Anthracyclines are an important class of natural antitumor drugs. They have a conservative aromatic tetracycline backbone that is substituted with different deoxyglucoses. The deoxyglucoses are crucial for the biological activity of many bacterial natural products after the proper modification from glycosyltransferases (GTs). The difficulty in obtaining highly purified active GTs has prevented biochemical studies on natural product GTs. In this paper, a new Escherichia coli fusion plasmid pGro7′, which introduces the Streptomyces coelicolor chaperone genes groEL1, groES and groEL2, was constructed. The glycosyltransferase DnmS from Streptomyces peucetius ATCC 27952 was co-expressed with the plasmid pGro7′, and unprecedented high-efficiency and soluble expression of DnmS in the E. coli expression system was realized. Subsequently, the reverse glycosylation reaction characteristics of DnmS and DnmQ were verified. We found that DnmS and DnmQ had the highest enzyme activity when they participated in the reaction at the same time. These studies provide a strategy for the soluble expression of GTs in Streptomyces and confirm the reversibility of the catalytic reaction of GTs. This provides a powerful method for the production of active anthracyclines and to enhance the diversity of natural products.

中文翻译：

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链霉菌ATCC 27952糖基转移酶DnmS的表达纯化及其糖基转移酶逆反应催化表征研究

蒽环类药物是一类重要的天然抗肿瘤药物。它们具有被不同脱氧葡萄糖取代的保守芳香四环素主链。经过糖基转移酶 (GT) 的适当修饰后，脱氧葡萄糖对许多细菌天然产物的生物活性至关重要。获得高度纯化的活性 GT 的困难阻碍了对天然产物 GT 的生化研究。本文构建了一个新的大肠杆菌融合质粒pGro7'，该质粒引入了天蓝色链霉菌伴侣基因groEL1、groES和groEL2。将来自 Streptomyces peucetius ATCC 27952 的糖基转移酶 DnmS 与质粒 pGro7' 共表达，实现了 DnmS 在大肠杆菌表达系统中前所未有的高效可溶性表达。随后，验证了DnmS和DnmQ的反向糖基化反应特性。我们发现当DnmS和DnmQ同时参与反应时酶活性最高。这些研究为 GTs 在链霉菌中的可溶性表达提供了策略，并证实了 GTs 催化反应的可逆性。这为生产活性蒽环类化合物和增强天然产物的多样性提供了一种有效的方法。