Combinatorial biosynthetic libraries from cyanobacteria

该实验室计划的总体目标是通过创新方法生产天然产品衍生物质库，并在针对癌症的“智能”生物测定系统中对这些物质进行评估。尽管许多科学和医学领域都在努力为这一复杂的疾病家族寻找有效的治疗方法，但癌症的发病率和发病率仍然看起来非常惨淡。该实验室计划将重点研究海洋蓝藻的独特化学成分，并将与本计划项目资助申请中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的轨道：1)我们的实验室将为实验室计划#1提供培养的海洋蓝藻，用于基因组DNA照射和随后的实验室计划#2的组合生物学研究；2)我们的实验室将为实验室项目#2提供关于生物合成途径和关键“剪裁”酶的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产物的基因；3)我们的实验室将与诺华的“智能”分析（实验室计划#3）相结合，从上述整合的分子生物学方法中分离和结构定义新的先导化合物；4)我们的实验室将使用从海洋蓝藻中克隆的独特剪裁酶，并通过蛋白质工程技术进行修饰，以生物转化其他与癌症相关的天然产物结构。二极管阵列高效液相色谱方法将用于在诺华“Smart”检测中显示活性的组合生物合成文库中去复制亲本结构、已知化合物和有害物质。

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.