Genetic dissection of the biosynthetic route to gentamicin A 2 by heterologous expression of its minimal gene set

作者：Je Won Park, Jay Sung Joong Hong, Niranjan Parajuli, Won Seok Jung, Sung Ryeol Park, Si‐Kyu Lim, Jae Kyung Sohng, Yeo Joon Yoon · 发表于：Proceedings of the National Academy of Sciences · 年份：2008 · DOI：10.1073/pnas.0803164105 · 被引用次数：63 · 研究领域：Microbial Natural Products and Biosynthesis、Carbohydrate Chemistry and Synthesis、RNA and protein synthesis mechanisms

Since the first use of streptomycin as an effective antibiotic drug in the treatment of tuberculosis, aminoglycoside antibiotics have been widely used against a variety of bacterial infections for over six decades. However, the pathways for aminoglycoside biosynthesis still remain unclear, mainly because of difficulty in genetic manipulation of actinomycetes producing this class of antibiotics. Gentamicin belongs to the group of 4,6-disubstituted aminoglycosides containing a characteristic core aminocyclitol moiety, 2-deoxystreptamine (2-DOS), and the recent discovery of its biosynthetic gene cluster in Micromonospora echinospora has enabled us to decipher its biosynthetic pathway. To determine the minimal set of genes and their functions for the generation of gentamicin A(2), the first pseudotrisaccharide intermediate in the biosynthetic pathway for the gentamicin complex, various sets of candidate genes from M. echinospora and other related aminoglycoside-producing strains were introduced into a nonaminoglycoside producing strain of Streptomyces venezuelae. Heterologous expression of different combinations of putative 2-DOS biosynthetic genes revealed that a subset, gtmB-gtmA-gacH, is responsible for the biosynthesis of this core aminocyclitol moiety of gentamicin. Expression of gtmG together with gtmB-gtmA-gacH led to production of 2'-N-acetylparomamine, demonstrating that GtmG acts as a glycosyltransferase that adds N-acetyl-d-glucosamine (GLcNA) to 2-DOS. Expression of g...