Single-chain protein modulator of luteinizing hormone, follicle-stimulating, thyrostimulin and human chorionic gonadotropin (options), a molecule of nucleic acid (options), vector (options), line transformed cells cho (options), the method of producing single-chain protein (options)

 

The invention relates to biotechnology and endocrinology and can be used to produce single-chain forms of the glycoprotein hormones. Single-chain protein is covalently linkedandorandorandsubunit of luteinizing hormone (LH), follicle-stimulating hormone (FSH), a hormone, thyroid-stimulating (TSH) and chorionic gonadotropin (CG). Amino acid sequences of subunits linked via a linker or without him. As the linker use the full link KTP, consisting of 112-118 to 145 amino acid subunit of human HCG. Single-chain protein is produced by culturing the cell line of Cho, transformed by a vector containing a nucleic acid encoding a single-chain protein. The invention allows to obtain a stable form of hormones. 15 C. and 9 C.p. f-crystals, 4 Il.

Description text in facsimile form (see the graphical part of) The

Claims

1. Single-chain protein, modulator (agonist or antagonist) of hormoneman, thyroid-stimulating (TSH) and human chorionic gonadotropin (HCG), formula-(linker)n-or-(linker)n-whereissubunit of LH, FSH, TSH or HCG or her option; n = 0 or 1;represents the amino acid sequence ofsubunit common to LH, FSH, TSH or HCG or her option; and named the single-chain protein is optionally glycosylated.

2. Single-chain protein under item 1, characterized in that the said linker is a peptide and subunitandconnected on the principle of "head to tail".

3. Single-chain protein in p. 2, characterized in that the linker includes a full link KTP consisting of amino acid residues 112-118 to 145subunit of human chorionic gonadotropin (HCG) or its variants.

4. Single-chain protein in p. 2, characterized in thatsubunit, orsubunit, or both of these subunits modificatory specified, the linker includes a full link KTP, a part of him or his option.

5. The nucleic acid molecule encoding a single-chain protein, a hormone modulator selected from the group consisting of LH, FSH, TSH and HCG, which is a DNA or RNA comprising the nucleotide sequence corresponding to the amino acid sequence of single-chain protein under item 1.

6. The vector for the production of single-chain protein modulator of LH, FSH, TSH or XT comprising the nucleotide sequence under item 5, functionally associated with a regulatory sequence that provides its expression in compatible cell host.

7. The cell line Cho transformed by the vector according to p. 6, which is the producer of single-chain protein under item 1.

8. Method of producing single-chain protein modulator (agonist or antagonist) of LH, FSH, TSH, or HCG, comprising culturing the cell line transformed with the vector under item 6 in the conditions for producing the specified single-chain protein, and need not be further released from the cell culture.

9. Single-chain protein hormone antagonist selected from the group consisting of luteinizing hormone (LH), follicle-stimulating hormone (FSH), a hormone, with NCAR)n-whereissubunit of LH, FSH, TSH or HCG or its variant, n = 0 or 1;
moreover, the specified single-chain protein is optionally glycosylated.

10. Single-chain protein under item 9, characterized in that the said linker is a peptide, andsubunits are connected by the principle of "head to tail".

11. Single-chain protein by p. 10, characterized in that n is 1, and the specified linker includes full KTP link consisting of amino acid residues 112-118 to 145 In the subunit of human chorionic gonadotropin (HCG) or its variants.

12. Single-chain protein by p. 10, characterized in that one or both-subunit is modified by inserting the full QFT link part of the link or its variant in a non-critical part, and/or in which the specified linker includes full KTP link, its part or its variant.

13. The nucleic acid molecule encoding a single-chain protein, a hormone modulator selected from the group consisting of LH, FSH, TSH and HCG, which is a DNA or RNA comprising the nucleotide sequence corresponding to the amino acids of the modulator (agonist or antagonist) LH, FSH, TSH, or HCG, which includes the nucleotide sequence for p. 13, functionally associated with a regulatory sequence that provides its expression in compatible cell host.

15. The cell line Cho transformed by the vector according to p. 14, which is the producer of single-chain protein under item 9.

16. Method of producing single-chain protein modulator (agonist or antagonist) of LH, FSH, TSH, or HCG, comprising culturing the cell line under item 15 in conditions that ensure the production of a specified protein, and optionally further its secretion from the cell culture.

17. Single-chain protein modulator (antagonist or agonist) hormone selected from the group consisting of luteinizing hormone (LH), follicle-stimulating hormone (FSH), a hormone, thyroid-stimulating (TSH) and chorionic gonadotropin (CG) formula:
-(linker)n-,
in whichrepresents the amino acid sequence ofsubunit common to LH, FSH, TSH and HCG or its variant;
n = 0 or 1,
moreover, the specified single-chain protein is optionally glycosylated.

18. About the g> subunits are connected by the principle of "head to tail".

19. Single-chain protein under item 18, characterized in that the linker includes full KTP link consisting of amino acid residues 112-118 to 145subunit of human chorionic gonadotropin (HCG) or its variants.

20. Single-chain protein under item 18, characterized in that one or boththe a subunit is modified by inserting the full QFT link part of the link or its variant in a non-critical part, and/or in which the specified linker includes full KTP link, part of the KTP link or option.

21. The nucleic acid molecule encoding a single-chain protein, a hormone modulator selected from the group consisting of LH, FSH, TSH and HCG, which is a DNA or RNA comprising the nucleotide sequence corresponding to the amino acid sequence of single-chain protein under item 17.

22. The vector for the production of single-chain protein modulator (agonist or antagonist) of LH, FSH, TSH, or HCG, comprising the nucleotide sequence according to p. 21, functionally associated with a regulatory sequence that provides its expression in compatible to echnolo protein in p. 17.

24. Method of producing single-chain protein modulator (agonist or antagonist) of LH, FSH, TSH, or HCG, comprising culturing the cell line, transformirovannykh vector for p. 22 under conditions that ensure the production of the specified single-chain protein and optional subsequent isolation from cell culture.

 

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