Method for recombinant polypeptide production

FIELD: pharmacology.

SUBSTANCE: method for antibody production, a method for production of a pharmaceutical preparation containing an antibody obtained by this method, a nucleic acid molecule, application of a vector containing such nucleic acid molecule, and application of a cell where the said nucleic acid molecule or vector is artificially introduced in the method for antibody production. This method for antibody production involves culturing a cell that expresses an antibody-enhancing sequence (APES) that has been artificially inserted into a cell and into which an exogenous DNA encoding the desired antibody was introduced to produce the desired antibody, and wherein APES is a nucleic acid molecule containing a nucleotide sequence that can bind to DNA or mRNA of NfkBia gene derived from a human, mouse, rat, or hamster by base pairing, and can suppress expression of the NfkBia gene.

EFFECT: invention allows production of an antibody at a high level with suppression of NfkBia expression.

16 cl, 29 dwg, 8 ex

 



 

Same patents:

FIELD: chemistry.

SUBSTANCE: invention relates to field of biotechnology, namely to internalisation of therapeutic molecules into cell, and can be applied in medicine. Obtained is composition for delivering molecules of nucleic acids into cells, containing at least one peptide with at least 92% identity to GAAEAAARVYDLGLRRLRQRRRLRRERVRA (SEQ ID NO: 2); IREIMEKFGKQPVSLPARRLKLRGRKRRQR (SEQ ID NO: 3); or YLKVVRKHHRVIAGQFFGHHHTDSFRMLYD (SEQ ID NO: 4), bound to one or several molecules of nucleic acids.

EFFECT: invention makes it possible to increase efficiency of delivery of molecules of nucleic acids into mammalian cell due to peptide, capable of internalisation into mammalian cell with efficiency, constituting at least 200% of efficiency of internalisation of peptide TAT, which has amino acid sequence GRKKRRQRRRPPQ (SEQ ID NO: 1).

8 cl, 16 dwg, 1 tbl, 8 ex

FIELD: biotechnology.

SUBSTANCE: method comprises immunoaffinity chromatography using mini-antibodies a-hLF-1 and a-hLF-4, which amino acid sequences are presented as SEQ ID NO:1 and SEQ ID NO:2. The invention may be used to obtain highly purified fraction of human lactoferrin.

EFFECT: invention enables to carry out with high efficiency the separation of proteins of lactoferrin of human and goat consisting in milk, using the single-domain mini-antibodies.

6 dwg, 2 ex

FIELD: chemistry.

SUBSTANCE: claimed inventions deal with a modified protein, nucleic acid, coding such protein, a vector, containing nucleic acid, and a carrier for biotin binding, which such protein is immobilised on. The characterised modified biotin-binding protein is obtained by the introduction of a mutation from one to several amino acid residues into a sequence, represented in SEQ ID NO:2, or an amino acid sequence, identical to the said sequence by 98% or more, and the presence of the biotin-binding activity, where at least one residue, selected from the group, consisting of residues from 1) to 4), presented below, is substituted with the residue of acidic amino acid or residue of neutral amino acid; 1) residue of arginine in position 104 SEQ ID NO: 2; 2) residue of lysine in position 141 SEQ ID NO: 2; 3) residue of lysine in position 26 SEQ ID NO: 2 and 4) residue of lysine in position 73 SEQ ID NO: 2.

EFFECT: claimed inventions make it possible to obtain the biotin-binding protein and can be applied for biotin binding.

14 cl, 6 dwg, 11 tbl, 3 ex

FIELD: medicine.

SUBSTANCE: invention refers to biotechnology and can be used for recombinant production of human tissue factor (hTF). Constructed is a plasmid pHYP-10ETFCS6 having a length of 5,912 b.p. with a physical map presented on Fig. 2, for expression in a bacterium of the genus Escherichia, which is a precursor of the mutant [C245S] hTF containing an inseparable N-terminal leader peptide containing a deca-histidine cluster and an enterokinase identification sequence fused in a frame with a sequence coding the above mutein fused in the frame with the sequence coding the additional inseparable C-terminal peptide containing the deca-histidine cluster. A method for producing the precursor of the mutein[C245S]hTF contains culturing the producing bacterium in a nutrient medium, recovering inclusion bodies, solubilising the precursor protein, performing a metal chelator chromatography in the denaturation environment, re-folding and diafiltration of the protein solution. A method for producing the mature mutein[C245S] hTF involves detecting the N-terminal leader peptide from the above mutein precursor with using enterokinase and recovering the target protein.

EFFECT: invention enables increasing the level of biosynthesis and yield of pro-coagulation active hTF.

9 cl, 5 dwg, 1 tbl, 7 ex

FIELD: chemistry.

SUBSTANCE: group of inventions relates to versions of a method for microbiological synthesis of hybrid protein E7-HSP70. Synthesis of protein E7(6)-HSP70, E7(11)-HSP70, E7(16)-HSP70 or E7(18)-HSP70 is carried out by cultivating the respective yeast strain Saccharomyces cerevisiae VKPM Y-3919, yeast strain Saccharomyces cerevisiae VKPM Y-3853, yeast strain Saccharomyces cerevisiae VKPM Y-4057 or yeast strain Saccharomyces cerevisiae VKPM Y-4058 in suitable conditions in a medium containing saccharose as a carbon source. The cultivation process is carried out in two phases. The first phase is carried out at 25-26°C and initial saccharose concentration of 25-30 g/l. Upon achieving saccharose concentration in the medium of 15 mg/l, the second phase of the cultivation process is carried out, where temperature is lowered and kept not higher than 23°C, pH is kept at 5.7-5.9, and saccharose concentration is maintained via exponential replenishment at 15 mg/l.

EFFECT: group of inventions enables to obtain the end product in amount of not less than 550 mg/l and enables to carry out the cultivation process for not more than 65 hours.

4 cl, 12 dwg, 4 ex

FIELD: biotechnologies.

SUBSTANCE: invention proposes a constructed plasmid for expression in a cell of a Chinese hamster, in the following sequence, which mainly contains the following elements: pUC plasmid replication beginning region; an open reading frame (ORF) of beta-lactamase providing immunity to ampicillin; procaryotic gene promoter bla; a section of terminal repetition of Epstein-Barr virus of a human being; a functional gene promoter of elongation factor 1 alpha of the Chinese hamster, 5' non-translated region of this gene and a non-transcribed region flanking this gene, coding Kozak sequence for cap-dependent initiation of translation; ORF of the gene of subunit 1 of complex of 2,3-epoxyreductase of vitamin K (VKORC1) of the Chinese hamster with stop codon; a functional terminator and a signal of polyadenilation of the gene of elongation factor 1 alpha of the Chinese hamster, 3' non-translated region of this gene and a non-transcribed region flanking this gene; a promoter of early genes of virus SV40; gene of immunity to a selective agent; and a polyadenilation signal and terminator of virus SV40. Cells of the Chinese hamster - producer of protein with Gla-domain are transformed by the obtained plasmid, which are used in a method of recombinant obtainment of proteins with Gla-domain.

EFFECT: invention allows increasing productivity of the above cells owing to increasing activity of native VKORC1.

12 cl, 12 dwg, 2 tbl, 8 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to the field of biotechnology, namely to novel analogues of insulin and can be used in medicine. The said insulin analogue is characterised by one of the following structures: Arg(A0), His(A8), Gly(A21), Arg(B31), Arg(B32)-NH2-insulin; His(A8), Gly(A21), Arg(B31), Arg(B32)-NH2-insulin; Arg(A0), Glu(A15), His(A8), Gly(A21), Arg(B31), Arg(B32)-NH2-insulin. The invention also relates to a method of obtaining the said analogue, a pharmaceutical composition and a medication, including its application for treatment of diabetes mellitus.

EFFECT: invention makes it possible to obtain the insulin analogue, characterized by the delayed release and steady and longer action, for instance, in comparison with human insulin or glargine insulin.

18 cl, 2 dwg, 24 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to biotechnology, namely to leukolectins, and can be used in medicine. What is prepared is the polypeptide leukolectin characterised by SEQ ID NO:1-8. The recombinant preparation is ensured by using a nucleic acid coding it and integrated into an expression vector which is used to transform a host cell. Testing absence-presence or determining an amount of the polypeptide leukolectin are ensured by using an antibody or an antigen-binding fragment of a variable region of the above antibody which is specifically bound to the polypeptide leukolectin. The polypeptide leukolectin or the nucleic acid coding it are used as ingredients of a pharmaceutical composition in therapy of pathological disorders of skin and mucous membranes.

EFFECT: invention enables treating or preventing autoimmune disorders of skin, inflammatory diseases of skin or mucous membrane, or injured skin in an animal effectively.

16 cl, 19 dwg, 3 tbl, 12 ex

FIELD: medicine.

SUBSTANCE: invention refers to biotechnology, more specifically to modified von Willebrand factor (VWF), and can be used in medicine. A recombinant method is used to preparing modified VWF fused in C-terminal of its primary translation product with N-terminal of albumin by the linker SSGGSGGSGGSGGSGGSGGSGGSGGSGGSGS. The prepared modified VWF is used as a part of the pharmaceutical composition for treating or preventing coagulation failure.

EFFECT: invention enables preparing the modified VWF which maintains its ability to N-terminal dimerisation and C-terminal multimerisation with a prolonged half-period of functional blood plasma occurrence as compared to the half-period of functional VWF occurrence.

17 cl, 5 dwg, 4 tbl, 11 ex

FIELD: chemistry.

SUBSTANCE: group of inventions relates to biochemistry. Disclosed is L-fucose α1→6 specific lectin, which is extracted from a basidiomycete or an ascomycete or an ascomycete, characterised by peak molecular weight of about 4500 m/z, determined via MALDI-TOF mass spectrometry analysis. The novel L-fucose α1→6 specific lectin has high affinity for a L-fucose α1→6 sugar chain, represented by an association constant of 1.0×104 M-1 or higher (at 25°C), and has an association constant of 1.0×103 M-1 or lower (at 25°C) with high-mannose sugar chains and/or glucolipids which do not contain an L-fucose α1→6 sugar chain. In one version, the disclosed L-fucose α1→6 specific lectin is a protein or a peptide which consists of an amino acid sequence selected from SEQ ID NO:2-6. The L-fucose α1→6 specific lectin is used for specific detection of a L-fucose α1→6 sugar chain and effective purification of the L-fucose α1→6 sugar chain or a sugar chain which does not contain L-fucose α1→6.

EFFECT: obtaining L-fucose α1→6 specific lectin.

16 cl, 38 dwg, 8 tbl, 4 ex

FIELD: medicine.

SUBSTANCE: invention relates to field of biochemistry, in particular to antibody or its antigen-binding fragment, which is specifically bind with human TNFα. Also disclosed are: separated molecule of nucleic acid, which codes said antibody, vector of expression, which contains said molecule of nucleic acid, and host cell, which contains said vector, for expression of said antibody. Disclosed are pharmaceutical composition for treatment of TNFα-mediated disease, which contains therapeutically effective quantity of said antibody, and method of treating TNFα-mediated disease with application of claimed composition.

EFFECT: invention makes it possible to effectively treat TNFα-mediated diseases.

16 cl, 9 dwg, 2 tbl, 2 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to the field of immunology. Claimed is the application of a monoclonal antibody against a protein of a human Fas ligand (CD95L, or Apo1L, or FasL) or its antigen-binding fragment for manufacturing a medication for the prevention and/or treatment of skin diseases, associated with the acantholysis of keratinocytes, in particular for the prevention and/or treatment of pemphigus, where the antibody contains amino acid sequences CDR of the antibody NOK-2, F919-7-3, F918-9-4 or F919-9-18 or is produced by the hybridoma ATCC PTA-5045, ATCC PTA-5533, ATCC PTA-5534 or ATCC PTA-5535.

EFFECT: application of the antibodies by the invention or their antigen-binding fragments provides the effective blocking of mechanisms, resulting in pemphigus lesions.

7 cl, 14 dwg

FIELD: chemistry.

SUBSTANCE: present invention relates to biotechnology and provides a α1,6-glucan-containing compound of Helicobacter pylori. The present invention also discloses a conjugate for inducing immune response against H.pylori, which contains said compound conjugated with a carrier protein. The present invention also discloses an immunogenic composition, use of said composition and a method of inducing immune response against H.pylori using said composition. The present invention also discloses immune serum for neutralising H.pylori in mammals, which is obtained by immunising said mammal with an immunogenic composition containing said immunogenic composition. The present invention discloses an antibody which recognises said α1,6-glucan-containing compound of H.pylori, use of said antibody and a method of inducing complement-mediated bacteriolysis of H.pylori strains which express α1,6-glucan using said antibody.

EFFECT: invention improves the effectiveness of immunogenic compositions against Hpylori.

27 cl, 8 dwg, 21 tbl, 11 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to medicine, namely to therapy and pharmaceutics, and deals with an influence on the functional activity of a cannabinoid receptor. For this purpose an activated-potentiated form of antibodies to the human cannabinoid receptor is introduced into an organism with the application of an activated-potentiated form of antibodies to an intact molecule or a polypeptide fragment of the human cannabinoid receptor of type I.

EFFECT: method ensures the effective correction of endocannabinoid system impairments with the absence of side effects.

4 cl, 1 tbl, 1 ex

FIELD: medicine.

SUBSTANCE: invention relates to medicine, namely to cardiovascular surgery and cardiology, and deals with complex correction of immunoinflammatory responses of cardiovascular bed. For this purpose, in case of presence of high level of circulating immune complexes and/or complement in patient, three sessions of plasmapheresis in accordance with conventional methods, and in case of low level of IgG and reduced phagocytic activity of neutrophils and monocytes, as well as in case of confirmed autoimmune process, a course of intravenous infusions of human polyvalent immunoglobulin is carried out in accordance with conventional schemes.

EFFECT: method provides reduction of both hypo- and hyperactive disorders in immune system of patients and resulting increase of efficiency of treatment of cardiovascular system diseases.

3 ex

FIELD: medicine.

SUBSTANCE: claimed invention relates to the field of biotechnology. Claimed are versions of a humanised anti-CD79b antibody, each of which is characterised by the presence of a light and heavy chain and a set of 6 CDR with a determined amino acid sequence. An epitope of the antibody from 11 amino acids is determined by the Biacore method. Disclosed are: an immunoconjugate of the antibody with a medication or means for inhibiting cell growth, where the antibody is bound with means covalently, and versions of the composition, based on an effective quantity of the immunoconjugate or the antibody, used for inhibiting B-cell proliferation; as well as a method of determining CD79b in a sample with the application of the antibody. Described are: an antibody-coding polynucleotide, as well as an expression vector and an isolated cell, containing the vector for obtaining the antibody. Disclosed are versions of applying the antibody or immunoconjugate for obtaining the medication for inhibiting the growth of CD79b-expressing cells for the treatment of an individual, affected with cancer, for the treatment of proliferative disease or for inhibiting B-cell proliferation.

EFFECT: invention provides novel antibodies, which can find further application in the therapy of proliferative CD79b-associated diseases.

91 cl, 8 tbl, 9 ex, 20 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention refers to biotechnology, more specifically to AXL signalling pathway inhibitors, and can be used in medicine. What is produced is a soluble AXL polypeptide version free from AXL transmembrane domain, which contains at least one amino acid modification in position No. n, wherein n is specified in 32, 72, 87, 92 or 127 or a combination thereof, wherein n+7 is described by numbering SEQ ID NO: 1 that are wild-type AXL sequences, wherein the above modification increases a AXL polypeptide binding affinity to protein 6 specifically inhibiting the growth (GAS6), which is twice as strong as the wild-type AXL polypeptide affinity. The polypeptide can be fused with Fc fragment and used in a method of treating, reducing or preventing tumour dissemination and invasion in a mammalian patient.

EFFECT: invention enables inhibiting the AXL/GAS6 signalling pathways effectively.

8 cl, 15 dwg, 6 tbl, 3 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: claimed invention relates to field of immunology. Claimed is application of monoclonal antibody against human Fas ligand protein (CD95L, or Apo1L, or FasL) or its antigen-binding fragment for manufacturing medication for prevention and/or treatment of skin diseases, associated with acantholysis of keratinocytes, in particular for prevention and/or treatment of pemphigus, where antibody contains amino acid sequences CDR of 3E1 antibody or is produced by hybridoma ATCC PTA-4017.

EFFECT: application of antibody in accordance with invention or its antigen-binding fragment provides more effective prevention of desmoglein (dsg3) cleavage in comparison with application of other antibodies.

11 cl, 16 dwg

FIELD: medicine, pharmaceutics.

SUBSTANCE: invention relates to the field of biochemistry, in particular to a humanised antibody to a tumour necrosis factor-α or its antigen-binding fragment Fab. Also disclosed are: gene, coding the protein of the antibody or Fab, genetic material, expressing the said antibody or Fab-fragment. Disclosed are: application of the antibody of Fab for obtaining a medication for the prevention or treatment of diseases, associated with the human tumour necrosis factor-α and a pharmaceutical composition for the treatment of diseases, associated with the human tumour necrosis factor-α, containing an effective quantity of the said antibody or Fab.

EFFECT: invention possesses a reduced immune response, in comparison with the pharmaceutical antibody Remicade, which makes it possible to treat diseases, associated with the human tumour necrosis factor-α, in an effective way.

23 cl, 5 dwg, 9 tbl, 7 ex

FIELD: medicine, pharmaceutics.

SUBSTANCE: presented invention refers to immunology. What is presented is a monoclonal anti-IFNAR1 antibodies with L234F, L235E and P331S Fc mutations of human IgG1 possessing a lower affinity to Fcgamma RI, Fcgamma RIIIA and c1q receptors as compared to a non-modified antibody. There are described the recovered nucleic acid providing expression of the above antibody containing a nucleotide sequence coding the antibody, and a pharmaceutical composition based on the above antibody.

EFFECT: using the invention provides the antibody possessing the lower affinity to Fcgamma RI, Fcgamma RIIIA and c1q receptors that provides reducing the undesired effector functions in treating chronic inflammation and autoimmune conditions.

9 cl, 34 dwg, 7 tbl, 36 ex

FIELD: medicine.

SUBSTANCE: invention relates to field of biotechnology and deals with method of separating mixture of DNA and proteins from epimastigote forms of strain TPAP/MX/2002/Albarrada of Trypanosoma cruzi culture. Characterised method includes obtaining biomass of said strain culture, with further washing of obtained biomass, its concentration to content of not less than 6×108 cells in 1 ml, introduction of synthetic preparation of human defensin α-1 with molecular weight 3.4 kDa into obtained concentrated biomass in amount 20 mcg/ml to its final concentration 3.97 mcM and exposure until pores are formed in lipid membranes. After that obtained suspension is shaken by shaker with further exposure at room temperature for not less than 24 hours, with the following centrifuging obtained medium with separation of mixture of DNA and proteins in liquid phase.

EFFECT: invention makes it possible to obtain product with reduced content of protein components and can be used in further obtaining of preparations.

2 cl, 2 ex

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