RUI: Molecular Analysis of PHB/V Biosynthesis

聚-B-羟基烷酸酯(PHA‘s)是一种存在于多种细菌中的聚合物，由于其具有可生物降解的热塑性性质，近年来引起了人们的极大兴趣。PHA聚合物的物理性质由其单体单元中的碳数决定。例如，聚β-羟基丁酸酯(PHB=C4单体单元)要灵活得多。为此，在PHB/V共聚物的开发中有相当多的正在进行的研究。产碱杆菌是第一批被证明能够产生PHB或PHB/V的细菌之一，具体取决于原料。我们的研究小组已经将红曲霉合成聚合物的基因克隆到大肠杆菌中，在那里我们利用它们来产生大量的PHB胞内沉淀物。在使用标准大肠杆菌菌株进行的初步实验中，即使改变原料以刺激PHB/V的合成，我们也无法检测到共聚物的合成。然而，最近我们能够使用一种大肠杆菌突变体进行共聚合成。初步结果表明，共聚物在大肠杆菌中的合成与在富营养化弧菌中发现的相似。这项研究的目的是利用新开发的大肠杆菌系统进一步发展我们对PHB/V合成和PHB/V遗传学的知识。这项研究将通过对主要产生PHB和PHB/V的细菌进行全面调查，克隆选定的PHB/V遗传途径，分析能够生产PHV的大肠杆菌中的这些途径，以及利用基因技术操纵共聚物的合成和结构来完成这项研究。

Poly-B-hydroxyalkanoates (PHA's), polymers found in many bacteria, have attracted considerable interest in recent years because they exhibit biodegradable thermoplastic properties. The physical properties of PHA polymers are dictated by the number of carbons in their monomeric unit. For instance, poly-beta-hydroxybutyrate (PHB= C4 monomeric unit) is much more flexible. For this reason, there is considerable ongoing research in the development of the PHB/V copolymer. The bacterium Alcaligenes eutrophus is one of the first bacteria shown to be capable of producing either PHB or PHB/V, depending on the feedstock. The genes for polymer synthesis in A. euthrophus have been cloned into Escherichia coli by our research group where we have employed them to produce large intracellular deposits of PHB. In initial experiments using standard E. coli strains we have not been able to detect copolymer synthesis, even when the feedstock is altered to stimulate PHB/V synthesis. However, recently we were able to obtain copolymer synthesis employing an E. coli mutant. Preliminary results indicate that copolymer synthesis in E. coli is similar to that found in A. eutrophus. The purpose of this research is to further develop our knowledge of PHB/V synthesis and PHB/V genetics using the newly develop E. coli system. This research will be done by conducting a comprehensive survey of bacteria that produce primarily PHB and PHB/V, cloning of selected PHB/V genetic pathways, analysis of these pathways in E. coli capable of PHV production, and manipulation of copolymer synthesis and structure using genetic techniques.