38 results about "Deamidation" patented technology

The invention belongs to the technical field of food, and provides a special enzyme composition for infant soybean milk powder and soybean milk powder thereof. A three-stage directional enzymolysis technology is adopted, the three-stage directional enzymolysis technology comprises glutamine transaminase deamidation pretreatment, bacillus subtilis protease-trypsin-papain first-stage hydrolysis and aminopeptidase-phytase-beta-glucosidase second-stage hydrolysis, and conductivity control, carbon dioxide ventilation in-situ complexation and spray drying processes are combined. The free amino nitrogen of the product is increased by more than or equal to 20%, the proportion of 200-1500Da peptide fragments is more than or equal to 70%, the median particle size is 50-180 microns, the water activity is less than or equal to 0.30, the instant dissolving time is less than or equal to 30 seconds at 25 DEG C under the water hardness of 15-200ppm, the settling rate is less than or equal to 3% after standing for 30 minutes, and the zeta potential is-10 to-25mV. The palatability contradiction between deep hydrolysis and bitterness inhibition of the bean-based infant formula powder, the brewing contradiction between high-calcium reinforcement and instant stability and the process controllability contradiction between three-stage enzymolysis and ionic strength accumulation are effectively solved, and the bean-based infant formula powder has wide application value in the fields of bean-based infant formula food and vegetable protein nutritional products.

The invention discloses a preparation method and application of a myofibrillar protein emulsion based on enzymatic modification. According to the technology, a myofibrillar protein aqueous suspension is subjected to enzymatic deamidation and glycosylation reaction through protein glutaminase and glucan, a protein-polysaccharide compound with good emulsifying performance is constructed, soybean oil serves as an oil phase, and emulsion systems with different oil phase volume fractions are constructed through a controllable homogenizing technology. The obtained emulsion has the characteristics of natural degradability, excellent rheological property, uniform particle size distribution, long-term physical stability and the like. Compared with a traditional chemical modification technology, the technology adopts a whole bio-based raw material combined enzyme method green technology, the problems of organic solvent residues and the like are effectively solved, and the production cost is reduced while the product safety is ensured. The emulsion has remarkable application advantages in the fields of functional food active ingredient delivery, drug controlled release carriers, cosmetic transdermal absorption systems and the like, and a new thought is provided for development of a biomacromolecule-based delivery system.

This invention relates to the field of lyophilized formulation technology, specifically disclosing an esupagglutide α-nano-lyophilized micelle and its preparation method. The micelles of this invention utilize polylactic-co-hydroxyacetic acid copolymer and sodium phosphatidylglycerol as micelle carriers, with arginine and cysteine ​​providing branching synergistic effects. Homogenization via film formation and low-temperature lyophilization yield lyophilized nano-micelle powder. The nano-micelle powder reduces protein oxidative decomposition during lyophilization, decreases impurity fragments, and reduces various non-fucose glycosylation, basic components (succinimide, methionine oxidation products), N-terminal deletion, Asn deamidation, and Met and Trp residue oxidation impurities that affect product quality. Product quality analysis and animal experiments after reconstitution show good resistance to impurities and degradation risk. The reconstituted esupagglutide α-nano-micelle system exhibits better stability and sustained-release properties, enhancing the hypoglycemic sustained-release effect 72 hours after injection, significantly improving upon conventional non-micelle formulations.

Methods of differentiating and quantifying modifications in the tau protein in bio-fluids to detect, classify, and treat a plurality of tauopathies are disclosed. The disclosed methods include liquid chromatographic mass spectrometric analyses on isolated tau protein fragments to identify and quantify deamidated or isomerized asparagines or aspartic acids indicative of a neuropathology associated with a tauopathy.

The invention discloses a method for deamidating and modifying peanut protein by using protein glutaminase, which comprises the following specific steps of: adding acetic acid / boric acid / NaOH buffer solution into peanut protein serving as a raw material while stirring to form protein dispersion liquid with certain substrate concentration; adding food-grade protein glutaminase into the peanut protein dispersion liquid for modification and termination; and carrying out vacuum freeze drying on the modified peanut protein dispersion liquid to obtain the deamidated modified peanut protein. Compared with unmodified peanut protein, the modified peanut protein obtained by the method disclosed by the invention has the advantages that the deamidation degree is obviously improved, the hydrolysis degree is not changed, and the functional properties such as solubility and foamability are all improved. The method for deamidating and modifying the peanut protein is simple in process and high in safety, can be used for large-scale industrial production and application, does not generate side reactions such as hydrolysis and crosslinking, and is beneficial to expanding the application scene of the peanut protein.

The invention relates to a Chryseobacterium-derived protein glutaminase PG3, a coding gene and an application thereof, and belongs to the technical field of biological genetic engineering. The amino acid sequence of the protein glutaminase PG3 is as shown in SEQ ID No. 1, and the cDNA sequence of the coding gene of the protein glutaminase PG3 is as shown in SEQ ID No. 2. A large amount of protein glutaminase PG3 from Chryseobacterium is prepared through an Escherichia coli expression system, and the protein glutaminase is used for carrying out enzymatic deamidation modification on the spotted silver carp myofibrillar protein PG3, so that the solubility of the spotted silver carp myofibrillar protein under a salt-free system condition is remarkably improved. The invention provides an important tool enzyme for protein deamidation modification, and lays a theoretical foundation for development of low-salt fish protein processed products.

Methods of differentiating and quantifying modifications in the tau protein in bio-fluids to detect, classify, and treat a plurality of tauopathies are disclosed. The disclosed methods include liquid chromatographic mass spectrometric analyses on isolated tau protein fragments to identify and quantify deamidated or isomerized asparagines or aspartic acids indicative of a neuropathology associated with a tauopathy.

The present application provides novel adeno-associated virus (AAV) vectors, AAV vectors with reduced capsid deamidation, and uses thereof. A recombinant adeno-associated virus (rAAV) vector includes an AAV capsid having a heterogeneous population of vp1 protein, a heterogeneous population of vp2 protein, and a heterogeneous population of vp3 protein. In particular, the present invention relates to a recombinant VP1 amino acid sequence comprising a capsid comprising a modified amino acid as compared to the encoded VP1 amino acid sequence, the capsid comprising a highly deamidated asparagine residue at an asparagine-glycine pair, and further comprising a plurality of other less deamidated asparagine residues, and optionally a glutamine residue.

The invention provides a protein glutaminase mutant with improved thermal stability and application thereof, and relates to the technical field of enzyme engineering. According to the invention, a female parent sequence of protein glutaminase is modified, the modification comprises saturated mutation and combinatorial mutation of 30th serine and 108th serine with the female parent sequence such as SEQ ID NO.1, the relative enzyme activity of the screened mutant and the residual enzyme activity after treatment at 60 DEG C are obviously improved compared with those of a wild type, and the mutant shows higher thermal stability. The residual enzyme activity of the combined mutant S30F / S108Q at the temperature of 60 DEG C is improved by 436.4% compared with that of a wild type, the deamidation degree of the combined mutant S30F / S108Q to soybean protein at the temperature of 55 DEG C is improved by 18.5% compared with that of the wild type, the problems that in the prior art, the thermal stability of protein glutaminase is poor, and industrial production and application cannot be met are solved, and the combined mutant S30F / S108Q has good application prospects in the fields of food processing, feed production and the like.

The invention provides a protein glutaminase mutant with synchronously improved enzyme activity and thermal stability and application of the protein glutaminase mutant, and belongs to the field of gene engineering and enzyme engineering. According to the invention, a series of mutants MT1-MT6 are obtained by carrying out molecular modification on a coding gene of protein glutaminase PG3 from Chryseobacterium sp. Compared with a wild type, the mutant MT6 has the advantages that the optimal catalytic temperature of the mutant MT6 is increased by 5 DEG C, the specific activity and the specific activity t65 DEG C 1 / 2 are 106.1 U / mg and 76.7 min respectively and are 2.69 times and 2.99 times of the specific activity t65 DEG C 1 / 2 of the wild type, the purpose of simultaneously improving the catalytic activity and the thermal stability of the protein glutaminase PG3 is achieved, and the application potential of the protein glutaminase in the industrial field is expanded. In addition, the mutant can significantly improve the foamability and stability of the myofibrillar protein of the spotted silver carp, and provides a potential enzyme resource for enhancing the functional characteristics of the protein through enzymatic deamidation modification.

The invention provides a protein glutaminase mutant as well as a coding gene and application thereof, and belongs to the field of gene engineering and enzyme engineering. According to the invention, a series of mutants MT1-MT8 are obtained by carrying out molecular modification on a coding gene of chitophage protein glutaminase PG5. Compared with a wild type, the mutant has the advantages that the optimal catalytic temperature is increased by 5 DEG C, the specific activity and t55 DEG C 1 / 2 are 31.1 U / mg and 216.2 min respectively and are 2.99 times and 3.57 times that of the wild type, and the application potential of protein glutaminase in the industrial field is expanded. Besides, the mutant can significantly improve dissolution of the myofibrillar protein of the spotted silver carp under the condition of a salt-free system, provides a potential enzyme resource for enzymatic deamidation modification and enhancement of protein functional characteristics, and has a wide market application prospect.

To provide a novel protein deamidation enzyme (protein asparaginase) that catalyzes a reaction of deamidation of an asparagine residue contained in a protein.SOLUTION: A protein characterized by having activity to catalyze a reaction in which an asparagine residue in a protein is deamidated, and by having one β-barrel domain at the C-terminus or having no β-barrel domain at the C-terminus.SELECTED DRAWING: Figure 1

The invention relates to the technical field of food processing, in particular to a modification method for improving solubility of soybean protein. The method comprises the following steps: carrying out heat treatment on a first solution containing soybean protein and inorganic acid; and performing cold plasma treatment on the first solution subjected to heat treatment to obtain a second solution. According to the physical and chemical synergistic superposition modification method provided by the embodiment of the invention, the cold plasma treatment is performed after the protein is subjected to deamidation modification by using the inorganic acid, so that the dissolvability is remarkably improved on the basis of not prolonging the deamidation reaction time, and the dissolvability is greatly improved. And moreover, the problem of protein molecular weight change caused by excessive deamidation is effectively solved, the deamidation reaction time is shortened, and industrial production is promoted.

The present invention provides isolated monoclonal antibodies that specifically bind LAG-3, and have optimized functional properties compared to previously described anti-LAG-3 antibodies, such as antibody 25F7 (US 2011 / 0150891 A1). These properties include reduced deamidation sites, while still retaining high affinity binding to human LAG-3, and physical (I.e., thermal and chemical) stability. Nucleic acid molecules encoding the antibodies of the invention, expression vectors, host cells and methods for expressing the antibodies of the invention are also provided, as well as immunoconjugates, bispecific molecules and pharmaceutical compositions comprising the antibodies. The present invention also provides methods for detecting LAG-3, as well as methods for treating stimulating immune responses using an anti-LAG-3 antibody of the invention. Combination therapy, in which the antibodies are co-administered with at least one additional immunostimulatory antibody, is also provided.

The invention relates to chitophage protein glutaminase PG5, a coding gene and application of the chitophage protein glutaminase PG5, and belongs to the technical field of biological genetic engineering. The amino acid sequence of the protein glutaminase PG5 is as shown in SEQ ID No. 1, and the cDNA sequence of the coding gene of the protein glutaminase PG5 is as shown in SEQ ID No. 2. A large amount of chitophage-derived protein glutaminase PG5 is prepared through an escherichia coli expression system, and the protein glutaminase PG5 is used for carrying out enzymatic deamidation modification on the myofibrillar protein of the spotted silver carp, so that the thermal reversible protein gel is successfully prepared in a salt-free system. An important tool enzyme is provided for protein deamidation modification, unprecedented flexibility is provided for processing adaptability and cold-chain storage and transportation of fish protein gel products, and the application range of the fish protein gel products in food design is greatly expanded.

The invention belongs to the technical field of aquatic product processing, and provides minced fillet soft gel and a preparation method thereof. The minced fillet soft gel is prepared from the following raw materials in parts by weight: 96-99 parts of minced fillet, 0.1-0.3 part of protein glutaminase, 1-2 parts of table salt and 0.5-1 part of composite phosphate. Protein glutaminase is added in the preparation process of the minced fillet gel, so that the hardness of the gel can be remarkably reduced, the moisture retention capability of the gel can be greatly improved, and the requirements of the special soft minced fillet gel for patients with dysphagia on texture and moisture are met; meanwhile, ultrasonic-assisted deamidation treatment is introduced, so that the working efficiency of the enzyme can be improved, the enzyme addition amount can be reduced to save the production cost, the gel hardness and the gel strength of the minced fillet can be further reduced, the water holding capacity is enhanced, the digestion characteristic of the minced fillet is improved, and finally, an easy-to-swallow food rich in high-quality protein is provided for people with dysphagia.

To provide compositions comprising AAV-based constructs for delivery of heterologous molecules which have stable receptor binding and / or stable capsid, avoid neutralizing antibodies and / or retain purity during storage.SOLUTION: A recombinant adeno-associated virus (rAAV) vector comprises an AAV capsid having a heterogeneous population of vp1 proteins, a heterogeneous population of vp2 protein and a heterogeneous population of vp3 proteins. The capsid contains modified amino acids as compared to the encoded VP1 amino acid sequence, the capsid containing highly deamidated asparagine residues at asparagine-glycine pair, and further comprising multiple other, less deamidated asparagine and optionally glutamine residues.SELECTED DRAWING: Figure 1A

Provided herein are methods for determining the serotype of a virus particle and / or or determining the heterogeneity of a virus particle (e.g., an AAV particle). In other embodiments, the invention provides methods to determine the heterogeneity of AAV particles. In some aspects, the invention provides viral particles (e.g., rAAV particles) with improved stability and / or improved transduction efficiency by increasing the acetylation and / or deamidation of capsid proteins.

The present invention provides deamidase compositions comprising a deamidase inhibitor, wherein the deamidase exhibits less self-deamidation and / or modification to other enzymes after storage.

Deamidation is usually viewed as a post-translational modification that sets an expiration date on proteins. Among apoptosis regulators of the Bcl-2 family, Bcl-xL shows a unique eligibility to be either singly or doubly deamidated. The inventors therefore analysed Bcl-xL deamidation state in platelets from mice models where platelets lifespan was manipulated. In parallel, the inventors compared human platelets obtained at steady state from healthy controls, to platelets newly synthesized after recovery from acute thrombocytopenia: they found that while expression levels of Asn52 monodeamidated Bcl-xL remains unchanged, Asn52Asn66 doubly-deamidated Bcl-xL is virtually absent in young platelets and accumulates in old platelets. Therefor the Asn52Asn66 doubly-deamidated Bcl-xL could be used as a reliable biomarker for determining the age of platelets.

The invention provides a novel composition containing an artificially synthesized corticotropin analogue and a preparation method and application thereof, the composition comprises N-25 deamidation ACTH (1-39) and further comprises an excipient, and the amino acid sequence of the N-25 deamidation ACTH (1-39) is shown as SEQ ID NO.4. The invention also provides a preparation method and application of the novel composition containing the artificially synthesized corticotropin analogue. The composition disclosed by the invention has a remarkable inhibition effect on rat neonatal spasm symptoms. The composition provided by the invention can reduce the risk of drug immunogenicity, reduce adverse reactions caused by impurities, improve the drug safety, and also can improve the accessibility of the drug to meet the clinical use requirements.

A recombinant adeno-associated virus (rAAV) vector comprising an AAV capsid having a heterogeneous population of vp1 proteins, a heterogeneous population of vp2 protein and a heterogeneous population of vp3 proteins. The capsid contains modified amino acids as compared to the encoded VP1 amino acid sequence, the capsid containing highly deamidated asparagine residues at asparagine-glycine pair, and further comprising multiple other, less deamidated asparagine and optionally glutamine residues.

The problem addressed by the present invention is to provide an enzyme agent for protein deamidation and protein crosslinking containing an enzyme having protein deamidation activity and transglutaminase activity, and the use thereof. The present invention provides an enzyme agent for protein deamidation and protein crosslinking that contains as an active ingredient an enzyme having protein deamidation activity and transglutaminase activity comprising an amino acid sequence having 90% or more sequence identity with the amino acid sequence of SEQ ID NO: 1.

The invention belongs to the technical field of biology, and discloses an artificially modified cephalosporin C acylase mutant and application thereof. The mutant is obtained by carrying out molecular evolution on wild-type enzyme through a DNA shuffling technology, the amino acid sequence of the mutant is shown as SEQ ID NO: 1, and compared with the wild-type enzyme, the mutant has remarkably improved catalytic activity and can efficiently catalyze cephalosporin C deamidation to generate 7-aminocephalosporanic acid. The invention also discloses a nucleic acid molecule for coding the acylase, a recombinant vector and a recombinant host cell containing the nucleic acid molecule, and application of the acylase or the recombinant host in production of 7-ACA by using cephalosporin C as a substrate.

The invention discloses flavor-improved pleurotus eryngii protein and a preparation method thereof, and belongs to the technical field of food processing. The method comprises the following steps: (1) crushing and sieving dried pleurotus eryngii to obtain pleurotus eryngii powder; (2) carrying out degreasing treatment on the pleurotus eryngii powder by using normal hexane so as to remove fat-derived peculiar smell precursor substances; (3) mixing the degreased pleurotus eryngii powder with water, extracting protein under an alkaline condition, and centrifuging to obtain supernate; (4) adjusting the pH value of the supernate to be neutral, adding glutaminase (PG) to carry out an enzymolysis reaction, and modifying a protein structure through a deamidation effect; and (5) inactivating the enzymatic hydrolysate, adjusting to an isoelectric point to precipitate protein, washing, neutralizing and drying to obtain a final product. The method has the remarkable advantages that a deamidase modification process is innovatively integrated into a protein extraction link, and structural modification, flavor improvement and extraction and purification are integrally completed through the synergistic effect of alkali extraction, acid precipitation and enzymolysis. The method effectively overcomes the defects that the inherent peculiar smell of the traditional pleurotus eryngii protein is obvious, and the processing performance (such as water solubility) is poor, the obtained product is pure in flavor, the functional characteristics are remarkably improved, and a new way is opened up for high-value application of the pleurotus eryngii protein in various healthy foods.

A method for preparing deamidated sweet whey protein powder via enzymatic deamidation; a whey protein beverage exhibiting high clarity and is essentially turbidity-free, and its method of preparation using the deamidated sweet whey protein powder.

The invention discloses a protein targeting editor and application thereof, and particularly relates to a protein editor for targeting a target N-glycosylated protein in a receptor biological cell and application of the protein editor. The constructed protein editor comprises three elements: a protein targeting peptide, a linker and a protein editing enzyme. The construction of the deglycosylated protein sequence editor is beneficial to removal of an N-sugar chain of a target protein, and deamidation of an asparagine residue at the site is carried out to convert the asparagine residue into an aspartic acid residue. This deglycosylation editing results in a significant reduction in the stability of the target protein and accelerates the rate of degradation thereof by proteasomes. Therefore, the invention provides a tool capable of performing deglycosylation protein sequence editing on an N-glycosylation modification site of a specific target protein in a cell, which is expected to reveal the functions of N-glycosylation modification and protein sequence editing of asparagine deamidation into aspartic acid in living organisms. And the method has the potential of serving as an alternative method for developing a protein degradation technology.

The invention particularly relates to a quantitative analysis method for deamidation modification in polypeptide. The deamidation of the polypeptide is a common degradation reaction and is also a main reason for reducing the activity of a protein product. According to the present invention, the content of the deamidated peptide in the protein product is quantitatively analyzed based on the peptide map analysis method, and the analysis method can be used for screening the suitable storage condition of the protein active component and optimizing the preparation formula, and has important significance on the analysis of the protein structure and the reduction of the activity loss.

The invention relates to the technical field of freeze-dried preparations, and particularly discloses an esupaglutide alpha nano freeze-dried micelle and a preparation method thereof.Polylactic-co-glycolic acid and sodium phosphatidyl glycerol are selected as micelle carriers, arginine and cysteine provide a branch knot synergistic effect, and the esupaglutide alpha nano freeze-dried micelle is prepared through film forming method homogenization and low-temperature freeze-drying. And freeze-dried nano-micelle powder is obtained. The nano-micelle powder reduces the oxidative decomposition of protein in the freeze-drying process, reduces impurity fragments, various non-fucose glycosylation, alkaline components (succinimide and methionine oxidation products) influencing the product quality, N-terminal deletion, Asn deamidation and generation of Met and Trp residue oxidation impurities, and improves the product quality. Product quality analysis and animal experiments after redissolution show good impurity resistance and degradation risk, the redissolved Esupaglutide alpha nano-micelle system shows better stability and slow release property, the hypoglycemic slow release effect after 72 hours of injection is enhanced, and compared with a conventional non-micelle preparation, the hypoglycemic slow release effect is remarkably improved.