RECOMBINANT PLASMID DNA pER-TA1 GyrA-AcSer CODING SERINE ACETYLTRANSFERASE CAPABLE OF in vivo ACETYLATION OF N-TERMINAL SERINE DEACETYL-THYMOSIN α1 AND HYBRID PROTEIN CAPABLE OF AUTOCATALYTIC BREAKDOWN TO FORM HUMAN THYMOSIN α1, STRAIN OF Eschrichia coli C3030/pER-TA1GyrA-AcSer PRODUCER OF SAID PROTEINS AND METHOD OF PRODUCING GENETICALLY ENGINEERED HUMAN THYMOSIN

摘要

FIELD: biotechnology.SUBSTANCE: invention relates to genetic and protein engineering and represents recombinant plasmid DNA pER-TA1GyrA-AcSer, intended for simultaneous biosynthesis of two proteins - hybrid polypeptide containing amino acid sequence of human thymosin α1 and intein, and enzyme E.coli - serine acetyl transferase, having molecular weight 5.04 MDa. Plasmid consists of BsaBI/NdeI DNA fragment of plasmid pTWTN-1, containing T7 promoter sequence and operator lacO; Ndei/PstI DNA fragment containing a sequence of hybrid protein, including adapted to said sites sequence of recombinant human thymosin α1, intein sequence Moss GyrA and sequence of chitin binding domain; Psti/NcoI DNA fragment coding reduced Shine-Dalgarno sequence and non-standard as codon-start amino acid valine; NcoI/XhoI DNA fragment containing a sequence of serine N-acetyl transferase; DNA fragment containing genetic marker of β-lactamase gene, determining resistance of transformed by plasmid pER-TA1GyrA-AcSer cells of E.coli to penicillin antibiotics; DNA fragment containing a sequence coding replication start point; DNA fragment coding sequence of lactose repressor LacI. Invention also relates to a strain Escherichia coli C3030/pER-TA1GyrA-AcSer, producing hybrid polypeptide, containing amino acid sequence of human thymosin α1 and intein, as well as serine acetyl transferase obtained by transformation of cells of Escherichia coli strain S3030 of recombinant plasmid DNA pER-TA1GyrA-AcSer.EFFECT: invention enables to obtain recombinant human thymosin Alpha 1 with high output and simple production technology.3 cl, 2 dwg, 3 ex