157 results about "Whole cell" patented technology

The invention discloses a biosynthesis system for high-yield 1, 6-hexamethylenediamine and application of the biosynthesis system, and belongs to the technical field of biology. According to the method, key genes which are excavated and optimized in a 1, 6-hexamethylenediamine biosynthetic pathway are co-expressed in a single cell module or a multi-cell module, and different enzyme elements expressed by the cell module are subjected to enzyme cascade reaction, so that 1, 6-hexamethylenediamine can be efficiently synthesized by taking caprolactam or 6-aminocaproic acid as a substrate; furthermore, after the carboxylic acid reductase MaCAR is mutated, the catalytic activity of the carboxylic acid reductase MaCAR can be remarkably improved, and the yield of the 1, 6-hexamethylenediamine is further improved; in addition, the biosynthesis system disclosed by the invention directly takes the cell module as a whole-cell catalyst, and an enzyme purification step is avoided, so that the biosynthesis system has the advantage of low cost.

The invention discloses an arginine decarboxylase diaA enzyme mutant and application thereof in preparation of butanediamine, and belongs to the field of bioengineering. According to the invention, rational charge overturning transformation is simultaneously carried out on a pentamer meridian oligomeric interface and a dimer latitudinal oligomeric interface, so that stable assembly and efficient catalysis of the decamer under the condition of neutral to alkaline pH (7.0-9.0) are realized. Wherein positive charges are introduced into a meridian interface to weaken electrostatic repulsion, and negative charges are introduced into a latitudinal interface to enhance dimer compactness and substrate transfer efficiency. The specific enzyme activity of the representative double mutant AdiAD110K / H736E at pH 8.0 is about 35 times that of a wild type, and the representative double mutant AdiAD110K / H736E keeps a complete decamer state in a pH range of 7.0-9.0. The yield of butanediamine is up to 145.9 g / L under the whole-cell catalysis of the mutant.

Provided are S-adenosylmethionine synthetase mutants and the use thereof, belonging to the technical field of biological enzyme engineering. Site-directed combinatorial mutations are performed on positions 189, 200, 234, 266 and 371 of S-adenosylmethionine synthetase MAT to obtain a series of mutants with improved enzyme activity, wherein the specific enzyme activity of I189V / V266H and V266H / N371R is increased by more than 10 times relative to the parent. The yields of S-adenosylmethionine synthesized by whole cells of recombinant bacteria expressing mutants I189V / V266H / N371R, V266H / Q234N / N371R and I189V / V266H / Q234N / N371R all reach 1550 mg / L or more. The provided enzyme mutants have broad application prospects in the production of S-adenosylmethionine and the construction of genetically engineered microorganisms for S-adenosylmethionine.

The invention discloses a high-efficiency lysine decarboxylase mutant and an application of the high-efficiency lysine decarboxylase mutant in synthesis of pentamethylene diamine. A series of mutants including a single mutant (such as N218G, T222V, C244P and the like), a double mutant and a three mutant are obtained by carrying out multi-site mutation on wild type lysine decarboxylase (SEQ ID NO.2) from Klebsiella grimontii. The catalytic efficiency of the mutant is remarkably improved, and the activity of the mutant is improved by 5.0-10.2 times compared with that of a wild type enzyme. According to the method disclosed by the invention, the substrate conversion rate of 99% or above can be realized within 2 hours through an optimized process of constructing an expression vector and a genetically engineered bacterium, inducing expression of lysine decarboxylase and synthesizing 1, 5-pentanediamine by utilizing a whole-cell catalysis technology. Compared with a traditional chemical method and a natural enzyme method, the method disclosed by the invention has the advantages of simplified process, high catalytic efficiency, low production cost and the like, and is suitable for producing bio-based 1, 5-pentamethylene diamine and nylon 56 salt.

The invention provides an application of inositol monophosphate in an enzyme catalysis reaction system for preparing inositol. The invention also provides a preparation method of the inositol, the starch is used as a substrate, glucan phosphorylase, glucose phosphate mutase, inositol-3-phosphate synthase and inositol monophosphate are added as enzyme catalysts to catalyze the substrate to generate the inositol, the inositol monophosphate is derived from Archaeoglobus fulgidus, the amino acid sequence is shown as SEQ ID NO: 12, or the amino acid sequence is shown as SEQ ID NO: 1, or the amino acid sequence is shown as SEQ ID NO: 2, or the amino acid sequence is shown as SEQ ID NO: 3, or the amino acid sequence is shown as SEQ ID NO: 4, or the amino acid sequence is shown as SEQ ID NO: 5, or the amino acid sequence is shown as SEQ ID NO: 6. The inositol monophosphate is derived from Thermococcus kodakarensis, and the amino acid sequence of the inositol monophosphate is as shown in SEQ ID NO: 13; or the inositol monophosphate is derived from Pseudothermotoga lettinae, and the amino acid sequence of the inositol monophosphate is as shown in SEQ ID NO: 14. Inositol-3-phosphate synthetase and inositol monophosphate with good performance are screened, high-concentration starch is efficiently catalyzed to be converted into inositol by a one-pot method in a whole-cell feeding mode, and NAD < + > and other coenzymes do not need to be used. Under a 60L reaction system, 200g / L of corn starch is catalyzed, the unit yield of the prepared inositol reaches 161g / L, and the method has a good industrial application prospect.

The invention discloses a method for preparing butanediamine and spermidine from whole cells, and belongs to the field of bioengineering. According to a rational copolymerization modification strategy, a meridian interface and a latitudinal channel interface of arginine decarboxylase are subjected to rational modification respectively or simultaneously, and a series of mutants are obtained. The optimal mutant AdiA H729D / E467K / H736E, which is subjected to double-interface synergistic modification, disclosed by the invention, keeps high activity in the whole neutral-alkaline range of pH (Potential of Hydrogen) of 7.0 to 9.0. The optimal pH of the arginine decarboxylase mutant is increased, the enzyme activity stability in a neutral pH range is improved, the arginine decarboxylase mutant is more suitable for the condition requirements of industrial microbial fermentation, and a foundation is laid for efficient synthesis of butanediamine. The spermidine yield is up to 153.3 mg / L by using the mutant to catalyze the whole cell.

The invention provides a method, device and system for evaluating the effect of intervening small molecules by tumors and a storage medium, and relates to the technical field of bioinformatics, the method comprises the following steps: obtaining a first transcriptome representation of a tumor cell population before target small molecule intervention and a second transcriptome representation of a normal tissue cell population corresponding to the tumors; inputting the first transcriptome representation and the condition vector into a prediction model to obtain a third transcriptome representation; calculating a first distance and a second distance; an intervention effect is evaluated based on a comparison of the first distance and the second distance. According to the method, the influence of the drug on the transcriptome of the whole cell population is efficiently predicted, and the distance between the transcriptome and the health state is quantitatively compared, so that a set of brand new standard for improving the evaluation target from the traditional'killing cell 'to'reversing disease state' is established, and an objective decision basis is provided for developing more accurate drugs.

The present application relates to the technical field of fermentation engineering, and provides an engineered Escherichia coli for producing uroporphyrin as well as a construction method and application thereof.The present application uses Escherichia coli to express porphyrin synthase of thermophilic bacteria, performs whole-cell catalysis of 5-ALA to synthesize uroporphyrin under high-temperature conditions, and completely shields endogenous regulation; further, the Escherichia coli is wrapped by COF material to prevent the loss of enzymes in the cell recovery process; and a protein scaffold based on dockerin-cohesin is constructed to further improve the yield of uroporphyrin III.The present application creates a brand-new uroporphyrin high-temperature whole-cell catalysis process, realizes the efficient production of uroporphyrin, and provides a basis and idea for the industrial production of uroporphyrin.

The invention discloses a diketopiperazine heterodimer as well as a biosynthesis method and application thereof, and belongs to the technical field of bioengineering. The biosynthesis method of the diketopiperazine heterodimer comprises the following steps: S1, culturing a recombinant cell containing P450 dimerization enzyme or a mutant thereof, and performing induced expression to obtain a whole-cell culture; and S2, adding a cyclic dipeptide substrate containing tryptophan into the whole-cell culture, carrying out a catalytic reaction, and separating and purifying the obtained product to obtain the diketopiperazine heterodimer. According to the invention, the P450 dimerization enzyme or the mutant thereof is used for catalyzing the biosynthesis of the tryptophan-containing diketopiperazine into the diketopiperazine heterodimer with different regioselectivity / stereoselectivity, the biosynthesis method is green and environment-friendly, the efficiency is high, and the variety of the diketopiperazine heterodimer is greatly expanded.

Disclosed are systems and methods for producing renewable alcohols by fermentative processes using cell lysate or a combination of cell lysate and whole cells, as well as the cell lysate itself. The disclosure also provides processes and systems for converting renewable alcohols to transportation fuels, such as, gasoline, jet fuel, and diesel fuels.

This application discloses an embedded whole-cell biosensor, its fabrication method, and its application in machine learning-assisted mercury ion detection, belonging to the field of sensor materials and detection technology. This sensor encapsulates *E. coli* expressing green fluorescent protein (GFP) in a polyacrylamide / hyaluronic acid (PAAM / HA) hydrogel matrix, enabling highly selective and sensitive detection of mercury ions. It effectively eliminates interference from competing metal ions and can be completely regenerated through EDTA-2Na treatment, allowing for repeated use without performance loss. It maintains a stable fluorescence response for up to three weeks, possesses excellent biocompatibility and biodegradability, minimizing the risk of secondary contamination, and facilitates rapid on-site analysis.

The invention provides a beta-glucosidase mutant from Paenibacillus phyllosphaerae, and a method for efficiently preparing rare ginsenoside F2 by using an enzyme method and a whole-cell method, and particularly provides a beta-glucosidase mutant from Paenibacillus phyllosphaerae and a method for efficiently preparing rare ginsenoside F2 by using a beta-glucosidase mutant from Paenibacillus phyllosphaerae. The method comprises the following steps: engineering-modifying beta-glucosidase by utilizing computer-aided design, constructing a mutant library, carrying out primary screening and secondary screening to obtain two high-activity mutants Q7D and G189A, constructing a combined mutant Q7D / G189A, transforming the combined mutant Q7D / G189A into genetically engineered bacterium escherichia coli for expression, and producing F2 by utilizing a whole-cell method. Finally, the highest yield of 18.01 g / L is achieved after the reaction is carried out for 10 h in a 5 L fermentation tank and is 6.82 times that of a shake flask, and the purity of F2 reaches up to 82.20% after the reaction is carried out for 24 h finally. The invention not only provides an efficient solution for large-scale production of rare saponin F2, but also provides a transferable framework for designing and engineering diversified glycoside hydrolase in customized biological catalysis.

The invention discloses an amylosucrase mutant and application thereof in synthesis of alpha-arbutin, the mutant is obtained by mutation of an amino acid sequence of amylosucrase derived from coprophilobacterium mediterranei, and the amino acid sequence of the mutant is shown as SEQ ID NO.4. The invention further discloses a preparation method of the amylosucrase mutant. Compared with a wild strain, the catalytic activity, the temperature stability and the substrate tolerance of the amylosucrase mutant disclosed by the invention are remarkably improved; the recombinant bacteria constructed by using the recombinant bacteria can be used as a whole-cell catalyst to efficiently catalyze a substrate to synthesize alpha-arbutin in a reaction system containing hydroquinone. Therefore, the amylosucrase mutant disclosed by the invention has a good application prospect.

The application provides a myo-inositol-3-phosphate synthase mutant, which is obtained by mutating a myo-inositol-3-phosphate synthase with an amino acid sequence as shown in SEQ ID NO: 4 at positions 55, 57 and 273 respectively. The application also provides a preparation method of myo-inositol. The inventors have screened a wild-type myo-inositol-3-phosphate synthase and a myo-inositol monophosphatase with good performance, which can be used in cooperation with a glucan phosphorylase and a glucose phosphate mutase to catalyze starch to generate myo-inositol more efficiently. The myo-inositol-3-phosphate synthase mutant is applied to in-vitro non-fermentation biosynthesis of myo-inositol, combined with myo-inositol monophosphatases E1 and E5, and in a way of feeding whole cells, so that starch can be converted into myo-inositol in a one-pot method, without using NAD + coenzymes, and has a good industrial application prospect.

The present application relates to the field of agricultural biotechnology, in particular to fusion P450 enzyme mutants, methods for improving the catalytic activity of VD3, and methods for producing 25(OH)VD3 by whole cells, wherein a fusion P450 enzyme VK1-CYP116B46-L21 and a self-sufficient fusion P450 enzyme mutant with improved catalytic activity for synthesizing calcidiol from vitamin D3 are constructed; using VD3 as a substrate, adding glucose, a cosolvent, and a recombinant Escherichia coli expressing the fusion P450 enzyme and glucose dehydrogenase after induction, the maximum yield of 25(OH)VD3 can reach 3.26 g / L within 24 h after the catalytic reaction; directly using whole cells as catalysts without purifying enzymes, introducing glucose dehydrogenase to regenerate cofactor NAD(P)H using glucose as a substrate, avoiding the addition of expensive cofactors, reducing production costs, and being conducive to industrial production.

The invention belongs to the technical field of biological catalysis, and particularly relates to a glycosyl transferase mutant and application thereof in catalytic synthesis of rebaudioside M. Engineering bacterium whole cells or broken enzyme liquid for co-expressing a glycosyl transferase G1 mutant, a glycosyl transferase C1 mutant and sucrose synthase SUS is used as a catalyst, Reb A is used as a substrate, sucrose is used as a glycoside donor, Reb M is synthesized under mild conditions, the space-time yield of the generated Reb M is up to 3.33 g / L / h, and the method has the advantages of being high in space-time yield, convenient to purify, green, environmentally friendly and the like, and is suitable for industrial production. The industrial application value is extremely high.

This invention belongs to the field of synthetic biology and microbial engineering, and discloses a chemical-biological method for preparing the isopentenyl bibenzyl compound 2-isopentenyl-3,3',4',5-tetrahydroxybibenzyl (NPB-1575) which has anti-ischemic stroke activity. The specific disclosed method includes: (1) chemically synthesizing the substrate dihydropiperidine; (2) constructing an engineered Escherichia coli strain based on the isopentenyl utilization pathway, co-expressing hydroxyethylthiazol kinase, isopentenyl phosphokinase, isopentenyl pyrophosphate isomerase and specific isopentenyl transferase, and efficiently synthesizing the isopentenyl donor; (3) using this strain to catalyze dihydropiperidine in whole cells, the yield in a 200L fermenter can reach 2.54g / L, which is nearly 40 times higher than the existing method; (4) obtaining NPB-1575 with a purity >98.5% by macroporous adsorption resin and normal / reversed phase chromatography and recrystallization. This method uses readily available raw materials, involves simple steps, is environmentally friendly, and yields high purity, laying the foundation for the development of NPB-1575 as a new drug for ischemic stroke and showing significant application potential.

The application discloses an ecDNA recognition method based on scATAC-seq data. The ecDNA recognition method provided by the application can accurately detect copy number amplification, breakpoint quantity and ecDNA feature quantity on the whole cell level. The method provided by the application can recognize specific circular oncogenes carried in tumors, analyze the distribution characteristics of the circular oncogenes in the genome, and reveal the overall composition of all circular DNAs in a single cell and the specific composition of a single circular DNA. In addition, the method can also verify the existence of the circular DNA through a staining experiment, so that the reliability of the detection result is ensured.

The application discloses an L-aspartate alpha-decarboxylase with improved substrate tolerance, and the L-aspartate alpha-decarboxylase is derived from Bacillus subtilis and is subjected to site-directed mutation, i.e., phenylalanine at the 4th site is changed into tryptophan, isoleucine at the 33rd site is changed into alanine, and isoleucine at the 88th site is changed into tryptophan, so as to obtain an enzyme mutant; the mutant is recombined into E. coli, and after expression is induced, the mutant catalyzes the substrate L-aspartate to generate beta-alanine. When the substrate addition concentration of the whole cell catalytic system is 60 g / L, the conversion rates of the three enzyme mutants T4W, I33A and I88M can reach about 90%, and the yields are 1.2, 1.3 and 1.2 times of that of the unmutated strain respectively, and the substrate tolerance is obviously improved; the combined mutation also has a certain improvement on the substrate tolerance, and the finding has important research value for industrial preparation of beta-alanine.

The application relates to the photovoltaic technical field and discloses a back contact cell and a photovoltaic module. The back contact cell comprises a silicon wafer substrate and an electrode layer arranged on the back surface of the silicon wafer substrate, and the electrode layer comprises a plurality of parallel electrode areas. Each electrode area is provided with a first grid line group and a second grid line group. The first grid line group comprises a first bus grid line extending along a first direction and a first main grid line connected with the first bus grid line, and the end point of the first main grid line is connected with a first fine grid line. The second grid line group comprises a second bus grid line extending along the first direction and a second main grid line connected with the second bus grid line, and the end point of the second main grid line is connected with a second fine grid line. The first main grid line and the second main grid line are arranged at intervals along the first direction and are staggered along a second direction, and the first fine grid line and the second fine grid line are arranged at intervals along the second direction and are staggered along the first direction. Therefore, the influence of the electric leakage channel on the whole cell is reduced, and the open-circuit voltage and the short-circuit current of the cell are improved.

The invention discloses a novel TOPCon battery preparation process and device, and relates to the technical field of battery processing, and the novel TOPCon battery preparation process comprises the following steps: S1, preparing a sheet; s2, scribing: segmenting the whole cell by using laser; s3, side face polishing is conducted, specifically, chain type etching and alkali washing are conducted on the cutting face, and the cutting face is corroded at the temperature of 30-60 DEG C through 10-30% of TMAH and 5-20% of IPA aqueous solution; s4, edge passivation is conducted, specifically, an aluminum oxide passivation film of 20-50 nm is deposited in an atomic layer deposition mode, and annealing is conducted for 30-60 min at the temperature of 250-300 DEG C; and S5, IV testing: testing the appearance, IV and EL performance of the battery piece, and then grading. Preferably, the flaking process comprises the steps of texturing and boron diffusion. According to the invention, a battery panel is driven to move in a manner of carrying liquid through a roller, and silicon, silicon nitride and silicon oxide on a cut surface of the battery can be corroded and polished by using a TMAH + IPA solution and an HF solution.

The invention discloses a guanidinoacetic acid N-methyltransferase mutant and application of the guanidinoacetic acid N-methyltransferase mutant in synthesis of creatine, and belongs to the technical field of bioengineering. The invention provides a method for preparing creatine by utilizing a guanidinoacetic acid N-methyltransferase mutant (Y134I / N170H) and catalyzing guanidinoacetic acid and S-adenosylmethionine in a whole-cell form. The genetically engineered bacterium for expressing the guanidinoacetic acid N-methyltransferase mutant is fermented for 24 hours under the conditions that the whole cell amount is 0.2 g, the pH value is 7.5, the reaction temperature is 37 DEG C, the concentration of guanidinoacetic acid is 80 mM, and the concentration of S-adenosylmethionine is 100 mM, the creatine yield is 7.34 g / L, and the molar yield is 69.96%. According to the method disclosed by the invention, the production cycle is shortened, the yield of creatine is increased, and the industrial process of producing creatine by an enzyme conversion method is accelerated.

The present invention relates to chimeric receptors that can be used in whole-cell sensors for detecting analytes of interest. The inventors showed that the DNA binding domains and downstream gene expression can be activated via dimerization of an artificial dimerization composed of a single chain variable domain. They demonstrated for the first time that an artificial bacterial receptor using an antibody-like domain can be activated and produce a transcriptional output upon ligand-binding. In particular, the present invention relates to a chimeric receptor polypeptide comprising: i) a first DNA binding domain, ii) at least one binding domain selected from the group consisting of heavy chain variable domain, camelid VHHs, or antibody mimetics having specificity for an analyte, and iii) a linker between the DNA binding domain and the binding domain.

The application provides a non-inducible engineering strain for producing alpha-matrix and a construction method and application thereof. The engineering strain of the application is a recombinant Escherichia coli, which contains an expression regulatory element and a sucrose phosphorylase gene, and the expression regulatory element includes a PgapA-infC tandem promoter, a Plpp-infC tandem promoter or a PalsR-infC tandem promoter. The engineering strain can greatly shorten the protein expression period without adding an inducer, and the bacterial body after fermentation culture is ended is used as a whole cell catalyst to synthesize alpha-matrix by glycosylation reaction of a substrate hydroquinone and a sugar donor (such as sucrose), and the molar conversion rate of the substrate hydroquinone is high, and the engineering strain has an industrial application prospect.

The invention relates to a tracer compound for whole-cell labelling, comprising: a non-streptavidin- binding biotin analogue, at least one connecting moiety comprising a functional group configured for a click-chemistry reaction, and at least one cross-linkable moiety, preferably wherein the cross-linkable moiety is a (para)formaldehyde cross-linkable group, and the non-streptavidin-binding biotin analogue enables the tracer compound to diffuse within a biological cell with essentially the same properties as biocytin and / or neurobiotin, preferably enabling the detection of cellular protrusions, such as dendrites, dendritic spines and axons. The invention further relates to methods for constructing such tracer compound and for labelling a cell therewith. The invention further relates to kits for carrying out the method according to the invention.

Photovoltaic module (1) comprising three cell string groups connected in series, wherein each of the three cell strand groups comprises four cell strands connected in parallel (14); a cell strand comprising the four cell strands (14) cells (15) that are electrically connected to each other and that are quarter cells (15) formed by cutting a whole cell (15); and a length of the cell (15) is represented as L1 and a width of the cell (15) as L2, where 46.675 mm ≤ L2 ≤ 53.25 mm.

The invention discloses an immobilized recombinant whole-cell catalyst and a method for preparing gamma-cyclodextrin from the immobilized recombinant whole-cell catalyst. The recombinant whole-cell catalyst is prepared by immobilizing recombinant genetically engineered bacterium whole-cells for expressing a high-selectivity gamma-cyclodextrin glucosyltransferase mutant (the amino acid sequence is as shown in SEQ ID NO: 2) in a calcium alginate carrier matrix through an embedding method. The invention also provides a preparation method of the catalyst and application of the catalyst in preparation of gamma-cyclodextrin by catalyzing starch conversion in multi-batch reaction. The immobilized whole-cell catalyst is simple in preparation process and low in cost, has the high enzyme activity utilization rate of free whole cells and the excellent operation stability of immobilized enzyme, the enzyme activity retention rate is still 85% or above after the immobilized whole-cell catalyst is repeatedly used for 8 times at the temperature of 60 DEG C, and the proportion of a gamma-cyclodextrin product can be kept not lower than 96%.

A cell system and an application thereof, and a method for activating a broad-spectrum cancer cell-specific T cell. The cell system includes a cancer cell-specific T cell extracted from a tumor-infiltrating lymphocyte, where the extraction includes steps of co-incubating the tumor-infiltrating lymphocyte or a T cell in the tumor-infiltrating lymphocyte and an antigen-presenting cell with a nanoparticle and / or a microparticle loaded with a whole-cell antigen of a cancer cell to activate a cancer cell-specific T cell, and then isolating the activated cancer cell-specific T cell from the tumor-infiltrating lymphocyte. The problem that broad-spectrum and polyclonal cancer cell-specific T cells in tumor-infiltrating lymphocytes cannot be effectively screened in clinical practice at present is overcome, broad-spectrum effector cancer cell-specific T cells with a specific tumor-killing function can be isolated from the tumor-infiltrating lymphocytes, which have the characteristics of easy isolation and high specificity, and can be used for cancer prevention and treatment.

Disclosed are a method for screening a riboswitch capable of specifically recognizing doxycycline as well as rational engineering and application of the riboswitch, and belongs to the field of gene expression regulation. The present disclosure provides a doxycycline specific riboswitch, and establishes a high-throughput screening method based on flow cytometry, and obtains the doxycycline riboswitch with further improved activation fold by way of computer-aided prediction of binding sites of the riboswitch and calculation of mutation sites. According to a dose-response curve, the riboswitch shows linear correlation at 40-100 μg / L, and meanwhile, a whole-cell sensor constructed is low in price, high in stability and simple to operate, and can be used for on-site rapid detection.

This invention discloses a method for hydroxylation modification of recombinant mussel foot protein and its application. The invention utilizes a whole-cell catalyst with tyrosinase catalytic activity to hydroxylate tyrosine residues in recombinant mussel foot protein. This method eliminates the need for cell disruption and protein purification steps. By immobilizing tyrosinase on the host cell surface through cell surface display, the recombinant mussel foot protein can directly contact the tyrosinase in the extracellular environment and undergo hydroxylation, thereby introducing dopamine groups into the protein to improve its underwater adhesion properties. This method is applicable to the hydroxylation modification of various types of recombinant mussel foot protein and has advantages such as high catalytic efficiency, good stability, simple operation, and low cost, making it valuable for industrial applications.