Elucidation of expression/regulation of genes involved in microbial polyhydroxylalkanoate (PHA)

为了阐明反硝化细菌和甲硫藻合成/降解PHA的调控机制，本论文从基因/酶/代谢水平进行了研究，取得了以下成果：1.研究了PHA在反硝化细菌和甲基化细菌中的合成/降解规律。在Phap下游发现了一种名为Phar的基因，该基因编码PHA颗粒相关蛋白Phasin。Phar是一种转录因子，与Phap启动子区域结合，也与PHA颗粒结合。结论：1.调节PHA颗粒的大小。参与PHA合成/降解的基因，即PHAA、PHAB、PHAC和phaZ几乎是结构性表达的，与PHA合成水平无关。对参与PHA胞内降解的两种不同类型的PHA解聚酶和PHB寡聚体水解酶进行了分离纯化，并对其生化特性进行了研究。PHA解聚酶是一种能直接从PHB聚合物中合成3-羟基丁酸酯单体的新型解聚酶。

With aim of elucidating regulation mechanism of PHA synthesis/degradation within denitrifyer and methylotroph, Paracoccus denitrificans, studies were carried out on genes/enzymes/metabolism levels, and the following have been achieved.1. A gene named phaR, was discovered downstream of phaP that encoded PHA granule-associated protein, phasin. PhaR was a transcription factor which bound promoter region of phaP and also bound the PHA granule. It regulated the size of PHA granule.2. Genes involved in PHA synthesis/degradation, i.e phaA, phaB, phaC, and phaZ, were expressed almost constitutively irrespective of PHA synthesis level.A metabolite, Coenzyme A, was proposed as a global regulator of PHA biosynthesis.3. Three enzymes involved in the intracellular degradation of PHA, i.e. two different kinds of PHA depolymerases, and PHB oligomer hydrolase, were purified and their biochemical characteristics were elucidated. One of the PHA depolymerases was a novel enzyme which yielded 3-hydroxylbutyrate monomer directly from PHB polymer.