Division of Bioengineering, Incheon National University, Incheon 22012, Republic of Korea; Research Center for Bio Materials & Process Development, Incheon National University, Incheon 22012, Republic of Korea.
Department of Bioengineering and Nano-Bioengineering, Graduate School of Incheon National University, Incheon 22012, Republic of Korea.
Int J Biol Macromol. 2024 Oct;278(Pt 2):134858. doi: 10.1016/j.ijbiomac.2024.134858. Epub 2024 Aug 18.
The iminosugar class of carbohydrate-active enzyme inhibitors has therapeutic applications in metabolic syndrome conditions, viral infections and cancer. Compared to chemical synthesis, microbial iminosugar production has benefits of cost, sustainability and optimization. In this study, the 1-deoxynojirimycin (DNJ) biosynthetic gene cluster from Bacillus velezensis MBLB0692, and its individual genes, were cloned into Corynebacterium glutamicum (Cg). Characterizations of the encoded aminotransferase GabT1, phosphatase Yktc1, and dehydrogenase GutB1, were performed with purified enzymes and whole cell biocatalysts bearing individual and clustered (TYB) genes. GabT1 showed a variable pattern in its half-reaction with a slow turnover. GutB1 was an alkaline dehydrogenase with a broad substrate specificity and no divalent ion dependency while the zinc-dependent phosphatase Yktc1 had substrate specificity that was both pH- and ion-dependent. The CgYktc1 and CgGutB1 whole cells were viable biocatalysts with wider ranges of substrates than their enzyme counterparts. The CgTYB cells produced mannosidase-inhibiting iminosugars corresponding to mannojirimycin dehydrate (162 m/z) and deoxymannojirimycin (164 m/z). Mannosidase inhibitors have been found to be effective in treating orphan diseases, cancer and viral infections, and their biosynthesis by recombinant C. glutamicum can be optimized for industrial production and novel drug development.
氨基糖类碳水化合物酶抑制剂在代谢综合征、病毒感染和癌症等疾病的治疗中有应用价值。与化学合成相比，微生物法生产氨基糖具有成本低、可持续和可优化等优势。本研究从解淀粉芽孢杆菌 MBLB0692 中克隆了 1-脱氧野尻霉素（DNJ）生物合成基因簇及其单个基因，并将其转入谷氨酸棒杆菌（Cg）。利用纯化酶和携带单个及簇状（TYB）基因的全细胞生物催化剂，对编码氨基转移酶 GabT1、磷酸酶 Yktc1 和脱氢酶 GutB1 的基因进行了表征。GabT1 在其半反应中表现出可变模式，其周转率较慢。GutB1 是一种碱性脱氢酶，具有广泛的底物特异性，不需要二价离子，而依赖锌的磷酸酶 Yktc1 具有依赖 pH 和离子的底物特异性。CgYktc1 和 CgGutB1 全细胞是有活力的生物催化剂，其底物范围比相应的酶更宽。CgTYB 细胞产生了甘露糖苷酶抑制剂氨基糖，对应甘露糖酐脱水物（162 m/z）和脱氧甘露糖（164 m/z）。已发现甘露糖苷酶抑制剂在治疗孤儿病、癌症和病毒感染方面有效，通过重组谷氨酸棒杆菌进行其生物合成可优化工业生产和新型药物开发。
