Funktionelle Genomik mit mRNA-Protein-Konjugaten und DNA Mikrochips

近年来，由于几个基因组计划的大力努力，快速自动基因组测序技术以惊人的速度发展。相比之下，对基因组翻译的信息--蛋白质--实际功能的高度平行研究方法的发展远远落后于此。这个项目将探索一种新的策略，用生物组合的方法研究蛋白质之间的功能相互作用。因此，Szostaks博士实验室开发的一种产生mRNA-蛋白质融合的方法将与DNA微阵列技术相结合。从一个cDNA文库开始，将产生一个相应的mRNA-蛋白质融合文库，其中每个蛋白质都与其编码的mRNA共价连接。这些融合经历了活性选择过程，特别是与特定蛋白质靶标的亲和力。由于结合了mRNA，选择性浓缩蛋白的遗传信息可以通过聚合酶链式反应进行扩增，并通过将荧光标记的聚合酶链式反应产物与适当的DNA微阵列杂交来读出。这一策略将允许在基因组水平上对几乎任何生物体的蛋白质-蛋白质和蛋白质-小分子相互作用进行筛选。该方法在寻找新药靶点和研究代谢途径方面具有很大潜力。

In recent years, technology for very fast automated genome sequencing has been developed at an amazing speed due to strong efforts in several genome projects. In contrast, development of methods for highly parallel investigation of the actual function of the genome`s translated message - the proteins - is in lagging far behind.This project will investigate a new strategy of functional interaction studies on proteins in a bio-combinatorial approach. Therefore, a method for the generation of mRNA-protein fusions, developed in Prof. Dr. Szostaks laboatory, will be combined with DNA microarray technology. Starting from a cDNA library, a corresponding mRNA-protein fusion library will be generated where every protein is covalently linked to its encoding mRNA. These fusions are subjected to an actvity selection procedure, specifically affinity to a certain protein target. As a result of the attached mRNA, the genetic information of the selectively enriched protein can be amplified by PCR an read out by hybridizing the fluorescently labeled PCR product with an appropriate DNA microarray. This strategy will allow screening for protein-protein and protein-small molecule interactions at genomic scale for virtually any organism. The method has great potential in the search for new drug targets an investigating metabolic pathways in general.