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

本研究的目的是阐明天蓝色链霉菌形态分化所需的类似洗涤剂的分子SapB的结构。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底物中。它们形成密集的丝状网络，直到发出信号分化成向上生长的疏水气生菌丝，然后分离成孢子，重新开始循环。气生菌丝的形成依赖于秃顶(BLD)基因的产物，BLD基因反过来调节SapB的产生；BLD突变株产生的光滑菌落只由营养底物菌丝组成，并阻止SapB的生物合成。当纯化的SapB被加入到任何BLD突变体中时，在富含菌基上的生长过程中观察到胞外互补。SapB在分化过程中具有结构性作用，通过释放菌落/空气界面的表面张力，发挥表面活性物质的作用，促进气生丝向上生长。SapB的基因(S)及其结构仍然难以捉摸。这项研究计划将使用生化方法来纯化和阐明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学和核磁共振研究确定，这些研究将分别在纽约州立大学石溪分校的查尔斯·伊登博士和弗朗西斯·皮卡德博士的实验室进行。对SapB结构的解析将促进对这种丝状细菌形态分化中细胞外通讯作用的理解。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.