Efficient production of recombinant cystatin C using a peptide-tag, 4AaCter, that facilitates formation of insoluble protein inclusion bodies in Escherichia coli

Masahiro Hayashi, Shigehisa Iwamoto, Shinya Sato, Shigeo Sudo, Mari Takagi, Hiroshi Sakai, Tohru Hayakawa

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Cystatin C is a cysteine protease inhibitor produced by a variety of human tissues. The blood concentration of cystatin C depends on the glomerular filtration rate and is an endogenous marker of renal dysfunction. Recombinant cystatin C protein with high immunogenicity is therefore in demand for the diagnostic market. In this study, to establish an efficient production system, a synthetic cystatin C gene was designed and synthesized in accordance with the codon preference of Escherichia coli genes. Recombinant cystatin C was expressed as a fusion with a peptide-tag, 4AaCter, which facilitates formation of protein inclusion bodies in E. coli cells. Fusion with 4AaCter-tag dramatically increased the production level of cystatin C, and highly purified protein was obtained without the need for complicated purification steps. The purity and yield of the final product was estimated as 87 ± 5% and 7.1 ± 1.1 mg/l culture, respectively. The recombinant cystatin C prepared by our method was as reactive against anti-cystatin C antibodies as native human cystatin C. Our results suggest that protein production systems using 4AaCter-tag could be a powerful means of preparing significant amounts of antigen protein.

Original languageEnglish
Pages (from-to)230-234
Number of pages5
JournalProtein Expression and Purification
Volume88
Issue number2
DOIs
Publication statusPublished - Mar 11 2013

Keywords

  • 4AaCter-tag
  • Bacillus thuringiensis
  • Biomarker for renal dysfunction
  • Cry4Aa toxin
  • Formation of insoluble protein inclusion bodies
  • Production of antigen protein
  • Recombinant cystatin C

ASJC Scopus subject areas

  • Biotechnology

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