2185 results about "Trypsin" patented technology

Recombinant protein complexes having human Factor VIII:C activity are expressed in a eukaryotic host cell by transforming the host cell with first and second expression cassettes encoding a first polypeptide substantially homologous to human Factor VIII:C A domain and a second polypeptide substantially homologous to human Factor VIII:C C domain, respectively. In the present invention, the first polypeptide may be extended having at its C-terminal a human Factor VIII:C B domain N-terminal peptide, a polypeptide spacer of 3-40 amino acids, and a human Factor VIII:C B domain C-terminal peptide. Expression of the second polypeptide is improved by employing an .alpha..sub.1 -antitrypsin signal sequence.

The present invention concerns compounds, compositions, and methods for the study, diagnosis, and treatment of diseases and conditions associated with alpha-1 antitrypsin (AAT) allelic variants that respond to the modulation of gene expression and/or activity. The present invention also concerns compounds, compositions, and methods relating to diseases and conditions associated with alpha-1 antitrypsin (AAT) allelic variants that respond to the modulation of expression and/or activity of genes involved in alpha-1 antitrypsin (AAT) gene expression pathways or other cellular processes that mediate the maintenance or development of alpha-1 antitrypsin (AAT) diseases and conditions such as liver disease, lung disease, and any other diseases or conditions that are related to or will respond to the levels of an alpha-1 antitrypsin (AAT) variant protein in a cell or tissue, alone or in combination with other therapies. Specifically, the invention relates to small nucleic acid molecules, such as short interfering nucleic acid (siNA), short interfering RNA (siRNA), double-stranded RNA (dsRNA), micro-RNA (mRNA), and short hairpin RNA (shRNA) molecules capable of mediating RNA interference (RNAi) against the expression disease related genes or alleles having alpha-1 antitrypsin (AAT) sequences.

The present invention provides methods and compositions for generating novel nucleic acid molecules through targeted spliceosomal mediated RNA trans-splicing. The compositions of the invention include pre-trans-splicing molecules (PTMs) designed to interact with a SERPINA1 target precursor messenger RNA molecule (target pre-mRNA) and mediate a trans-splicing reaction resulting in the generation of a novel chimeric RNA molecule (chimeric RNA). In particular, the PTMs of the present invention include those genetically engineered to interact with SERPINA1 target pre-mRNA so as to result in correction of SERPINA1 genetic defects responsible for AAT deficiency. The PTMs of the invention may also comprise sequences that are processed out of the PTM to yield duplex siRNA molecules directed specifically to mutant SERPIN A1 mRNAs. Such duplexed siRNAs are designed to reduce the accumulation of toxic AAT protein in liver cells. The methods and compositions of the present invention can be used in gene therapy for correction of SERPINA1 disorders such as AAT deficiency.

Compounds of formula (I) are useful as inhibitors of trypsin like serine protease enzymes such as thrombin, factor VIIa, factor Xa, TF/FVIIa, and trypsin. These compounds could be useful to treat and/or prevent clotting disorders, and as anticoagulating agents.

The present invention demonstrated the potential use of Lactobacillus johnsonii D115 as a probiotic, as a prophylactic agent or as a surface treatment of materials against human and animal pathogens such as Brachyspira pilosicoli, Brachyspira hyodysenteriae, Shigella sonnei, Vibrio cholera, Vibrio parahaemolyticus, Campylobacter jejuni, Streptococcus pneumoniae, Enterococcus faecalis, Enterococcus faecium, Clostridium perfringens, Yersinia enterocolitica, Escherichia coli, Klebbsiella pneumoniae, Staphylococcus aureus, Salmonella spp., Bacillus cereus, Aspergillus niger and Fusarium chlamydosporum. The proteineous antimicrobial compound was partially characterized and found to be heat tolerant up to 121° C. for 15 min, and acid tolerant up to pH1 for 30 min at 40° C. The compound is also stable to enzymatic digestion, being able to retain more than 60% antimicrobial activity when treated with pepsin and trypsin.

A protein expression vector containing (a) a nucleotide sequence encoding an IgG(κ) or a trypsin secretory signal peptide, (b) a nucleotide sequence encoding a polyhistidine amino acid sequence, (c) a nucleotide sequence encoding an amino acid sequence comprising amino acid residues 36–40 of SEQ ID NO:19 (Asp-Asp-Asp-Asp-Lys), which is cleavable by an enterokinase, and (d) a cloning site into which a nucleotide sequence encoding a target protein can be inserted, wherein (a), (b), (c) and (d) are assembled within the vector in the order recited.

Embodiments herein illustrate methods and compositions for treating medical disorders. In certain embodiments, compositions and methods relate to reducing, inhibiting or treating graft rejection, transplant rejection or diabetes in a subject. Other embodiments herein relate to compounds including naturally occurring and synthetic mutant compositions of alpha-1 antitrypsin, wherein the alpha-1 antitrypsin has no significant serine protease inhibitor activity.

A novel method of treating and preventing diseases is provided. In particular, compositions and methods of blocking diseases associated with aberrant levels of nitric oxide and facilitated by a serine proteolytic (SP) activity are disclosed, which consist of administering to a subject a therapeutically effective amount of a compound having a serine protease inhibitory activity. Among effective compounds are alpha1-antitrypsin and synthetic drugs mimicking some or all of the actions of alpha1-antitrypsin.

The invention discloses a comprehensive extraction process of holothurian polypeptides and polysaccharides. The invention is characterized in that: fresh holothurians are taken as raw materials, gelatinized and hydrolyzed under the effect of composite trypsinases; precipitation and supernatant are respectively collected through centrifugation of the holothurians; ethanol is added into the supernatant for centrifugation; supernatant obtained is mixture of the holothurian polypeptides; centrifugal precipitations obtained for two times are synthesized, and A.S1398 purified neutral proteases are added for hydrolysis and centrifugation; ethanol is added into the supernatant for centrifugation; supernatant obtained for two times is synthesized into holothurian polypeptide mixture, and precipitation is crude polysaccharides; the holothurian polypeptide mixture is dissolved in water, and ultrafiltration membranes with different apertures are used for separation and purification of subsections of different molecular weights; holothurian polypeptide powders of different molecular weight sections are obtained after debitterizing, decolorization and freeze drying; crude polysaccharides are mixed into 5 percent aqueous solution; after centrifugation, decolorization and repeated centrifugation, precipitation is colleted and washed, and aqueous solution is mixed and dialyzed; ethanol is added for repeated precipitation for at least three times, and purified holothurian polysaccharides are obtained. The invention simultaneously prepares the holothurian polypeptides with different molecular weight sections and the holothurian polysaccharides, and has the advantages of high yield, high purity, high safety and mild reaction conditions.

The present invention comprises a method of protecting organs or tissue susceptible to reperfusion-induced dysfunction after ischemia. The method comprises parenterally administering to a patient a therapeutical composition containing natural alpha-1 acid glycoprotein, natural alpha-1 antitrypsin or a mixture thereof. Alternatively, organ or tissue transplants can be contacted with natural alpha-1 acid glycoprotein, natural alpha-antitrypsin or mixtures by perfusing or flushing them with a solution containing natural alpha-1 acid glycoprotein, natural alpha-1 antitrypsin or mixtures thereof in a concentration of 0.1 to 5 g/l.

The invention relates to the technical field of biology, and discloses a moringa extract as well as a preparation method of the moringa extract and a moringa feed additive. According to the preparation method, water is added into moringa leaves, the pH value and the temperature are reduced, then, biological enzymes are adopted for enzymolysis, next, heating digestion is adopted to obtain water extraction liquid, ethanol precipitation is carried out after water extraction liquid concentration, obtained precipitates are the moringa extract, the biological enzymes are protease, cellulose and pectinase, and the protease is papain, pepsase, trypsin or serrapeptass. The enzymolysis effects of the specific protease, cellulose and pectinase is utilized for hydrolyzing large blocks of complicated-molecular-structure ingredients of the moringa leaves per se into smaller water-soluble high-bioavailability small molecular active substances, and then, high-yield and high-quality moringa extracts are obtained through a water extraction and ethanol precipitation method.

The invention discloses a method and process for extracting animal micro-molecule polypeptides and amino acids by utilizing a complex enzyme. The method comprises the following steps: (1) crushing animal tissues so that the animal tissues can pass through a sieve of 80-200 meshes, and adding water to prepare a suspension with the concentration of 8-30 percent; (2) raising the temperature to be 35-40 DEG C, regulating the pH value to be 8.0-9.0 by using sodium carbonate, adding a complex enzyme I accounting for 0.5-2.0 percent of the animal tissues, performing ultrasonic enzymolysis at 14-16 kHz for 1-2 hours so that the proteins between cells are subjected to full enzymolysis, regulating the pH value to be 6.5-7.5, adding a complex enzyme II accounting for 0.5-3.0 percent of the animal tissues, fully and uniformly stirring, controlling the temperature of the water bath to be 37-45 DEG C, and performing ultrasonic enzymolysis at 15-17 kHz for 1-6 hours; (3) raising the temperature of the enzymolysis solution to be 90-100 DEG C to inactivate the enzyme for 20-30 minutes after the enzymolysis is finished; (4) filtering the solution subjected to enzymolysis through a 0.22mu m microfiltration membrane; and (5) concentrating the filtrate to the relative density of 1:1.0-1:1.2 to prepare the concentrated solution, wherein the complex enzyme II consists of Alcalase, Flavourzyme, Protamex, neutral protease and papain. According to the method, the enzymolysis efficiency can be improved, and the production cost can be reduced.

The invention discloses a method for extracting polysaccharide from a cordyceps militaris medium. The cordyceps militaris is crushed and mixed with water, is extracted in an enzyme liquor and an alkali liquor under 30 DEG C to 80 DEG C, and an obtained solution is centrifugalized. An insoluble substance is extracted repeatedly, and supernatants are combined. Amylase enzymolysis is performed on the obtained product under 50 DEG C to 75 DEG C. After centrifugation, a supernatant is condensed to be with a sugar content of 1.5 percent to 3.0 percent. The supernatant is added with ethanol and alcohol precipitation is performed on the obtained product under 0 DEG C to 4 DEG C, a product is obtained by drying the obtained product after centrifugation. After deproteinization and dialysis, crude cordyceps polysaccharide is obtained. A single enzyme method, a double enzyme method, a compound enzyme method and a supersonic wave enzyme method can be adopted, and protease, trypsin, etc. can be adopted. The method optimizes the technology route for extracting the cordyceps polysaccharide, enhances an obtaining rate and an extraction efficiency of the cordyceps polysaccharide. Extraction, separation and purification of the cordyceps polysaccharide are carried out effectively. The method provides a solution approach for the utilization of agricultural wastes, and provides a technology base for further exploitation of the cordyceps polysaccharide.

The invention discloses a giant salamander active peptide and an application. The giant salamander active peptide is prepared by the following steps: conducting enzymatic hydrolysis on giant salamander meat which serves as a raw material by virtue of a complex enzyme of marine alkaline protease and papain; separating an enzymatic hydrolysate by virtue of a trypsin immobilized ultra-filtration membrane separator; and then conducting separation and purification by virtue of Sephadex LH-20 molecular sieve chromatography and high performance liquid chromatography, so that the giant salamander active peptide is obtained. The giant salamander active peptide is capable of effectively scavenging free radicals and boosting body immunity, and meanwhile, the active peptide can also promote proliferation of skin fibroblasts; therefore, the giant salamander active peptide has a broad application prospect in the fields of food, medicines and cosmetics.

The invention relates to a quantitative detection method for thermal-denaturation and non-denaturation bovine alpha-lactalbumin in milk and milk products by applying an enzymolysis-liquid chromatography and mass spectrometry combination technology. The quantitative detection method comprises the steps as follows: taking a certain amount of milk or milk samples, dissolving and diluting the milk or milk samples in water to obtain solution with total protein content being about 1mg/mL; after volume metering, correctly sucking 500 mu L, adding an internal standard substance, reacting disulfide bond with dithiothreitol (DTT), alkylating to protect sulfydryl produced in reaction by iodoacetamide (IAA), and then conducting constant-temperature and constant-time enzymolysis with trypsin; and separating enzymolysis products by reversed phase liquid chromatography, conducting detection with a mass spectrum multiple reaction monitoring (MRM) manner, and calculating the result by an internal standard method. The quantitation limit of the method is 0.001g/100g; when adding amount is 0.2, 1.7 and 5.0g/100g, the recovery rate is 98.9-110.8% (n is equal to 6) and repeatability: RSD (Relative Standard Deviation) is smaller than 7.6%; and the quantitative detection method can be applicable to the quantitative detection of samples with different contents of bovine alpha-lactalbumin.

The present invention relates to a method for isolating and culturing adult stem cells derived from human amniotic membrane in high yield, and more particularly to a method for obtaining a large amount of adult stem cells, the method comprising obtaining amniotic epithelial cells from human amniotic tissue in high yield by treatment with dithiothreitol (DTT) and a low concentration of trypsin and culturing the amniotic epithelial cells in a medium containing a Rho-associated kinase inhibitor. The human amniotic epithelial cell-derived stem cells are easily extracted compared to existing therapeutic stem cells such as umbilical cord blood stem cells and bone marrow stem cells, the yield and proliferation thereof are significantly increased by DTT treatment, the addition of the ROCK inhibitor or the replacement of medium. Thus, the method can be used to efficiently prepare adult stem cells.

The invention relates to a method for producing a soybean peptide. In the method, fermented soybean meal is taken as a raw material and then animal-derived protease is utilized to produce the soybean peptide. The method comprises the following steps of: 1) enzymolyzing liquid, namely mixing the fermented soybean meal and water in a weight ratio of 1:(4-8), adding trypsinase, and preserving heat and enzymolyzing; 2) killing enzyme at high temperature; 3) filtering, namely filtering and separating enzymatic hydrolysate to obtain crude soybean protein; 4) decolorizing and deodorizing; 5) refining; and 6) spray-drying, and sterilizing to obtain a finished product, wherein the decolorization and deodorization are realized through filtration by a pretreated macroporous resin. The method has theadvantages that: 1) the fermented soybean meal is taken as the raw material, so that a protease inhibitor in the raw material is eliminated; 2) the prehydrolysis of soybean protein in the fermented soybean meal can be completed; and 3) the amount of the additional enzyme is reduced greatly, so that the cost is effectively reduced and the higher hydrolysis rate is obtained.