Plant Biofilm Inhibitors to Discover Biofilm Genes

描述（由申请人提供）：为了表征不影响细菌生长的无毒、有效的抗生物膜剂，提出了一项研究计划来研究天然存在的海洋呋喃酮和其他植物衍生化合物抑制大肠杆菌形成的能力。大肠杆菌生物膜。此外，生物膜形成的遗传基础将使用时间的DNA微阵列研究，敲除，和NMR结构测定所发现的生物膜调节蛋白。该项目将发现E.大肠杆菌生物膜形成的两个互补的方法，使用DNA微阵列：直接识别生物膜基因和抑制生物膜与植物化合物。此外，该项目将确定植物来源的 生物膜抑制剂影响生物膜，并将发现其它抑制生物膜的植物衍生化合物。 被发现的生物膜基因将成为其他类型化合物的丰富靶标，这些化合物可用于防止工业和医学中的生物膜。 该项目的具体目标是： 1.从红色的Delisea pulchra中合成天然呋喃酮，并使用共聚焦显微镜和96孔快速测定法评估这些呋喃酮和其他植物衍生化合物防止生物膜形成的能力 2.评价植物源抑制剂对大肠杆菌基因表达的影响。使用DNA微阵列（确定哪些基因受到影响，而抑制剂不影响生长） 3.用DNA微阵列发现E.大肠杆菌（淹没生物膜） 4.通过抑制与呋喃酮的通讯，使用DNA微阵列发现细胞间信号传导基因 5.研究基因芯片在生物膜形成方面阐明的基因的重要性， 使用用绿色荧光蛋白标记的敲除突变体并通过标记生物膜调节蛋白的启动子的结构 6.使用NMR确定重要的生物膜调节蛋白的结构和功能 7.基于遗传信息（所述基因 诱导/抑制）和蛋白质信息识别（通过NMR）

DESCRIPTION (provided by applicant): To characterize non-toxic, effective anti-biofilm agents that do not affect bacterial growth, a research program is proposed to investigate the ability of naturally-occurring marine furanones and other plant-derived compounds to inhibit the formation of E. coli biofilms. In addition, the genetic basis of biofilm formation will be investigated using temporal DNA microarray studies, knock-outs, and NMR structure determinations of the discovered biofilm regulatory proteins. This project will discover the genes required for E. coli biofilm formation by two complementary approaches using DNA microarrays: direct identification of biofilm genes and by inhibiting biofilms with plant compounds. In addition, this project will identify how the plant-derived biofilm inhibitors influence biofilms, and will discover other plant-derived compounds which inhibit biofilms. The biofilm genes that are found will be rich targets for other classes of compounds that may be used to prevent biofilms in industry and medicine. The specific objectives of this project are: 1. synthesize natural furanones from the red alga Delisea pulchra and evaluate these furanones and other plant-derived compounds for their ability to prevent biofilm formation using confocal microscopy and 96-well rapid assays 2. evaluate the impact of the plant-derived inhibitors on the gene expression of E. coli using DNA microarrays (identify which genes are affected while the inhibitor does not affect growth) 3. discover biofilm genes using DNA microarrays for E. coli (submerged biofilm) 4. discover cell-to-cell signaling genes using DNA microarrays by inhibiting communication with furanones 5. study the importance of the genes elucidated with the gene chips in terms of biofilm formation and architecture using knock-out mutants that are labeled with the green fluorescence protein and by tagging the promoters of biofilm regulatory proteins 6. determine the structure and thereby the function of the important biofilm regulatory proteins using NMR 7. deduce a model for bacterial biofilm formation based on the genetic information (which genes induced/repressed) and protein information discerned (via NMR)