Expression of recombinant endochitinase from the Antarctic bacterium, Sanguibacter antarcticus KOPRI 21702 in Pichia pastoris by codon optimization
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- Expression of recombinant endochitinase from the Antarctic bacterium, Sanguibacter antarcticus KOPRI 21702 in Pichia pastoris by codon optimization
- Other Titles
- 코돈최적화에 의한 남극세균 상귀박터 안탁티쿠스 코프리 21702 유래의 내재키티네이즈의 피키아 파스토리스 내에서의 재조합 발현
- Koh, Hye Yeon
Lee, Sung Gu
Na, Deuk Chae
Yim, Joung Han
Lee, Hong Kum
Han, Se Jong
- Biochemistry & Molecular Biology; Biotechnology & Applied Microbiology
- Antarctica; Codon optimization; Endochitinases; Pichia pastoris; Recombinant
- Issue Date
- Koh, Hye Yeon, et al. 2010. "Expression of recombinant endochitinase from the Antarctic bacterium, Sanguibacter antarcticus KOPRI 21702 in Pichia pastoris by codon optimization". PROTEIN EXPRESSION AND PURIFICATION, 71(1): 108-114.
- An endochitinase was previously purified and the gene was cloned from the psychrophilic Antarctic bacterium,Sanguibacter antarcticus (KCTC 13143). In the present study, recombinant endochitinase,rChi21702, was expressed using a yeast expression system (Pichia pastoris) and codon optimization. The expressed rChi21702 was purified by Phenyl-Sepharose column chromatography. Optimal expression yielded 1-mg purified enzyme from 1-L bioreactor culture. When p-NP-(GlcNAc)2 was used as a substrate, the specific activity of the enzyme was determined to be 20 U/mg. In vitro assays and thin-layer chromatography demonstrated that the recombinant enzyme has endochitinase activity that produces diacetyl-chitobiose as a dominant end product when chitooligomers, colloidal chitin, and the chromogenic p-NP-(GlcNAc)2 are used as substrates. Optimal activity for rChi21702 was observed at 37oC and a pH of 7.6. Interestingly, rChi21702 exhibited 63% of optimal activity at 10oC and 44% activity at 0oC. Taken together, the results indicate that rChi21702 has psychrotolerant endochitinase activity even after recombinant expression in yeast cells.
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