High-Level Expression of A β-Mannanase AuMan5A<sup>loop/H321G</sup> in Kluyveromyces lactis GG799

doi:10.3969/j.issn.1673-1689.2019.07.004

Home > Archive>Volume 38, Issue 7, 2019 >26-33. DOI:10.3969/j.issn.1673-1689.2019.07.004

High-Level Expression of A β-Mannanase AuMan5Aloop/H321G in Kluyveromyces lactis GG799
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                        10.3969/j.issn.1673-1689.2019.07.004
                    
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To prepare the food-grade β-mannanase，a β-mannanase mutant gene Auman5A^loop/H321G was expressed in Kluyveromyces lactis GG799. Auman5A^loop/H321G，gene constructed based on a glycoside hydrolase family 5 β-mannanase gene（Auman5A） from Aspergillus usamii，was inserted into the signal peptide α-MF downstream of the expression plasmid pKLAC1，forming a recombinant plasmid pKLAC1-Auman5A^loop/H321G. Then，pKLAC1-Auman5A^loop/H321G was electroporated into Kluyveromyces lacti GG799. After induction by galactose and enzyme activity analysis，a transformant GG799/Auman5A< sup >loop/H321G expressing the highest recombinant β-mannanase（reAuMan5A^loop/H321G） activity of 54.9 U/mL was selected for subsequent studies. The apparent molecular weight of reAuMan5A^loop/H321G was estimated to be about 55 000 by SDS-PAGE analysis. Furthermore，the β-mannanase activity produced by GG799/Auman5A^loop/H321G was increased by 254.6% after primary optimization of the initial pH and loading volume of YPG medium as well as the species，initial concentration and supplementary mode of inducer. As a result，the high-level expression of reAuMan5A^loop/H321G in K. lactis was actualized at the optimized fermentation process.

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TANG Shihan, LI Xueqing, YUAN Fengjiao, LI Jianfang, WU Minchen. High-Level Expression of A β-Mannanase AuMan5A^loop/H321G in Kluyveromyces lactis GG799[J]. Journal of Food Science and Biotechnology,2019,38(7):26-33.

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