Biotechnology and Bioprocess Engineering

, Volume 24, Issue 2, pp 298–307 | Cite as

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

  • Seung Woo Lee
  • Nam Hyeon Kang
  • Jang Won ChoiEmail author
Research Paper


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


anti-obesity aprE promoter Bacillus subtilis brown adipocyte granulocyte-colony stimulating factor thermogenic activity 


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This study was supported by Daegu University Research Grant 2014.


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Copyright information

© The Korean Society for Biotechnology and Bioengineering and Springer 2019

Authors and Affiliations

  • Seung Woo Lee
    • 1
  • Nam Hyeon Kang
    • 2
  • Jang Won Choi
    • 1
    Email author
  1. 1.Department of BioindustryDaegu UniversityGyeongsanKorea
  2. 2.Department of BiotechnologyDaegu UniversityGyeongsanKorea

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