Hwang, Dohyeon Kim, Sang Ah Yang, Eun Gyeong Song, Hyun Kyu Chung, Hak Suk 2024-01-20T03:30:21Z 2024-01-20T03:30:21Z 2021-09-05 2016-10 1046-5928 https://pubs.kist.re.kr/handle/201004/123618 Since human Caspase-3, a member of the cysteine protease family, plays important roles not only in the apoptosis pathway as an executioner protein, but also in neurological disorders as a critical factor, biomedical researchers have been interested in the development of modulators of caspase-3 activity. Such studies require large quantities of purified active caspase-3. So far, purification of soluble caspase-3 from full-length human caspase-3 in Escherichia coli (E. coli) yields only several mg from a liter of culture media. Therefore, a number of alternative strategies to purify active caspase-3 have been described in the literature, including refolding and protein engineering. In this study, we systematically study the effects of host E. coli strains and growth conditions on purifications of active caspase-3 from full-length human caspase-3. Using a combination of conditions that include use of the C41(DE3) strain, low-temperature expression, and auto-induction that induces caspase-3 expression depending on metabolic state of the individual host cell, we are able to obtain 14-17 mg caspase-3 per liter of culture, an amount that is about 7 times larger than published results. This optimized expression and purification method for caspase-3 can be easily scaled up to facilitate the demand for active enzyme. (C) 2016 Elsevier Inc. All rights reserved. English ACADEMIC PRESS INC ELSEVIER SCIENCE A facile method to prepare large quantities of active caspase-3 overexpressed by auto-induction in the C41(DE3) strain Article 10.1016/j.pep.2016.06.004 1 PROTEIN EXPRESSION AND PURIFICATION, v.126, pp.104 - 108 PROTEIN EXPRESSION AND PURIFICATION 126 104 108 N scie scopus 000380579400015 2-s2.0-84975827991 Biochemical Research Methods Biochemistry & Molecular Biology Biotechnology & Applied Microbiology Biochemistry & Molecular Biology Biotechnology & Applied Microbiology Article ESCHERICHIA-COLI PROTEIN-PRODUCTION APOPTOSIS Caspase-3 Auto-induction C41(DE3) Soluble expression Enzyme kinetics