| 摘要 |
FIELD: biochemistry, chemistry of proteins. SUBSTANCE: invention relates to protein characterizing the following properties: (a) molecular mass about 60 kDa under reducing conditions, about 60 kDa and 120 kDa under nonreducing conditions at electrophoresis in polyacrylamide gel in the presence of sodium dodecyl sulfate (SDS-PAGE); (b) high affinity to cation-exchange column and column with heparin; (c) biological activity of inhibition of differentiation and/or maturation of osteoclasts being this activity is decreased in heating at 70 C for 10 min or at 56 C for 30 min and indicated activity is lost in heating at 90 C for 10 min; (d) internal amino acid sequences presented in SEQ ID NO : 1, 2 and 3; and (e) optionally having N-terminal amino acid sequences presented in SEQ ID NO : 7. Invention relates to method of such proteins by culturing human fibroblasts, purification of protein by combination of column ion-exchange chromatography, affinity column chromatography and column chromatography with reversed phase. Invention relates also to proteins with amino acid sequence presented in SEQ ID NO : 4, cDNA encoding amino acid presented in SEQ ID NO : 4; cDNA with nucleotide sequence presented in SEQ ID NO : 6 encoding protein OCIF with signal peptide having amino acid presented in SEQ ID NO : 5; protein expressed from cDNA encoding amino acid sequence presented in SEQ ID NO : 4; protein with biological activity of inhibition of differentiation and/or maturation of osteoclasts that is obtained as amino acid sequence expressed by cDNA having at least 80% of identity with amino acid sequence presented in SEQ ID NO : 4; method of preparing protein inhibiting development of osteoclasts by methods of genetic engineering and using cDNA encoding amino acid sequence presented in SEQ ID NO : 4: (a) molecular mass about 60 kDa under reducing conditions and about 60 kDa and 120 kDa under nonreducing conditions at electrophoresis in polyacrylamide gel in the presence of sodium dodecyl sulfate (SDS-PAGE); (b) high affinity to cation-exchange column and to column with heparin; (c) inhibitory activity of differentiation and/or maturation of osteoclasts that is decreased at heating at 70 C for 10 min or at 56 C for 30 min being indicated activity is lost at heating at 90 C for 10 min; (d) internal amino acid sequence presented in SEQ ID NO : 1-3; cDNA encoding protein OCIF or its variants; genome DNA with nucleotide sequence presented in SEQ N : 104 or 105; polyclonal antibody with specific affinity raised to OCIF; monoclonal antibody with specificity to OCIF obtained by hybridoma technology using OCIF as antigen and characterizing the following properties: molecular mass 150 000 Da; method of determination of concentration of protein OCIF. EFFECT: improved method of proteins preparing. 19 cl, 16 tbl, 15 dwg, 26 ex
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