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

本研究的目的是阐明洗涤剂样分子，SapB，所需的细菌天蓝色链霉菌形态分化的结构。这种微生物的发育周期始于孢子萌发，导致多核营养菌丝延伸到固体基质中。这些菌丝形成一个密集的丝状网络，直到发出信号分化成向上生长的疏水性气生菌丝，然后分隔成孢子，开始新的循环。 气生菌丝的形成依赖于秃（bld）基因的产物，该基因反过来调节SapB的产生; bld突变体产生仅由营养基质菌丝组成的光滑菌落，并且在SapB生物合成中被阻断。当纯化的SapB加入到任何bld突变体中时，在丰富培养基上生长期间观察到细胞外互补。SapB在分化中具有结构作用，作为表面活性剂通过释放在殖民地/空气界面处的表面张力来促进气丝的向上生长。SapB的基因及其结构仍然难以捉摸。这项研究计划将使用生物化学方法来纯化和阐明SapB的结构。结构研究将从SapB纯化开始开始。然后，将分别在斯托尼布鲁克的纽约州立大学的Charles Iden和弗朗西斯Picard博士的实验室通过质谱和核磁共振研究确定其结构。SapB结构的解析将促进对这种丝状细菌中细胞外通讯在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.