深海链霉菌SCSIO ZJ46来源的抗菌环肽desotamides的生物合成研究

Natural products derived from marine microorganisms are important sources for drug discovery and development. Desotamides A-D are cyclic peptides produced by Streptomyces scopuliridis SCSIO ZJ46 isolated from the deep South China Sea. They possess novel structural elements including L-allo-isoleucine and formylkynurenine/kynurenine, and showed strong antibacterial activities against Streptococcus Natural products derived from marine microorganisms are important sources for drug discovery and development. Desotamides A-D are cyclic peptides produced by Streptomyces scopuliridis SCSIO ZJ46 isolated from the deep South China Sea. They possess novel structural elements including L-allo-isoleucine and formylkynurenine/kynurenine, and showed strong antibacterial activities against Streptococcus pnuemoniae and methicillin-resistant Staphylococcus epidermidis. However, the biosynthetic mechanism of desotamides and their novel structural elements has not been elucidated yet. A combination of multiple techniques will be applied to: 1) obtain the gene cluster governing desotamides biosynthesis, 2) analyze and identify the unique genes involved in the biosynthesis of the novel L-allo-isoleucine and formylkynurenine/kynurenine structural units, 3) overexpress the unique genes and perform biochemical assay of the enzymes responsible for the biosynthesis/or tailoring the two aforementioned two unique structural units. Thus, this project will pave the way for the discovery of new biocatalyst and generation of new antibacterial derivatives using combinatorial biosynthesis.

海洋微生物来源的天然产物是药物创制的重要源泉。Desotamides A-D是我们从一株南海深海链霉菌Streptomyces scopuliridis SCSIO ZJ46中筛选分离到的4个环肽类抗生素，其化学结构新颖，含有非天然的L-别异亮氨酸和甲酰基犬尿氨酸/犬尿氨酸结构单元，对肺炎链球菌和耐甲氧西林的表皮葡萄球菌具有较强的抑制活性。目前，desotamides及其独特结构单元的生物合成机制尚不清楚。本项目拟综合运用多门学科专业技能：1）获得desotamides生物合成基因簇，2）挖掘生物合成基因簇中与L-别异亮氨酸、甲酰基犬尿氨酸/犬尿氨酸生物合成和修饰相关的新功能基因，3）对新功能基因进行蛋白表达和酶反应动力学表征，以阐明desotamides及其“非天然”氨基酸结构单元的生物合成机制。本项目的开展，将为新型功能酶催化剂的挖掘和利用组合生物合成技术获得新结构活性衍生物奠定基础。

Desotamides A-D是我们从一株南海深海链霉菌Streptomyces scopuliridis SCSIO ZJ46中筛选分离到的4个环肽类抗生素，化学结构新颖，含有“非天然”氨基酸结构单元L-别异亮氨酸和甲酰基犬尿氨酸/犬尿氨酸，对肺炎链球菌和耐甲氧西林的表皮葡萄球菌具有较强的抑制活性。本项目通过对产生菌SCSIO ZJ46进行全基因组扫描测序和生物信息学分析鉴定了desotamides完整生物合成基因簇；通过将基因簇在异源宿主中进行异源表达，获得新结构衍生物desotamide G；利用PCR-Targeting和接合转移技术对基因簇中的生物合成前体合成基因、调控基因和转运基因进行突变，共构建7株双交换突变株；通过对突变株进行发酵和HPLC分析，对突变株产生的新中间体化合物进行分离、鉴定，明确各基因在desotamides生物合成过程中的功能，阐明了特殊结构单元L-别异亮氨酸的生物合成机制，推导了desotamides生物合成途径；首次发现了负责特殊结构单元L-别异亮氨酸生物合成的氨基酸转移酶和异构酶，并对它们进行了体外酶促反应机理的研究。研究成果在国际主流期刊杂志发表SCI论文2篇，其中1篇发表在化学生物学国际权威期刊杂志J. Am. Chem. Soc.上，被同期JACS Spotlights（2016, 138, 461）和Natural Products Reports (2016, 33, 530)作为研究亮点评述。申请发明专利1项。本项目为新型功能酶催化剂的挖掘和利用组合生物合成技术开发、改造desotamides类化合物提供了理论基础和技术支持。