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54 results about "Insulin Precursor" patented technology
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Method for purifying gene-recombinant insulin precursor
ActiveCN101029077AHigh recovery rateHigh purityFungiPeptide preparation methodsActivated carbonPichia pastoris
A process for purifying the genetically recombined insulin precursor features that the activated carbon as adsorbent and cationic chromatography are used to remove the pigment from fermented liquid and purify the active protein. Its recovery rate is more than 85% and its purity is more than 95%.
Owner:YICHANG HEC CHANGJIANG PHARMA CO LTD
Processes for making acylated insulin
InactiveUS7402565B2High cracking ratePeptide/protein ingredientsPeptide sourcesYeastProteinase activity
Owner:NOVO NORDISK AS
Expression of a human insulin precursor in P. pastoris
Methylotrophic recombinant yeast strain producing human insulin precursor, the strain comprising, in its genome, a copy of a first DNA construction and a second DNA construction, wherein the constructions are capably of directing the expression and secretion of human insulin precursor of the formula B(1-30)-Y1-Y2-A(1-21), wherein Y1 is lysine or arginine; Y2 is lysine or arginine; B(1-30) is the B peptide of the human insulin; and B(1-21) is the A peptide of human insulin, and wherein the strain is yeast Pichia Pastoris. DNA constructions and method for obtaining the strain are also provided.
Owner:LAB BETA SA
Method for high-density fermentation of pichia pastoris of insulin precurosor protein
ActiveCN106282274AReduce contentGood removal effectMicroorganism based processesInsulinsPichia pastorisGrowth phase
The invention relates to the field of microorganisms and discloses a method for high-density fermentation of pichia pastoris of insulin precurosor protein. The method comprises the steps of expanding culture of strain, fermentation culture and thallus collection. Fermentation culture is divided into a batch growth phase, a glycerinum replenishment phase and a methyl alcohol induction phase. By controlling parameters including age, replenishing rate and pH value of strain during fermentation, the concentration of glycosylation 2 can be effectively reduced by 60+ / -8%, and the unit yield of insulin precurosor protein can be increased by 25% or above. Furthermore, the method has the advantages of being simple, easy to operate, low in cost and the like.
Owner:YICHANG HEC CHANGJIANG PHARMA CO LTD
Purification and enzyme digestion transformation method of recombinant human insulin precursor
ActiveCN105111304ASimplified purification stepsImprove the purity yieldPeptide preparation methodsInsulinsProtein targetEnzyme digestion
The invention discloses a purification and enzyme digestion transformation method of a recombinant human insulin precursor and belongs to the field of preparation of recombinant human insulin and analogues thereof. In the invention, fermentation supernatant of a secretorily expressed recombinant human insulin precursor is purified by using a cation exchange chromatography column, two different washing solutions are used during washing of chromatography, the elution of target proteins is performed more quickly, the elution time is shortened, the precipitation of the target proteins is reduced and the volume of eluted samples is reduced; an eluting solution with pH 7.0 to 9.0 is used during elution, phase change operation does not need to be performed to eluted products, the eluted products are directly added into trypsin for enzyme digestion transformation and the transformation efficiency is high. The method disclosed by the invention is simple to operate, the primary purification and phase change operation of the fermentation supernatant are completed through one step of chromatography purification, the purity of the samples can be improved from 14 percent to 95 percent, the purified samples are directly subjected to enzyme digestion, the enzyme digestion transformation efficiency is above 95 percent, the production process is simplified and the cost is saved.
Owner:SHANDONG EHUA BIOLOGICAL PHARMA +1
Processes for making acylated insulin
InactiveUS20060264606A1High cracking rateIncrease cleavage ratePeptide/protein ingredientsPeptide sourcesInsulin PrecursorPancreatic hormone
A method is provided which allows high yields of acylated insulin. The method comprises expressing a singe-chain insulin precursor, preferably in yeast, cleaving the single-chain insulin precursor with a suitable protease which will open the peptide bond between the C-terminal amino group in the B-chain and a connecting peptide connecting the B chain with the A-chain, acylating the two-chain insulin intermediate in the ε-amino group in LysB29 and subjecting the acylated two-chain insulin intermediate to a proteolytic enzyme which will cleave of the N-terminal extension on the B- and A-chains on the precursor molecule.
Owner:NOVO NORDISK AS
Quick-acting insulin aspart precursor protein and preparation method for quick-acting insulin
InactiveCN105418755AHigh expressionStable structureFungiMicroorganism based processesInsulin aspartInsulin Precursor
The invention provides an insulin aspart precursor. The amino acid sequence is Z1(XY)nZ2Z3Z4-B(1-29)-C1C2C3-A(1-21), wherein Z1, X and Z2 are Asp or Glu respectively, Y is Ala or Gly, n is an integer from 1-10, Z3 is Pro, Glu or Asp, Z4 and C3 are Lys or Arg respectively, C1 and C2 are Glu, Asp, Ala or Gly respectively, B(1-29) is insulin aspart B chain 1-29 bit amino acid, and A(1-21) is insulin aspart A chain 1-21 bit amino acid. The invention also provides a coding gene, a cloning vector or an expression vector containing the coding gene, a transformant, and a method for preparing quick-acting insulin by utilization of the quick-acting insulin aspart precursor protein. The expression level of the product is high, the fermentation expression product purity is high, the extraction method is simple and the yield is high.
Owner:ZHUHAI JINBAIKANG BIOLOGICAL TECH CO LTD
Inter mass optimization during insulin precursor fermentation
InactiveCN101029323AGrowth adjustmentIncrease concentrationMicroorganism based processesFermentationInsulin PrecursorFermentation
A method for supplementing and optimizing feed during fermentation of insulin precursor is carried out by supplementing methanol to determine its fluid acceleration ratio, computing while adjusting for specific growth speed ration by difference of final fermented strain density and real-time strain density and fermentation period and controlling growth speed ratio. It has more protein concentration.
Owner:SUNSHINE LAKE PHARM CO LTD
Method for producing human recombinant insulin
InactiveUS20120058513A1Increase productionQuality improvementPeptide/protein ingredientsRecombinant DNA-technologyInclusion bodiesInsulin Precursor
The invention relates to biotechnology and can be used for producing human recombinant insulin for preparing medicinal agents for the treatment of pancreatic diabetes. A variety of recombinant plasmid DNAs which contain an artificial gene and encode the human insulin precursor is proposed. The biosynthesis of a hybrid polypeptide is induced using isopropyl-thiogalactopyranoside so that the post-induction level of the hybrid polypeptide is equal to or greater than 25% of the total cellular protein. According to the claimed procedure, human insulin is produced by cultivating a producer strain containing one of the recombinant plasmids, isolating inclusion bodies, solubilizing and renaturing the fusion protein, and enzymatically degrading and chromatographically purifying said protein. The invention simplifies the process for producing human recombinant insulin and increases the yield thereof.
Owner:LIABILITY MAKO
Codon-optimized insulin degludec precursor gene and expression method thereof
ActiveCN109295067AHigh expressionIncrease productionFungiPeptide/protein ingredientsInsulin PrecursorInsulin degludec
The invention relates to the technical field of genetic modification and protein expression, particularly to a codon-optimized insulin degludec precursor gene and an expression method thereof, whereinthe insulin degludec precursor gene is optimized based on the gene sequence represented by SEQ ID NO:1, and the optimizations comprise one or a plurality of the following cases that (a) the amino acid codon CAG at the site 4 is replaced with CAA; (b) the amino acid codon CAC at the site 5 is replaced with CAT; (c) the amino acid codon GTC at the site 12 is replaced with GTT; and (d) the amino acid codon CGT at the site 22 is replaced with AGA. According to the present invention, the amino acid sequence encoded by the optimized insulin degludec precursor gene sequence is consistent with the original gene sequence while the expression level of the gene sequence in yeast is much higher than the expression level of the original gene sequence.
Owner:JILIN HUISHENG BIOPHARMACEUTICAL CO LTD
Recombinant insulin and insulin analogue precursor purification method
InactiveCN105153294AGood technical effectAvoid time costPeptide preparation methodsInsulinsPurification methodsCentrifugation
The invention relates to the field of insulin production methods and discloses a recombinant insulin and insulin analogue precursor purification method. The invention solves the problems that conventional chromatographic packing cannot tolerate high-salinity sample loading, the amount of used organic reagent is great, the cost is high and the product purity is not high in the purification process of recombinant expressed insulin precursors and insulin analogue precursors. The invention adopts the technical scheme that the method comprises the steps of performing pH regulation and centrifugation to centrifuged fermentation supernatant, then directly loading a sample and performing adsorption, separation, purification and elution through a chromatographic column prepared by using any packing of Capto S, Capto MMC, Uni PMM S and Uni MSP to finally obtain high-purity recombinant insulin and insulin analogue precursors. Compared with the existing purification method, the recombinant insulin and insulin analogue precursor purification method has the advantages that the operation is simple, the yield is high, the spent time is short, the environmental influence is small, and the product production cost of the existing insulin and insulin analogues can be greatly reduced.
Owner:JINAN KANGHE MEDICAL TECH
Novel process for genetic engineering preparation of insulin and insulin analogs
The invention provides a novel process for genetic engineering preparation of insulin and insulin analogs, which comprises enzyme cutting insulin precursor with parenzyme, then isolating insulin, wherein the insulin precursor comprises the following elements from amino end to carboxyl end, (1) spacing peptide element with formula (Xaa)mZ, (2) insulin B chain element, (3) joint peptide element with formula (Xaa)nZ, and (4) insulin A chain element.
Owner:费俭
Preparation method of insulin
The invention provides a preparation method of insulin, relates to a method for transforming an insulin precursor into an active insulin compound, preferably a method for improving long-acting insulin; and particularly provides a method, wherein lysine residues in Arg-Arg precursor insulin can be reversibly closed, and parenzyme can cut the reversibly closed Arg-Arg precursor insulin.
Owner:VALIN TECH
Human insulin/analogue conjugate with continuous blood sugar reduction function and high rate of combination with receptor
ActiveCN102675452AUnderstand clearlyPeptide/protein ingredientsMetabolism disorderInsulin activityHigh rate
The invention relates to a human insulin / analogue conjugate with a continuous blood sugar reduction function and a high rate of combination with a receptor, and discloses a method for preparing a reconstructed human insulin / analogue conjugate by reconstructing a human insulin precursor by utilizing a methylotrophic yeast expression, performing purification, enzyme digestion, chromatography or synthesis to prepare a reconstructed human insulin and a DesB30 analogue thereof and coupling a side chain amino group of a 29th-bit lysine of a B chain of the reconstructed human insulin precursor and a polyethyleneglycol acetamide bond. The prepared reconstructed human insulin conjugate has the characteristics of high rate of combination with the insulin receptor, remarkable continuous blood sugar reduction effects and long in-vivo half-life period, and is applied to the treatment of I-type and II-type diabetes.
Owner:CHONGQING FAGEN BIOMEDICAL +1
Chimeric protein containing an intramolecular chaperone-like sequence
InactiveUS20020164712A1Improves insulin precursor foldingEasy to foldHydrolasesPeptide/protein ingredientsInsulin PrecursorAmino acid
The present invention relates to a chimeric protein containing an intramolecular chaperone (IMC) like sequence linked to a target protein, preferably an insulin precursor. The present invention also relates to a process for obtaining a correctly folded insulin-precursor-containing chimeric protein, comprising, inter alia, contacting an incorrectly folded chimeric protein containing an IMC like sequence linked to an insulin precursor with at least one chaotropic auxiliary agent. The present invention further relates to an assay for screening an amino acid sequence for the ability to improve folding of an insulin precursor using a chimeric protein containing an IMC like sequence linked to an insulin precursor.
Owner:TONGHUA GANTECH BIOTECH
Process for preparing insulin compounds
ActiveUS7396903B2Short reaction timeLow amountPeptide/protein ingredientsPeptide preparation methodsCompound aOrganic solvent
A preferred way of converting insulin precursors into insulin compounds is to perform an enzymatic peptide cleavage in an aqueous medium and, thereafter, without removal of the intermediate product formed, to add an amino acid ester or a peptide ester and an organic solvent so that the desired coupling takes place.
Owner:NOVO NORDISK AS
New monomeric insulin and its medicinal composition and prepn process
InactiveCN1468864AEasy to makeSuitable for mass productionPeptide/protein ingredientsMetabolism disorderYeastInsulin activity
Owner:SHANGHAI C A S SHENGLONGDA BIOTECH GRP
Preparation method of insulin detemir or insulin detemir analogue
ActiveCN105440125AEasy sample loading for purificationAcylation reaction time is shortInsulinsThreonineCombinatorial chemistry
The invention discloses a preparation method of insulin detemir or an insulin detemir analogue and belongs to the field of preparation of insulin analogues. According to the preparation method, threonine-free 30th-site human double-chain insulin precursor desB30 protein of a B chain is taken as a raw material, epsilon-amino of B29th-site lysine is selectively acylated by active ester through one step in the alkaline environment with pH ranging from 8.0 to 11 under the condition that N-terminal alpha-amino of double chains of the desB30 protein is not protected, and the insulin detemir or the insulin detemir analogue is generated efficiently. According to the preparation method of the insulin detemir or the insulin detemir analogue, parameters such as the pH value, the raw material ratio, the acylation time and the like of the acylation reaction of the insulin precursor are optimized, the yield of the insulin detemir or the insulin detemir analogue is remarkably increased, the preparation method has the advantages of high acylation efficiency, a few by-products, short acylation reaction time, a few acylation steps, simple process, low production cost and the like, and one simple and efficient method is provided for large-scale industrial preparation of the insulin detemir or the insulin detemir analogue through one-step acylation.
Owner:SHANDONG EHUA BIOLOGICAL PHARMA
Preparation method of recombinant human insulin
The invention discloses a preparation method of recombinant human insulin. The preparation method comprises the following steps of: performing an enzyme digestion reaction on a recombinant human insulin precursor in a mixed solution of an organic solution and water to obtain a human insulin solution containing desB30; centrifuging or suction filtrating the human insulin solution containing desB30 to obtain water-containing powder of desB30 human insulin; performing a transpeptidation reaction on the water-containing powder of desB30 human insulin to obtain human insulin ester, wherein the final concentration of the desB30 human insulin in a transpeptidation reaction system is 12.1mM-33mM, the mol ratio of the desB30 human insulin to threonine ester or a derivative thereof is 1:(15-150), and the mass ratio of the desB30 human insulin to trypsin is (10-80):1; degreasing the recombinant human insulin ester to obtain the recombinant human insulin. The method is suitable for industrial production of converting yeast-expressed proinsulin into the human insulin, high in yield, simple to operation and low in cost.
Owner:ZHUHAI UNITED LAB
Chaperone protein containing an intramolecular chaperone-like sequence and its application to insulin production
InactiveUS6924120B1Improves insulin precursor foldingEasy to foldFungiBacteriaProtein targetChimera Protein
The present invention relates to a chimeric protein containing an intramolecular chaperone (IMC) like sequence linked to a target protein, preferably an insulin precursor. The present invention also relates to a process for obtaining a correctly folded insulin-precursor-contain chimeric protein, comprising, inter alia, contacting an incorrectly folded chimeric protein containing an IMC like sequence linked to an insulin precursor with at least one chaotropic auxiliary agent. The present invention further relates to an assay for screening an amino acid sequence for the ability to improve folding of an insulin precursor using a chimeric protein containing an IMC like sequence linked to an insulin precursor.
Owner:BEIJING YUANHEGENZE TECHOLOGY CO
Method for purifying gene-recombinant insulin precursor
ActiveCN101029077BHigh recovery rateHigh purityFungiPeptide preparation methodsActivated carbonPichia pastoris
A process for purifying the genetically recombined insulin precursor features that the activated carbon as adsorbent and cationic chromatography are used to remove the pigment from fermented liquid and purify the active protein. Its recovery rate is more than 85% and its purity is more than 95%.
Owner:YICHANG HEC CHANGJIANG PHARMA CO LTD
Insulin precursor precipitation recovery method for removing amino acid residue at 30th of chain B
ActiveCN108164594AImprove sedimentation efficiencyHigh yieldPeptide preparation methodsInsulinsRecovery methodDivalent metal ions
The invention provides an insulin precursor precipitation recovery method for removing amino acid residue at 30th of a chain B. The method comprises carrying out digestion on an insulin precursor through trypsin at 30 DEG C for 1-2h, adding divalent metal ions into a reaction system subjected to digestion, adjusting pH of the reaction system to close to the isoelectric point of the reaction product, and collecting the precipitates, wherein the divalent metal ions comprise water-soluble divalent metal ions of Ni<2+>, Ca<2+>, Mg<2+>, Zn<2+>, Cu<2+>, Fe<2+> and Mn<2+>, a mole ratio of the divalent metal ions to the insulin precursor is in a range of 0.5:1 to 4:1, pH is in a range of 4.0 to 6.0, the precipitation temperature is 0-30 DEG C and the precipitation time is 0-12h. The method is freeof treatment on a digestion reaction system, realizes direct isoelectric point precipitation and has simple processes. Compared with the prior art, the method reduces the residual rate of the productin the supernatant by 10 times or more. The insulin precursor precipitation recovery method has obvious advantages.
Owner:ZHUHAI JINBAIKANG BIOLOGICAL TECH CO LTD
Preparation method of insulin
InactiveCN102628072AFull effectImprove digestion efficiencyPeptide preparation methodsInsulinsEnzymatic digestionInsulin products
The invention relates to a preparation method of insulin. The method which comprises two steps of chromatography purification can improve the enzymatic digestion efficiency, and the controllability in an enzymatic digestion process, so a complete reaction of trypsin and carboxypeptidase B is realized, and insulin precursors not reacted with the carboxypeptidase B can be thoroughly removed, so the purity and the quality of final insulin products are improved, the purity of the final insulin products can reach above 99.7%, and the recovery rate is high, thereby industrialized production can be carried out.
Owner:GAN&LEE PHARMA
Supplemented culture medium of proinsulin produced by fermentation and supplemented culture optimization method
ActiveCN101643764AEasy to control growth rateControl growth rateMicroorganism based processesFermentationMicroorganismEscherichia coli
The invention relates to a supplemented culture medium of proinsulin produced by fermentation and a supplemented culture optimization method, belonging to the field of microorganism engineering. Aiming at the defects of low fermentation density and expression amount, and the like of the prior proinsulin-presenting process by colon bacilli, the invention provides a supplemented culture medium containing a DMEM-high-glucose culture medium and a batch supplementation optimization method for applying the supplemented culture medium to proinsulin fermented by colon bacilli. The culture method has easily controlled process and strong universality and improves the supplemented culture medium of a proinsulin precursor presented by colon bacilli and the supplemented culture optimization method.
Owner:LUNAN PHARMA GROUP CORPORATION
Enzyme digestion conversion method of recombinant human insulin precursor
PendingCN112625116AHigh purityReduce difficultyInsulinsFermentationEnzyme digestionInsulin Precursor
Owner:SHANDONG EHUA BIOLOGICAL PHARMA +1
Genetically engineered bacterium for expressing insulin precursor, and preparation method and application of genetically engineered bacterium
PendingCN111662836AIncrease productionLow costFungiMicroorganism based processesPichia pastorisNucleotide
The invention provides a genetically engineered bacterium for expressing an insulin precursor. The genetically engineered bacterium is an engineered bacterium in which an expression vector of an insulin precursor gene is integrated in pichia pastoris, and the nucleotide sequence of the insulin precursor gene is shown as SEQ ID NO.1 in a sequence table. The invention also provides a preparation method of the insulin precursor. The strain yield of the genetically engineered bacterium fermented at a shake flask level is higher than the strain yield reported in the prior art. Through combination of the genetically engineered bacterium and the preparation method of the insulin precursor, the yield of the insulin precursor obtained by fermentation tank-level fermentation is remarkably increased.Besides, a culture medium used in the preparation method of the insulin precursor is relatively low in cost, and the risk of animal-derived virus pollution is avoided, so that cost of the preparationmethod is remarkably reduced.
Owner:SHANGHAI INST OF PHARMA IND +1
Purification and enzymatic conversion method of recombinant human insulin precursor
ActiveCN105111304BFast elutionReduce precipitationPeptide preparation methodsInsulinsProtein targetEnzyme digestion
Owner:华润昂德生物药业有限公司 +1
Fermentation medium of pichia pastoris for improving expression of insulin precursor and fermentation method
ActiveCN111411049AClear compositionThe composition is clear and the source is clearFungiMicroorganism based processesPichia pastorisPhosphate
The invention belongs to the technical field of biopharmaceuticals, and discloses a fermentation medium of pichia pastoris for improving the expression of insulin precursor and a method for fermentedproduction of the insulin precursor. The fermentation medium of pichia pastoris in the present invention adopts a phosphate buffer system, supplemented by inorganic salts such as ammonium sulfate andferrous sulfate, as well as nitrogen sources such as urea and ammonium dihydrogen phosphate. The fermentation medium has definite compositions with stable sources and does not require fed-batch of nitrogen sources in the induction stage. According to the method for fermented production of the insulin precursor, a glycerol / methanol mixed solution is adopted for induction of culture after initial culture and glycerol fed-batch culture. A certain proportion of glycerol is fed by batches in the induction stage, so as to accelerate the growth of microorganisms. As time increases, the mixing ratio of glycerol gradually decreases while methanol gradually increases, and a large number of microorganisms enter the high-speed expression period in a short time, so that a target product can obtain a higher level of expression, greatly shortening the culture time and avoiding influence on product quality caused by excessive by-products.
Owner:BEIJING HUIZHIHENG BIOTECHNOLOGY CO LTD +1
Supplemented culture medium of proinsulin produced by fermentation and supplemented culture optimization method
ActiveCN101643764BEasy to control growth rateControl growth rateMicroorganism based processesFermentationMicroorganismEscherichia coli
Owner:LUNAN PHARMA GROUP CORPORATION
Method for preparing insulin or insulin derivative precursor
ActiveCN109735589AAvoid issues that affect grooming efficiencyEasy to operatePeptide preparation methodsInsulinsInsulin PrecursorDigestion
The invention discloses a method for preparing insulin or an insulin derivative precursor. The method comprises the following steps that S1, protease is added to fermentation broth of insulin precursor protein; S2, the pH value and temperature of the solution treated in S1 are adjusted, so that protease is subjected to digestion, and a solution containing the insulin or insulin derivative precursor is obtained after digestion is carried out. The method for preparing the insulin or insulin derivative precursor can further comprises the step that S3, the solution containing the insulin or insulin derivative precursor obtained after digestion is carried out is subjected to isoelectric precipitation recycling operation, and the higher-purity insulin or insulin derivative precursor is obtained.Compared with the prior art, according to the scheme, the method for preparing the insulin or insulin derivative precursor is easy and convenient to operate, low in equipment investment and high in technological process yield and has good industrial application prospects.
Owner:ZHUHAI JINBAIKANG BIOLOGICAL TECH CO LTD
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