis, an aromatic isopentenyl transferase coding gene which is optimized by codons and has menadione substrate specificity is over-expressed in yarrowia lipolytica. The construction method of the yarrowia lipolytica for biosynthesis of menadione provided by the invention is simple to operate, the constructed yarrowia lipolytica can obtain a menadione product with relatively high concentration by taking commercially available menadione as a substrate, the biosynthesis steps are relatively short, and the yarrowia lipolytica has relatively high production and application values.

The invention provides a metabolic engineering transformation method of high-yield coenzyme Q10 of rhodobacter sphaeroides and application of transformed engineering bacteria. The method comprises the following steps: knocking out geranyl and geranyl pyrophosphate synthase gene crtE participating in a carotenoid synthesis route; performing overexpression on a farnesyl pyrophosphate synthase gene ispA; a histidine kinase coding gene prrB is mutated, and the mutant is introduced. According to the invention, a combined regulation strategy suitable for targeted regulation is found, the metabolic flux synthesized by the coenzyme Q10 is accurately optimized, and the capability of producing the coenzyme Q10 by the rhodobacter sphaeroides is greatly improved.

The application discloses recombinant escherichia coli for producing isopulegol and a construction method, and the construction method is as follows: a plasmid 1 is obtained by replacing an expression module of pETDuet-1 with an EM nucleotide fragment; a plasmid 2 is obtained by replacing an expression module of pRSFDuet-1 with the EM nucleotide fragment; an optimized geraniol synthase gene ObGES is integrated into the plasmid 1, and an optimized geranylgeranyl diphosphate synthase gene AgGPPS is integrated into the plasmid 1 to obtain a plasmid 3; an optimized EcGeDH is integrated into the plasmid 3 to obtain a plasmid 4; an optimized SsOYE2.6 W78Y / C113I is integrated into the plasmid 2, and an optimized ZmSHC F486C is integrated into the plasmid 2 to obtain a plasmid 5; the plasmids 4 and 5 are introduced into escherichia coli to obtain the recombinant escherichia coli for producing isopulegol; and the recombinant escherichia coli of the application can obtain isopulegol after fermentation.

The invention provides a method for detecting geranyl pyrophosphoric acid in water by headspace solid-phase microextraction / gas chromatography-mass spectrometry, which utilizes the characteristic that geranyl pyrophosphoric acid is easy to decompose into linalool under an acidic condition, adsorbs linalool by headspace solid-phase microextraction, and indirectly quantifies geranyl pyrophosphoric acid in combination with a gas chromatography-mass spectrometry technology. The method is easy to operate, low in instrument cost, high in sensitivity and suitable for detecting geranyl pyrophosphoric acid in water bodies such as source water, the generation risk of 2-methylisoborneol can be pre-judged through the content of geranyl pyrophosphoric acid, and technical support is provided for water supply safety.

The invention relates to an aerobic methanophilic engineering bacterium for producing taxadiene as well as a construction method and application of the aerobic methanophilic engineering bacterium. A strong promoter P16200 is used for expressing a taxadiene synthase gene TXS in host bacteria; 1-deoxy-xylulose-5-phosphate synthase genes dxs1 and dxs2, geranyl pyrophosphate synthase gene ispA and isopentenyl pyrophosphate isomerase gene Bsidi from bacillus subtilis in a taxadiene synthesis route are overexpressed, the metabolic flux of the route in which a target product is located is improved, meanwhile, 6-glucose phosphate dehydrogenase zwf1 is overexpressed, supply of NADPH (nicotinamide adenine dinucleotide phosphate) is enhanced, and the yield of the target product is increased. Therefore, the material flow and the reducing power are synergistically enhanced; bioconversion from methane to taxadiene is achieved for the first time, the yield of the taxadiene of the constructed engineering methanophilic bacteria is obviously increased, the engineering methanophilic bacteria have industrial application prospects, and a new way is provided for efficient synthesis of terpenoids.

The invention belongs to but is not limited to the technical field of plant genetic engineering, and discloses a Nerolidol / Linalool synthetase gene MnTPS6 of machilus rubescens and an application of the Nerolidol / Linalool synthetase gene MnTPS6. The machilus nerolidol / Linalool synthetase gene MnTPS6 is a key gene for synthesis of the nerolidol / Linalool in plant genetic engineering, synthesis of the nerolidol / Linalool can be effectively regulated and controlled, expression protein of the machilus nerolidol / Linalool synthetase gene MnTPS6 is applied to synthesis and preparation of the nerolidol / Linalool, the yield is high, and the machilus nerolidol / Linalool synthetase gene MnTPS6 has important application value. Substrates of farnesyl pyrophosphate (FPP) and geranyl pyrophosphate (GPP) are added to the expression protein of the induced machilus nigrum Nerolidol / Linalool synthetase gene MnTPS6, the catalytic activity of the recombinant protein is detected, and a GC-MS (Gas Chromatography-Mass Spectrometer) detection component shows that the result shows that the MnTPS6 is a bifunctional enzyme for synthesizing Nerolidol / Linalool.

The present invention relates to a protein, comprising the following 1)-3): 1) an amino acid sequence as shown in SEQ ID NO.1; 2) a sequence having the same function as the sequence as shown in SEQ ID NO.1 and obtained by replacing, deleting or adding one or more amino acids; 3) a fusion protein having the same function as the protein as shown in SEQ ID NO.1 and connected to a tag at the N-terminus or C-terminus; and also discloses the use of the AarTPS89 gene in catalyzing the precursor substance geranyl pyrophosphate GPP to generate borneol diphosphate, and further generating borneol through the catalytic action of phosphatase; wherein the AarTPS89 gene sequence is as shown in SEQ ID NO.2.

The invention belongs to but is not limited to the technical field of plant genetic engineering, and discloses a beta-Copaene / Geraniol synthetase gene MnTPS11 of Machilus machilus and an application of the beta-Copaene / Geraniol synthetase gene MnTPS11. The beta-Copaene / Geraniol synthase gene MnTPS11 of Machilus machilus provided by the invention is a key gene for synthesis of beta-Copaene / Geraniol in plant genetic engineering, synthesis of beta-Copaene / Geraniol can be effectively regulated and controlled, expression protein of the beta-Copaene / Geraniol synthase gene MnTPS11 is applied to synthesis and preparation of beta-Copaene / Geraniol, the yield is high, and the beta-Copaene / Geraniol synthase gene MnTPS11 has important application value. Substrates of farnesyl pyrophosphoric acid (FPP) and geranyl pyrophosphoric acid (GPP) are added to the expression protein of the induced Machilus machilus beta-Copaene / Geraniol synthetase gene MnTPS11, the catalytic activity of the recombinant protein is detected, and a GC-MS (Gas Chromatography-Mass Spectrometer) detection component shows that the result shows that the MnTPS11 is a bifunctional enzyme for synthesizing beta-Copaene / Geraniol.

The present application relates to the field of synthetic biology technology, and specifically discloses a cannabinol-producing microbial cell, its construction method, and application. A cannabinol-producing microbial cell includes a host cell, and the host cell includes a synthetic pathway for geranyl pyrophosphate, a synthetic pathway for olivate, and an isopentenyl transferase mutant. The construction method is as follows: based on the geranyl pyrophosphate-producing host cell yCAN10, the olivate synthesis pathway is constructed to form yCG02; a copy gene of the olivate synthase OLS is integrated into yCG02 to obtain yCG03; an acetyl-CoA carboxylase mutant ACC1 (S659A, S1157A) is integrated into yCG03 to obtain yCG04; and an isopentenyl transferase mutant is integrated into yCG04 to obtain yCG05, i.e., a cannabinol-producing microbial cell. The application is to produce cannabinol from a carbon source, which alleviates the problem of a large gap in cannabinol resources in the current market.

The application relates to an application of a geranylgeranyl pyrophosphate reductase in preparation of tocopherol, wherein a nucleotide sequence coding the geranylgeranyl pyrophosphate reductase is shown as SEQ ID No. 11, and the application comprises the following steps: under the assistance of coenzyme iron-oxidation-reduction protein and iron-oxygen-reduction protein-NADP+ reductase, the geranylgeranyl pyrophosphate reductase catalyzes the reduction of delta-tocotrienol to generate delta-tocopherol. The geranylgeranyl pyrophosphate reductase has the function of reducing the unsaturated isoprene side chain of tocotrienol, can catalyze the reduction of tocotrienol to generate tocopherol, does not need to depend on the geranyl pyrophosphate precursor in the existing tocopherol biosynthesis pathway, significantly improves the tocopherol synthesis efficiency, and provides a new strategy for the biosynthesis of tocopherol and other tocopherols.

Provided is a nucleic acid comprising a recombinant bacterial or archaeal geranyl pyrophosphate synthase (GPPS) gene, codon optimized for production in yeast. Also provided is a yeast cell comprising an expression cassette comprising the above nucleic acid. Additionally provided is a method of producing a terpene or cannabinoid in a yeast, the method comprising incubating the above yeast cell in a manner sufficient to produce the terpene or cannabinoid.

The invention discloses an application of geranyl-geranyl pyrophosphate synthetase in simple renal cyst, provides a development relationship between the geranyl-geranyl pyrophosphate synthetase and the simple renal cyst, and provides a new direction for targeted therapy of SRC by inducing formation of the renal cyst due to deletion of the geranyl-geranyl pyrophosphate synthetase. The invention further discloses that the variation frequency of the rs3806394 located in the GGPS1 promoter is increased in SRC patients, and the research on the pathological mechanism of SRC occurrence is facilitated.

The invention discloses a tobacco terpenoid synthase gene NtTPS2184 and application thereof, and belongs to the technical field of plant molecular biology, the nucleotide sequence of the tobacco terpenoid synthase gene NtTPS2184 is shown as SEQ ID NO.1, the invention provides a recombinant expression vector, the recombinant expression vector is obtained by positively inserting the tobacco terpenoid synthase gene NtTPS2184 between BamHI and SacI sites of pGEX4.0 plasmid, and the recombinant expression vector is used for expressing the tobacco terpenoid synthase gene NtTPS2184. The invention also provides an application of the tobacco terpene synthase gene NtTPS2184 or the recombinant expression vector in the synthesis of linalool and trans-nerolidone by a catalytic substrate, and the terpene synthase gene NtTPS2184 provided by the invention can be massively expressed by using escherichia coli genetically engineered bacteria, so that the yield of the tobacco terpene synthase gene NtTPS2184 is increased, and the yield of the tobacco terpene synthase gene NtTPS2184 is increased. In the presence of a substrate geranyl pyrophosphate, the monoterpene synthase exerts the activity of the monoterpene synthase to catalytically synthesize linalool; and in the presence of a substrate farnesyl pyrophosphate, the activity of the sesquiterpene synthase is exerted, and the trans-nerolidinol is catalytically synthesized.

The present invention relates to the field of bioengineering technology, and particularly relates to an engineered Saccharomyces cerevisiae strain, a method for constructing the same, and applications thereof. The Saccharomyces cerevisiae strain with the preservation number of CGMCC No. 26221 provided by the present invention is obtained by screening on the basis of the chassis strain Saccharomyces cerevisiae SyBE_Sc14D18 after site-directed mutagenesis, atmospheric and room temperature plasma mutagenesis, and adaptive laboratory evolution induced by hydrogen peroxide. On the basis of this strain, the expression of geranyl pyrophosphate synthase TmCrtE is increased and the expression of the Ura3 gene is restored to obtain an engineered Saccharomyces cerevisiae strain with high lycopene yield.

Disclosed are techniques for monoterpenes biosynthesis in engineered microorganisms having a mevalonate pathway, where the microorganisms express an engineered enzyme. The engineered enzyme may include one or more peptides that are selected to alter the active site pocket volume of the engineered enzyme as compared to a wild-type farnesyl pyrophosphate synthase. The engineered enzyme may include a first peptide (such as a linker peptide) fused to a C-terminus of a farnesyl pyrophosphate synthase. Optionally, a second peptide (such as a Myc-tag or a HA-tag) be fused to the first peptide. The method may include exposing the engineered microorganism to a carbon source (such a glucose). The method may include allowing the engineered microorganism to generate (i) a first product from the mevalonate pathway and (ii) the engineered enzyme. The method may include forming a monoterpene by allowing the first product and the engineered enzyme to interact.

The invention provides a method for producing an isoprenoid or a precursor thereof by microbial fermentation. Typically, the method involves culturing a recombinant bacterium in the presence of a gaseous substrate whereby the bacterium produces an isoprenoid or a precursor thereof, such as mevalonic acid, isopentenyl pyrophosphate, dimethylallyl pyrophosphate, isoprene, geranyl pyrophosphate, farnesyl pyrophosphate, and / or pinene. The bacterium may comprise one or more exogenous enzymes, such as enzymes in mevalonate, DXS, isoprenoid alcohol, or terpene biosynthesis pathways.

The invention belongs to the technical field of plant genetic engineering, and discloses a machilus nanmu TPS synthetase gene MnTPS7 and an application of the machilus nanmu TPS synthetase gene MnTPS7. The Machilus davidii Sesquiabinene / Linalool synthetase gene MnTPS7 provided by the invention is a key gene for synthesis of Sesquiabinene / Linalool in plant genetic engineering, synthesis of Sesquiabinene / Linalool can be effectively regulated and controlled, expression protein of the Machilus davidii Sesquiabinene / Linalool synthetase gene MnTPS7 is applied to synthesis and preparation of Sesquiabinene / Linalool, the yield is high, and the Machilus davidii Sesquiabinene / Linalool synthetase gene MnTPS7 has important application value. According to the present invention, substrates such as farnesyl pyrophosphate (FPP) and geranyl pyrophosphate (GPP) are added to the expression protein of the induced machilus machilus Sesquiabinene / Linalool synthase gene MnTPS7, the catalytic activity detection of the recombinant protein is performed, the gas chromatography-mass spectrometer (GC-MS) detection component shows that the main catalytic product of the FPP is Sesquiabinene, the main catalytic product of the GPP is Linalool, and the MnTPS7 is the bifunctional enzyme for synthesizing the Sesquiabinene / Linalool;

Provided herein are compositions and methods for producing myrcene by culturing genetically modified microbial host cells that express a myrcene synthase and optionally a geranyl pyroplosphate synthase. Also provided herein are isolated nucleic acid molecules that encode myrcene synthase variants derived from the Ocimum species myrcene synthase, which comprise one or more amino acid substitutions that improve in vivo performance of myrcene synthase in genetically modified microbial host cells. Also provided herein are isolated myrcene synthase variants that exhibit an improved activity for converting geranyl diphosphate into myrcene.

The invention discloses a saccharomyces cerevisiae engineering bacterium for efficiently synthesizing mogroside V based on cytoplasm and peroxisome double-cell-region indoor and a construction method of the saccharomyces cerevisiae engineering bacterium. Wherein the construction method comprises the following steps: integrating acetoacetyl coenzyme A thiolase ERG10, 3-hydroxy-3-methylglutaryl coenzyme A synthetase ERG13, truncated 3-hydroxy-3-methylglutaryl coenzyme A reductase tHMG1, phosphomevalonate kinase ERG8, mevalonate pyrophosphate decarboxylase ERG19, pentenyl pyrophosphate isomerase IDI1 and geranyl pyrophosphate synthase ERG20 at a site of a saccharomyces cerevisiae genome GAL80, and carrying out enzymolysis, so as to obtain a saccharomyces cerevisiae strain; a squalene synthase ERG9, an epoxy squalene cyclization enzyme ERG1 and the like; aBC efflux protein PDR11, isopentene phosphokinase AtIPK, geranyl pyrophosphate synthase ERG20, endogenous squalene synthetase ERG9, epoxy squalene cyclization enzyme ERG1 and the like are integrated at a site Exg1 of a saccharomyces cerevisiae genome. The yield of MG-V synthesized by shake flask fermentation of the saccharomyces cerevisiae engineering bacteria constructed by the invention reaches 117.6 mg / L, and the saccharomyces cerevisiae engineering bacteria have the advantages of high yield, strong metabolic flux directionality, wide industrial application prospect and the like.

The invention provides a method for producing a terpene or a precursor thereof by microbial fermentation. Typically, the method involves culturing a recombinant bacterium in the presence of a gaseous substrate whereby the bacterium produces a terpene or a precursor thereof, such as mevalonic acid, isopentenyl pyrophosphate, dimethylallyl pyrophosphate, isoprene, geranyl pyrophosphate, farnesyl pyrophosphate, and / or farnesene. The bacterium may comprise one or more exogenous enzymes, such as enzymes in mevalonate, DXS, or terpene biosynthesis pathways.

Disclosed herein are isoprenyltransferases capable of producing bakuchiol or an analog thereof from geranyl pyrophosphate (GPP) and cinnamic acid, cumaric acid, caffeic acid and / or ferulic acid. Also disclosed herein are engineered cells expressing an isoprenyltransferase capable of producing bakuchiol or an analog thereof, and methods of producing bakuchiol or an analog thereof using the isoprenyltransferase and the cells.

A prenyltransferase capable of producing bakuchiol or its analogues from geranyl pyrophosphate (GPP) and cinnamic acid, coumaric acid, caffeic acid and / or ferulic acid is disclosed herein. Manipulated cells expressing a prenyltransferase capable of producing bakuchiol or its analogues, as well as methods of using the prenyltransferase and cells to produce bakuchiol or its analogues, are also disclosed herein.

The invention discloses recombinant escherichia coli for producing (-)-isopulegol as well as a construction method and application thereof. The construction method comprises the following steps: firstly, constructing a plasmid 1 and a plasmid 2; the method comprises the following steps: constructing a plasmid 1 and a plasmid 2, constructing a plasmid 3 on the basis of the plasmid 1, constructing a plasmid 4 on the basis of the plasmid 2, and knocking out geranyl pyrophosphate synthase gene AgGPPS and limonene synthase gene McLMS in the plasmid pJBEI-6409 to obtain a plasmid 5; a plasmid 3, a plasmid 4 and a plasmid 5 are introduced into escherichia coli DH5alpha to obtain recombinant escherichia coli 7 for producing the (-)-isopulegol, and experiments prove that the (-)-isopulegol is obtained by mixed fermentation of the recombinant escherichia coli 6 and 7, and the shake flask yield of the (-)-isopulegol reaches 12.4 mg / L.

This disclosure generally relates to the use of enzyme combinations or recombinant microbes comprising same to make isoprenoid precursors, isoprenoids and derivatives thereof including prenylated aromatic compounds. Novel metabolic pathways exploiting Claisen, aldol, and acyloin condensations are used instead of the natural mevalonate (MVA) pathway or 1-deoxy-d-xylulose 5-phosphate (DXP) pathways for generating isoprenoid precursors such as isopentenyl pyrophosphate (IPP), dimethylallyl pyrophosphate (DMAPP), and geranyl pyrophosphate (GPP). These pathways have the potential for better carbon and or energy efficiency than native pathways. Both decarboxylative and non-carboxylative condensations are utilized, enabling product synthesis from a number of different starting compounds. These condensation reactions serve as a platform for the synthesis of isoprenoid precursors when utilized in combination with a variety of metabolic pathways and enzymes for carbon rearrangement and the addition / removal of functional groups. Isoprenoid alcohols are key intermediary products for the production of isoprenoid precursors in these novel synthetic metabolic pathways. These precursors can be modified to various isoprenoid products through prenyl transferase, terpene synthase, or terpene cyclases. The production of prenylated aromatic compounds is achieved through prenyl transfer of the hydrocarbon units of isoprenoid precursors to polyketides.

The invention discloses thiogeranyl geranyl pyrophosphate GGSPP and application thereof in preparation of liver cancer drugs, the structural formula of the GGSPP is as shown in formula (I): # imgabs0 # GGSPP specifically binds and activates human fructose-1, 6-diphosphatase 1FBP1 protein, up-regulates the enzymatic activity of FBP1, and reverses the Warburg effect of tumor cells, thereby inhibiting proliferation, migration and development of hepatocellular carcinoma. The invention not only clarifies the key effect of the GGSP-FBP1 axis in the occurrence and development of liver cancer, but also provides an efficient, specific and mechanism-clear anti-liver cancer candidate drug, opens up a new direction for the treatment of metabolism-related tumors, and has important scientific significance and clinical application prospects.

The invention provides a method for producing an isoprenoid or a precursor thereof by microbial fermentation. Typically, the method involves culturing a recombinant bacterium in the presence of a gaseous substrate whereby the bacterium produces an isoprenoid or a precursor thereof, such as mevalonic acid, isopentenyl pyrophosphate, dimethylallyl pyrophosphate, isoprene, geranyl pyrophosphate, farnesyl pyrophosphate, and / or pinene. The bacterium may comprise one or more exogenous enzymes, such as enzymes in mevalonate, DXS, isoprenoid alcohol, or terpene biosynthesis pathways.

The invention relates to the technical field of metabolic engineering, in particular to a recombinant saccharomyces cerevisiae strain for producing cannabinoid phenolic acid at high yield by using sugar as well as a construction method and application of the recombinant saccharomyces cerevisiae strain. According to the invention, firstly, reverse beta-oxidation pathway genes ReBktB, CnPaaH1, CaCrt and TdTer are introduced, so that the problem that cannabinoid phenolic acid biosynthesis depends on a hexanoic acid substrate is solved; and then the gene GAL80 of the GAL4 inhibitory protein is knocked out, and the GAL4 is over-expressed, so that the overall expression level of the pathway is up-regulated. Then, a mercaptase gene Ckth1A and a hydroxyacyl coenzyme A dehydrogenase gene Ckhbd1 are introduced, so that the supply of the hexanoyl coenzyme A is improved. Furthermore, the endogenous farnesyl pyrophosphate synthetase gene ERG20 is down-regulated, and the consumption of geranyl pyrophosphate by a branch path is reduced. Finally, the expression level of part of genes in saccharomyces cerevisiae cells is regulated and controlled through gene editing, and the purposes of synthesizing cannabinoid phenolic acid from the beginning by using saccharomyces cerevisiae and taking sugar as a substrate and improving the yield of cannabinoid phenolic acid are achieved.

The invention provides an Escherichia coli engineering bacterium for improving the yield of astaxanthin by using yhaK as well as a construction method and application of the Escherichia coli engineering bacterium. The engineering bacterium comprises a recombinant plasmid containing a yhaK gene, a crtS gene for coding astaxanthin synthetase, a crtR gene for coding cytochrome reductase, a crtYB gene for coding lycopene cyclase and a gene segment for coding farnesyl / geranyl pyrophosphate synthase IspA, the escherichia coli engineering strain ELZSS02 E.coli Bl21 (DE3): pTrc low-MvaE-MvaS / pET-RPlyc-crtS / pACYC-crtR-crtYB-IspA-yhaK is finally obtained, compared with wild escherichia coli, the expression quantity of MEP pathway genes in the engineering strain is remarkably increased, the yield of astaxanthin is also remarkably increased, and the escherichia coli engineering strain has good application value.

Provided are various enzymes for cannabinoid synthesis and methods for their preparation and use. Also provided are recombinant polypeptides having prenyltransferase activity and / or geranyl diphosphate:olivetolate geranyltransferase (GOT) activity, and engineered recombinant microorganisms for the production of CBGA, and methods for producing cannabigerolic acid (CBGA) or an analogue thereof, the methods comprising the step of reacting a heterologously expressed prenyltransferase with geranyl diphosphate (GPP) and an acid such as olivetolic acid (OA).

The invention provides a yarrowia lipolytica engineering strain with high yield of beta-carotene and construction of the yarrowia lipolytica engineering strain, and belongs to the technical field of synthetic biology and metabolic engineering. According to the invention, a multi-copy geranyl-geranyl pyrophosphate synthase gene, a compartmentalized localization expression beta-carotene synthetic pathway related enzyme gene, a diacylglycerol acyltransferase gene, a cell cycle regulation transcription factor, an oxidative stress regulation related gene, and a cell wall and membrane lipid homeostasis regulation related gene are integrated; and regulating foreign protein folding and expression related genes to obtain the yarrowia lipolytica engineering strain for efficiently biosynthesizing beta-carotene by taking glucose as a substrate. The yield of beta-carotene of the constructed yarrowia lipolytica engineering strain is high, the yield at the shake flask level can reach 3253.2 mg / L, and the yield can be increased to 26.5 g / L when fed-batch fermentation is carried out on a 5L scale. The method has great practical significance and economic value for industrial and commercial application of high-value terpenoid products.