Development of a recombinant d-mannose isomerase and its characterizations for d-mannose synthesis

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• 作者：Xing Hu^a ; ^b ; Peng Zhang^b ; Ming Miao^a ; ^b ; Tao Zhang^a ; ^b ; Bo Jiang^a ; ^b ; ^{bjiang@jiangnan.edu.cn" class="auth_mail" title="E-mail the corresponding author}
• 关键词：d-mannose isomerase ; Gene expression ; Biotransformation
• 刊名：International Journal of Biological Macromolecules
• 出版年：2016
• 出版时间：August 2016
• 年：2016
• 卷：89
• 期：Complete
• 页码：328-335
• 全文大小：1527 K

文摘

d-Mannose isomerase (MIase) catalyzes the conversion of d-fructose to d-mannose. In this study, the MIase encoding gene (yihS) from Escherichia coli BL21 contains an ORF of 1242 bp, was cloned and expressed in Bacillus subtilis WB800. This heterologous expression resulted in a hexamer with a molecular weight of 274.5 kDa and T_m of 61.4 °C. Efficient MIase secretory expression by the robust recombinant B. subtilis was achieved with activity of 51.2 U/ml (d-mannose forming). Its optimal temperature and pH were 45.0 °C and 7.0, respectively. Using d-fructose as the substrate, K_m, k_cat and catalytic efficiency value of kinetic reaction were 203.7 ± 6.7 mM, 27.7 ± 0.7 s⁻¹ and 136.0 ± 2.9 M⁻¹ s⁻¹, respectively. The production of d-mannose reached about 150 g/l with approximately 25% turnover yield under the optimum conditions. These results demonstrated that B. subtilis was a promising candidate of MIase expression system for d-mannose production.