BIOCHEMICAL CHARACTERIZATION OF BACTERIAL CYTOKINESIS

描述（由申请人提供）：细胞分裂将在丝状原核生物链霉菌中进行生化分析。该物种对本研究有利，因为其生长和生存能力不需要细胞分裂，已经存在一组分裂无效突变体，并且已经准备好了对该项目成功至关重要的试剂。越来越多的证据表明，分裂蛋白的功能是一种以有序方式组装的复合体；然而，直接的生化支持复合物是缺乏的。许多分裂蛋白含有至少一个跨膜片段，使复合体的生化表征更具挑战性。因此，现在是应用膜蛋白生化技术的时候了，这需要在该领域进行培训。具体来说，我将使用三种独特但互补的生化攻击来识别链霉菌细胞分裂复合体中的蛋白质：交联，免疫沉淀和柱层析。这些技术已经成功地定义了其他整体膜蛋白机器。我也希望用蛋白质组学鉴定细胞分裂复合体的新成分。这个项目的最终结果将独特地装备我的实验室继续在细胞分裂领域取得重大进展。

DESCRIPTION (provided by applicant): Cell division will be biochemically analyzed in the filamentous prokaryote, Streptomyces coelicolor. This species is advantageous for this study because growth and viability do not require cell division, a collection of division null mutants already exists, and reagents vital to the success of this project have been prepared. Mounting evidence suggests that the division proteins function as a complex that is assembled in an ordered fashion; however direct biochemical support for the complex is absent. Many division proteins contain at east one membrane-spanning segment, making the biochemical characterization of the complex more challenging. Therefore, the time has come to apply membrane protein biochemical techniques, which will require training in the area. Specifically, I will identify the proteins in the Streptomyces cell division complex using three unique but complementary biochemical attacks: cross-linking, immune precipitation, and column Chromatography. These techniques have successfully defined other integral membrane protein machines. I also hope to identify novel components of the cell division complex using proteomics. The net result of this project will uniquely equip my laboratory to continue to make significant strides in the cell division field.