23 results about "Phosphoketolase" patented technology

The invention relates to a recombinant pichia pastoris strain capable of producing astaxanthin at high yield as well as a construction method and application of the recombinant pichia pastoris strain. The recombinant pichia pastoris strain is obtained by expressing ATP (adenosine triphosphate) citrate lyase (ACL) and acetyl-CoA synthase (ACS) or phosphoketolase (PK) and phosphotransacetylase (PTA), choline kinase (CK), inositol polyphosphate kinase (IPK) and vitreoscilla hemoglobin (VHB) in host bacteria. Wherein the host bacterium is a pichia pastoris gene modified strain PP-LC2. The astaxanthin production performance of the recombinant strain is verified on the basis of comparison of precursor supply pathways, IUP pathway construction and improvement of the oxygen supply capacity of the engineering strain, and the astaxanthin production capacity of the pichia pastoris is further improved. The construction method of the recombinant pichia pastoris is simple, the synthesis of astaxanthin can be better promoted, the engineering strain can efficiently synthesize the astaxanthin by utilizing methanol through amplification fermentation of a 5L fermentation tank, and industrial production is facilitated.

The invention belongs to the technical field of biosynthesis, and particularly relates to a strain for producing sclareol as well as a construction method and application of the strain. In order to solve the problem of microbial synthesis of sclareol, phosphoketolase in an NOG pathway is mutated and optimized, phosphoketolase NaXpk from nanoarchaea archaeon is subjected to site-directed mutagenesis, a sclareol synthesis pathway is constructed in an escherichia coli host, the influence of NaXpk before and after mutation on the yield of sclareol is compared, and the yield of sclareol is improved. The S472F mutant and the S472Y mutant are determined, the S472F mutant and the S472Y mutant are applied to a sclareol production path, the sclareol yield of the finally obtained production strains can reach 623.3 mg / L and 655.2 mg / L, and the effect is remarkable.

Modified bacteria that produce L-glutamic acid are provided wherein a heterologous polynucleotide of phosphoketolase (XFP) is contained in its genome as compared to non-modified bacteria. The phosphoketolase is derived from bifidobacterium. In addition, methods of increasing the yield of L-glutamic acid using the modified bacteria are also provided.

The present disclosure describes a recombinant yeast host cell having a first engineered metabolic pathway to convert fructose-6-phosphate (F-6-P) into acetyl-coenzyme A. The first engineered metabolic pathway comprises: a genetic modification for expressing a heterologous phosphoketolase capable of converting fructose-6-phosphate into erythrose-4-phosphate, and at least one genetic modification for decreasing native 6- phosphofructo-2-kinase activity classified under Enzyme Commission No. 2.7.1.105. Optionally, the recombinant yeast host cell comprises a second engineered metabolic pathway for convert acetyl-coA into a fermentation product. The present disclosure further describes a process for making a fermentation product. The process comprises contacting the recombinant yeast host cell described herein with a carbohydrate source under a condition allowing the conversion of at least a part of the carbohydrate into a fermentation product.

The invention discloses a genetically engineered bacterium for synthesizing N-acetyl blue as well as a construction method and application of the genetically engineered bacterium. The genetically engineered bacterium is prepared by knocking out a lactic dehydrogenase gene ldh and inserting a phosphoketolase gene xfpK on a knockout site, knocking out a pyruvate oxidase gene poxB and inserting a phosphate transacetylase gene pta on a knockout site of a microorganism for producing N-acetyl blue, and knocking out an aconitase inhibition factor gene acnR, inserting a glutamate dehydrogenase gene gdhA on a knockout site, knocking out a glutamine synthetase adenylation transferase gene glnE, and inserting an alpha-ketoglutarate dehydrogenase inhibition factor gene odhI on a knockout site to obtain the strain. According to the method, the supply of the precursor acetyl coenzyme A is enhanced by introducing a non-oxidative glycolysis pathway, the pH dynamic regulation system balances the metabolic flux, and the double breakthrough of the yield and the carbon recovery rate of the N-acetyl blue is realized by combining a two-stage fermentation process.

The invention discloses construction and application of a recombinant escherichia coli engineering strain with high yield of beta-arbutin, and belongs to the technical field of gene engineering and biological engineering. According to the invention, a key gene for synthesizing beta-arbutin is regulated and expressed by adopting a temperature-sensitive promoter; an enzyme mutant aroGS180F capable of resisting feedback inhibition is introduced; an exogenous phosphoketolase pathway is introduced to enhance E4P supply; an acetic acid branch pathway gene is knocked out, so that the formation of a by-product acetic acid is reduced, and the yield of beta-arbutin of the constructed engineering strain can reach 6.83 g / L. The high-yield beta-arbutin escherichia coli recombinant strain constructed by the invention lays a foundation for the subsequent biosynthesis of beta-arbutin and derivatives, and has potential value and significance for the development of synthetic biology.

The present invention belongs to the fields of genetic engineering and enzyme engineering, and discloses a phosphoketolase mutant derived from Bifidobacterium adolescentis and its application in the production of metabolites. The present invention uses a site-directed mutagenesis method to mutate the 480th histidine of the phosphoketolase PKT derived from Bifidobacterium adolescentis to lysine, significantly improving the enzyme activity of the phosphoketolase. This effectively solves the problem of the current low catalytic performance of phosphoketolase on fructose-6-phosphate. The enzyme has been successfully applied to the fermentation production of bacitracin, ectoine, and γ-aminobutyric acid, providing a new approach to improving the production level of metabolites.

Provided is a modified bacterium for producing L-glutamic acid, which contains a heterologous polynucleotide encoding a phosphoketolases (XFP) in the genome thereof compared to a non-modified bacterium. The phosphoketolases is derived from Bifidobacterium. In addition, further provided is a method for increasing the yield of the L-glutamic acid by means of using the modified bacterium.

The invention discloses a genetically engineered bacterium for producing 1, 3-butanediol and application of the genetically engineered bacterium, and belongs to the technical field of genetic engineering. The invention discloses a genetically engineered bacterium for producing 1, 3-butanediol, which is characterized in that JM109 (DE3) is used as a starting strain, and a citrate synthase gene gltA is weakened; the method comprises the following steps: knocking out a D-lactic dehydrogenase gene ldhA and a pyruvate dehydrogenase-ubiquinone gene poxB; according to the invention, a phosphoketolase gene xfp and phosphotransacetylase genes pta, PhaA, PhaB, B1d *, yqhd, fdh1, nadk, pntA and pntB are overexpressed. When the genetically engineered bacterium constructed by the invention is used for producing the 1, 3-butanediol, the conversion rate is high, the economical efficiency is good, the sustainability is realized, and the industrial production is easy.

The invention relates to a recombinant pichia pastoris strain for producing coupled xylitol by NADPH (nicotinamide adenine dinucleotide phosphate) dependent growth and a construction method of the recombinant pichia pastoris strain, and belongs to the technical field of microbial fermentation. The recombinant pichia pastoris strain provided by the invention contains an NADPH dependent xylitol dehydrogenase gene, a sugar phosphatase gene, a D-arabinol-4-dehydrogenase gene, a fructose-1, 3, 4-triazole-1, 3, 4-triazole-1, 3-triazole-1, 3-triazole-1, 3-triazole-1, 3- The gene sequence is as follows: a 1, 6-diphosphatase gene, a phosphoketolase gene, a phosphate transacetylase gene, a glucose-6-phosphate dehydrogenase gene, a 6-phosphogluconolactonase gene, a 6-phosphogluconate dehydrogenase gene and a ribulose-5-phosphate epimerase gene; gene for expressing xylulokinase is knocked out or down-regulated: NADH dependent type xylitol dehydrogenase gene, ribose-5-phosphate isomerase gene, glutamate dehydrogenase gene, phosphofructokinase gene and phosphoglucose isomerase gene. The yield of xylitol produced by fermentation of the strain is far higher than that in the prior art.

This invention belongs to the field of biosynthesis technology, specifically relating to a strain for producing perillaldehyde, its construction method, and its application. Addressing the microbial synthesis problem of perillaldehyde, this invention mutates and optimizes the phosphotransketase in the NOG pathway, using a strain derived from nanoarchaemonas (…). Nanoarchaeota archaeon Site-directed mutagenesis was performed on the phosphoketolase NaXpk, and a perillaldehyde synthesis pathway was constructed in E. coli. The effects of NaXpk mutation on perillaldehyde yield were compared to identify the S472F and S472Y mutants. These mutants were then applied to the perillaldehyde production pathway, and the resulting production strains achieved perillaldehyde yields of 623.3 mg / L and 655.2 mg / L, respectively, demonstrating significant effectiveness.

The present invention discloses a genetically engineered bacterium producing 1,3-butanediol and its application, belonging to the field of genetic engineering technology. A genetically engineered bacterium producing 1,3-butanediol disclosed in the present invention, with JM109 (DE3) as the starting strain, weakens the citrate synthase gene gltA; knocks out the D-lactate dehydrogenase gene ldhA and the pyruvate dehydrogenase ubiquinone gene poxB; overexpresses the phosphoketolase gene xfp, the phosphotransacetylase gene pta, PhaA, PhaB, Bld*, yqhd, fdh1, nadk, pntA, pntB gene. The genetically engineered bacterium constructed using the present invention produces 1,3-butanediol with high conversion rate, good economy, sustainability and ease of industrial production.

The invention discloses a saccharomyces cerevisiae genetically engineered bacterium for efficiently synthesizing mogroside V through multi-dimensional cytoplasm adaptability and a construction method of the saccharomyces cerevisiae genetically engineered bacterium. The construction method of the saccharomyces cerevisiae genetically engineered bacterium comprises the following steps: integrating truncated 3-hydroxy-3-methylglutaryl coenzyme A reductase tHMG1, pentenyl pyrophosphate isomerase IDI1, squalene synthetase ERG9, epoxy squalene cyclization enzyme ERG1, choline kinase ScCK, isopentenyl phosphokinase AtIPK, phosphoketolase BbPK, phosphotransacetylase PTA and dihydroxyacetone hydrolase EcHAD at a site of a saccharomyces cerevisiae genome GAL80, and carrying out enzymolysis, so as to obtain the saccharomyces cerevisiae genetically engineered bacterium. The invention relates to a preparation method of a cytochrome P450 enzyme, which comprises the following steps of: preparing a cycloepoxide hydrolase SgEPH3, a cytochrome P450 enzyme CYP87D18 and a cytochrome P450 enzyme reductase AtCPR1; an ABC efflux protein PDR11, a glycosyl transferase UGTMG1, a sucrose synthase Susy and a glycosyl transferase SgUGT94-289-3 are integrated at a site Exg1 of a saccharomyces cerevisiae genome. The saccharomyces cerevisiae genetically engineered bacterium constructed by the invention realizes a higher level of mogroside V yield synthesized from the beginning in microorganisms, and has important industrial application potential.

The invention discloses a method for increasing glutamic acid yield. Modified bacteria that produce L-glutamic acid are provided wherein a heterologous polynucleotide of phosphoketolase (XFP), and one or more of a modification of increased activity of the gluAA1T, icd gene, aceE gene and / or pyc gene, are included in the genome as compared to non-modified bacteria. In addition, methods of increasing the yield of L-glutamic acid using the modified bacteria are also provided.

The invention discloses a genetically engineered bacterium for producing O-acetyl-L-homoserine as well as a construction method and application of the genetically engineered bacterium, the construction method is characterized in that a strain OAH1 is taken as a chassis bacterium, and a genome of the strain OAH1 is subjected to at least one modification from (1) to (4) through the following steps: (1) enhancing and regulating the expression of a gene of an acetyl coenzyme A module; (2) overexpressing a heterologous phosphoketolase coding gene fxpK and a pantothenate kinase coding gene coA; (3) strengthening an OAH transfer module; and (4) carrying out overexpression on the heterologous homoserine acetyltransferase coding genes cmetX and cgmetX, so as to obtain the homoserine acetyltransferase gene. According to the engineering strain, effective accumulation of OAH is achieved, the shake flask yield reaches 17.8 g / L, the fed-batch fermentation yield of a 5L fermentation tank reaches 105.3 g / L, the sugar-acid conversion rate reaches 55%, and a foundation is laid for subsequent construction of high-yield OAH engineering bacteria.

The invention relates to phosphoketolase MaPKT and SzPKT with a function of catalyzing formaldehyde to synthesize acetyl phosphate and application of phosphoketolase MaPKT and SzPKT in CO2 biosynthesis of ethanol, and belongs to the technical field of enzyme mining and biological catalysis application. A phosphoketolase specific protein sequence database and an oligopeptide module sequence library are established through an oligopeptide module recognition (PPR) technology, two phosphoketolase PKTs capable of catalyzing formaldehyde to synthesize acetyl phosphate are efficiently and accurately excavated, further research finds that the formaldehyde tolerance of MaPKT is improved, and the catalytic efficiency of MaPKT on formaldehyde is 8 times that of glycolaldehyde. Based on this, the invention also constructs an efficient condensation approach for synthesizing ethanol by artificially biologically converting CO2, and has potential application prospects.

The present application provides phosphoketolase mutants, related products thereof and applications thereof in production of acetyl phosphate, acetyl-CoA and / or 3-hydroxypropionic acid. The phosphoketolase mutants provided by the present application have high catalytic capacity for both six-carbon ketose and five-carbon ketose, and can catalyze the conversion of six-carbon ketose and five-carbon ketose into acetyl phosphate with high efficiency, and then efficiently generate subsequent products acetyl-CoA and 3-hydroxypropionic acid, thereby having great application potential in production of acetyl-CoA and products (such as 3-hydroxypropionic acid) taking acetyl-CoA as a precursor.

Disclosed in the present invention are a recombinant yeast strain for synthesizing an aromatic compound, and a construction method for and use of the recombinant yeast strain. According to the recombinant yeast strain for synthesizing the aromatic compound, Saccharomyces cerevisiae QL35 is used as a chassis, heterologously introduced phosphoketolase replaces endogenous non-oxidized phosphopentose pathway of Saccharomyces cerevisiae cells, a necessary substrate, namely, erythrose 4-phosphate, of the aromatic compound is synthesized, and acetyl phosphate is generated at the same time. Pyruvate kinase is further knocked out, the decomposition of phosphoenolpyruvic acid into pyruvic acid is blocked, and at the moment, the recombinant strain can only be converted into acetyl coenzyme A by means of acetyl phosphate to drive the growth of cells. Finally, a metabolic pathway is transformed into coupling product production and cell growth, and a recombinant yeast strain with high aromatic compound yield is obtained by means of adaptive evolution. The strain reduces the production cost by using glucose and an inorganic salt culture medium, and the growth-coupled recombinant yeast strain has the advantage of maintaining stable genetic character in fermentation process.

The invention discloses a method for reducing CO2 emission of escherichia coli, enhancing CO2 immobilization and increasing the yield of L-threonine, and belongs to the technical field of biology. According to the method, CO2 emission of escherichia coli is reduced by introducing a non-oxidative glycolysis pathway, and the yield of L-threonine and the sugar-acid conversion rate are improved by enhancing CO2 fixation efficiency; specifically, a phosphoketolase mutant derived from pseudomonas aeruginosa, an acetyl coenzyme A synthetase mutant derived from enterobacter faecalis and a carbonic anhydrase mutant derived from thiobacillus thermophilus are introduced, expression of pfkA is weakened, more carbon sources are intercepted to a non-oxidative glycolysis pathway, and ackA genes are knocked out, so that production of acetic acid is reduced. Finally, the obtained engineering strain THRHS-6 is subjected to fed-batch fermentation in a 5L fermentation tank to accumulate 165.3 g / L of L-threonine, and the sugar-acid conversion rate is 0.63 g / g.

The present invention provides non-natural microorganisms containing enzymatic pathway and / or metabolic modifications for increasing carbon flux by acetyl-CoA or by oxaloacetic acid and acetyl-CoA. Embodiments of the invention include a microorganism having a pathway for obtaining acetyl-CoA and oxaloacetic acid (PK pathway) comprising a phosphoketolase. The microorganism also has (i) a genetic modification that enhances the sugar uptake activity of a non-phosphotransferase (non-PTS) system, and / or (ii) a genetic modification in the electron transport chain (ETC) of the microorganism that increases the efficiency of ATP production that increases the availability of reducing equivalents, or both. The microorganism may optionally include (iii) a genetic modification that maintains, attenuates, or eliminates the sugar uptake activity of a phosphotransferase system (PTS). The increased carbon flux by acetyl-CoA and oxaloacetic acid may be used for the production of biologically derived compounds, and the microorganisms may further include pathways capable of producing biologically derived compounds.

Described herein are engineered cells including ones having synthetic methylotrophy which include an NADH-dependent enzyme capable of converting G3P to 3PG (e.g., B. methanolicus gapN) and / or fructose-1,6-bisphosphatase, along with hexulose-6-phosphate synthase, 6-phospho-3-hexuloisomerase, a phosphoketolase, or a combination thereof. Engineered cells of the disclosure beneficially maintain adequate pool sizes of phosphorylated C3 and / or C4 compounds, and / or provide increased levels of NADPH. As such, the modifications allow for the generation of C6 compounds from C1 (e.g., a methanol feedstod) and C5 compounds, the regeneration of C5 compounds from C6 compounds by carbon rearrangement, and an improved balance between regeneration of C5 compounds and lower glycolysis. In turn, this allows the engineered microorganism to generate sufficient quantities of metabolic precursors (e.g., acetyl-CoA) which can be used in a bioproduct pathway, and the engineered cells can include further modifications to those pathway enzymes allowing for production of a desired bioproduct.

The present invention belongs to the field of biotechnology, and specifically relates to a recombinant Escherichia coli for producing mevalonic acid and its application. The present invention uses Escherichia coli MG1655 as the host cell, and transfers the HMG-CoA synthase MvaS gene and HMG-CoA reductase MvaE gene derived from Lactobacillus casei, the acetyl-CoA acetyltransferase atoB gene derived from Escherichia coli, the phosphoketolase fxpk gene derived from Bifidobacterium adolescentis, and the fructose-1,6-bisphosphatase fbp gene derived from Escherichia coli into the Escherichia coli host cell. The recombinant Escherichia coli constructed by the present invention can be used for the synthesis of mevalonic acid. Using glucose as the substrate, after fermentation in a 5 L fermenter at 37 °C for 48 hours, the yield of mevalonic acid can reach 87.86 g / L. It has the advantages of short fermentation time and high yield, meeting the requirements of industrial production.

The invention relates to a method for production of acetyl phosphate from renewable or next generation feedstocks, comprising enzymatically converting a non-phosphorylated carbohydrate into acetyl phosphate and a product, with a phosphoketolase. In embodiments, the phosphoketolase comprises at least one mutation in its active site preferably enabling an increased affinity and / or increased phosphoketolase activity for a non-phosphorylated carbohydrate, such as i) D-fructose, (ii) D-erythrulose or (iii) glycolaldehyde, compared to a wild- type phosphoketolase sequence. The invention further relates to a phosphoketolase enzyme comprising in its active site the mutation(s) H548N, N549D, H256Y, T2A:16T:H260Y:H548N, H548Y, H260Y:H548Y, H256Y:H260Y:H548Y, H142N:E153D and / or T2A:16T:H260Y:H548Y and its use for producing acetyl phosphate from a non-phosphorylated carbohydrate. The invention further relates to non-human organisms, preferably a microorganism, comprising a gene encoding and optionally expressing said phosphoketolase enzyme and compositions comprising the same.