Heterologous Expression and Purification of Endo-Beta-N-Acetylglucosaminidase from Ogataea minuta

doi:10.3969/j.issn.1673-1689.2018.12.013

Home > Archive>Volume 37, Issue 12, 2018 >1313-1318. DOI:10.3969/j.issn.1673-1689.2018.12.013

Heterologous Expression and Purification of Endo-Beta-N-Acetylglucosaminidase from Ogataea minuta
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                        10.3969/j.issn.1673-1689.2018.12.013
                    
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To express active endo-beta-N-acetylglucosaminidase（ENGase）from methylotrophic yeast Ogataea minuta（Endo-Om） in Escherichia coli. The Endo-Om was overexpressed by using pET system. After optimizing host strainand culture condition，we purified the recombinant protein by using Ni2+ metal chelating column.Using the purified Endo-Om，ENGase activity against fluorescent-labeled oligosaccharide was measured. The recombinant protein was well expressed by IPTG induction，however，the majority remained in the insoluble fraction. The insolubility of the recombinant was improved by culturing the E. coli cells in Overnight ExpressTM Instant LB Medium（Merck Millipore） at 16 ℃. The purified protein showed hydrolysis activity to the fluorescent- labeled oligosaccharide. The functional Endo-Om was successfully obtained in this study，it could accelerate the study on its structural and biological function.

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JIA Yuanling, KITAJIMA Toshihiko, LI Zijie, GAO Xiaodong, NAKANISHI Hideki. Heterologous Expression and Purification of Endo-Beta-N-Acetylglucosaminidase from Ogataea minuta[J]. Journal of Food Science and Biotechnology,2018,37(12):1313-1318.

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Online: January 04,2019
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