Combinatorial biosynthetic libraries from cyanobacteria

该实验室计划的总体目标是通过创新方法产生天然产物衍生物质库，并在针对癌症的“智能”生物测定系统中评估这些物质。尽管科学和医学的许多领域都在努力寻找针对这一复杂疾病家族的有效治疗方法，但癌症的发病率和发病率仍然非常低迷。该实验室计划将重点研究海洋蓝细菌的独特化学性质，并将与该计划项目拨款申请的其他实验室计划的努力相结合，克隆天然产物生物合成基因，并在方便的宿主生物体委内瑞拉链霉菌中过度表达这些基因。在这一追求中将采取四个平行轨道：1）我们的实验室将为实验室计划#1提供培养的海洋蓝藻，用于实验室计划#2中的基因组DNA曝晒和随后的组合生物学工作； 2) 我们的实验室将向实验室计划#2提供有关关键“定制”酶的生物合成途径和部分蛋白质序列的信息，以帮助克隆与癌症和多样性产生高度相关的特定海洋蓝藻代谢物的基因； 3) 我们的实验室将与诺华“智能”检测（实验室计划#3）集成，以分离和结构定义源自上述集成分子生物学方法的新先导化合物； 4）我们的实验室将使用从海洋蓝细菌中克隆并通过蛋白质工程技术进行修饰的独特定制酶来生物转化与癌症相关的其他天然产物结构。二极管阵列 HPLC 方法将用于消除母体结构、已知化合物和组合生物合成文库中的有害物质，这些物质在诺华“Smart”测定中显示出活性。

The overall goal of this Laboratory Program is to produce libraries of natural product-derived substances by innovative methods and to evaluate these in "Smart" bioassay systems targeted to cancer. Despite intensive effort in many fields of science and medicine to find effective treatments for this complex family of diseases, the incidence and morbidity due to cancer continues to look very bleak. This Laboratory Program will focus on the unique chemistries of marine cyanobacteria, and will integrate with the efforts of the other laboratory programs of this Program Project Grant application to clone natural products biosynthetic genes and over express these in a convenient host organism, Streptomyces venezuelae. Four parallel tracks will be taken in this pursuit: 1) our laboratory will provide cultured marine cyanobacteria to Laboratory Program #1 for genomic DNA insolation and subsequent combinatorial biology efforts in Laboratory Program #2; 2) our laboratory will provide information to Laboratory Program #2 on biosynthetic pathways and partial protein sequences of key "tailoring" enzymes to aid in the cloning of genes of specific marine cyanobacterial metabolites of high relevance to cancer and diversity generation; 3) our laboratory will integrate with Novartis "Smart" assays (Laboratory Program #3) to isolate and structurally define new lead compounds deriving from the above integrated molecular biological approaches; and 4) our laboratory will use unique tailoring enzymes cloned from marine cyanobacteria and modified by protein engineering techniques to biotransform additional natural product structures of cancer relevance. Diode-array HPLC methods will be used to dereplicate parent structures, known compounds, and nuisance substance in the combinatorial biosynthetic libraries showing activity in the Novartis "Smart" assays.