Functional Secretion of Granulocyte Colony Stimulating Factor in Bacillus subtilis and Its Thermogenic Activity in Brown Adipocytes

摘要: Granulocyte colony-stimulating factor (G-CSF) is involved in generation of colonies hematopoietic cells by stimulation bone marrow cells, resulting the survival, proliferation, differentiation, and function neutrophil precursors mature neutrophils. To express secrete G-CSF Bacillus system, gene was amplified using gene-specific primers template DNA (pUNO1-hGCSFb). As a result, PCR product 530 bp obtained, sequenced, analyzed BLAST program. The open reading frame encoded 204-amino acid polypeptide precursor consisting 30-residue signal sequence 174-residue polypeptide. Recombinant secretion vector (pRBAS-CSF3, 6.96 kb) constructed pRBAS which contains promoter (0.45 (87 bp) alkaline protease (aprE), ribosome binding site (GGAGAGGG) transferred to subtilis LKS. protein secreted optimized increase productivity (19% total proteins, 0.12 mg/ml). culture broth harvested followed metal chelating, ethanol precipitation, gel chromatography. purified tested for thermogenic activity cultured brown adipocytes. activated HIB1B adipocytes increasing expression levels fat-specific marker proteins (UCP1 PRDM16) genes (Lhx8, Ppargc1, Prdm16, Ucp1, Zic1) This result suggests that per se or G-CSF-containing foodstuff can be used treat obesity.