RUI: Genetic and Biochemical Analysis of Extracellular Complementation in Streptomyces coelicolor

本研究的目的是阐明一种清洁剂样分子SapB的结构，SapB是大肠链霉菌形态分化所必需的。这种微生物的发育周期始于外孢子的萌发，导致多核营养菌丝延伸到固体基质中。它们形成一个密集的丝状网络，直到信号分化成向上生长的疏水气生菌丝，然后分离成孢子，开始新的循环。空生菌丝的形成依赖于秃顶基因的产物，秃顶基因反过来调节SapB的产生；突变体产生光滑的菌落，仅由营养基质菌丝组成，并在SapB生物合成中受阻。当纯化的SapB添加到任何突变体在富培养基上生长时，观察到细胞外互补。SapB在分化中具有结构性作用，作为表面活性剂，通过释放菌落/空气界面的表面张力，促进气丝向上生长。SapB的基因及其结构仍然是未知的。本研究计划将使用生化方法纯化和阐明SapB的结构。结构研究将从SapB纯化开始。然后，它的结构将通过质谱分析和核磁共振研究来确定，这些研究将在博士的实验室进行。查尔斯·伊登和弗朗西斯·皮卡德，分别在纽约州立大学石溪分校。SapB结构的解析将促进对胞外通讯在这种丝状细菌形态分化中的作用的理解。此外，这项研究将增加我们对细胞间通讯在一般发育过程中的作用的认识。本科生将参与该项目并接受分子和细胞生物学方面的培训。

The goal of this research is to elucidate the structure of a detergent-like molecule, SapB, required for morphological differentiation in the bacterium Streptomyces coelicolor. The developmental cycle of this microbe begins with the germination of exospores resulting in the extension of multinucleate vegetative hyphae into a solid substrate. These form a dense network of filaments until signaled to differentiate into upwardly growing hydrophobic aerial hyphae, which then septate into spores to begin the cycle anew. Aerial hyphae formation is dependent on the products of the bald (bld) genes, which in turn regulate the production of SapB; bld mutants produce smooth colonies consisting only of vegetative substrate hyphae and are blocked in SapB biosynthesis. Extracellular complementation is observed when purified SapB is added to any bld mutant during growth on rich medium. SapB has a structural role in differentiation, functioning as a surfactant to facilitate upward growth of aerial filaments by releasing the surface tension at the colony/air interface. The gene(s) for SapB and its structure remain elusive. This program of research will use a biochemical approach to purify and elucidate the structure of SapB. Structural studies will begin with SapB purification. Its structure will then be determined by mass spectrometry and nuclear magnetic resonance studies to be performed in the laboratories of Drs. Charles Iden and Francis Picard, respectively at the State University of New York, Stony Brook. The resolution of the structure of SapB will advance the understanding of the role of extracellular communication in the morphological differentiation in this filamentous bacterium. In addition, this research will increase our appreciation of the role of intercellular communication in developmental processes in general. Undergraduate students will be involved in the project and trained in Molecular and Cell Biology.