利用微生物生產不同種類聚羥基烷酸(PHAs)

Producing different polyhydroxyalkanates by microorganisms

唐世珍(Shih-Chen Tang)

指導教授 ： 康世旭

元智大學/工程學院/生物科技暨生物資訊研究所/碩士(2007年)

https://doi.org/10.6838/YZU.2007.00090

摘要

先前的研究指出，TCA cycle中的succinyl-CoA可以轉變成methylmalonyl-CoA, methylmalonyl-CoA去掉一個CO2後形成propionyl-CoA，最後經由PhaA與PhaB代謝成PHAs(polyhydroxyvalerate, PHV)（Paulo et al., 2003），但目前並沒有研究指出可以用生物產生高純度PHV的方法。本研究主要是探討提供不同的碳源下Bacillus sp.是否會產生不同種類的PHAs，以及將只合成中長碳鏈PHA的P. putida KT2442利用基因轉質的技術，使其能同時合成短碳鏈及中長碳鏈PHA。
Bacillus sp.在不同的碳源下，如succinate、glucose、sucrose、糖蜜及有機酸，發現除了有機酸不生長外，其他都可以合成胞內的脂質堆積。在氣相層析儀和核磁共振儀的分析中顯示，在以glucose和 sucrose為碳源時胞內的脂質堆積只有poly-ß-hydroxybutyrate (PHB)，累積量約25 ％，以succinate為碳源時胞內的脂質堆積只有PHV，累積量約15 ％，這樣的結果是非常重要的。
我們將Bacillus sp.所產的PHV經純化後，再利用GPC、TGA與DSC作物裡性質及化學性質的分析。又將B. megaterium的PHA合成基因，大小約4.1kb，轉質入帶大桿菌中，以glucose與succinate培養，再以氣相層析儀分析，發現當分別以glucose及succinate為碳源時，改質過的大腸桿菌皆產生PHV。
另外，利用基因轉質技術在P. putida KT2442中放入R. eutropha的PHA operon，在以葡萄糖酸鈉與月桂酸及油酸為碳源培養下，以氣相層析儀分析中顯示，改質過的P. putida KT2442可以同時生成短碳鏈的PHB與中長碳鏈PHA。進一步探討能代謝succinate的細菌是否也會有與Bacillus sp.相同之結果。

關鍵字

聚羥基烷酸,succinate

並列摘要

Polyhydroxyalkanoic acids (PHAs), are biosynthetic polyester for carbon source storage, produced by a variety of bacteria and archaea under nutrient-unbalance condition with excess carbon source. PHAs have the chemical properties similar to polypropylene (PP) and are biodegradable, insoluble in water, non-toxic, biocompatible, piezoelectric thermoplastic. These features make PHAs are suitable for many applications. According the monomer composition, PHAs are classified into two types, short chain length PHAs (scl-PHAs) with C3–C5 hydroxyacids and medium chain length PHAs (mcl-PHAs) with C6–C16 hydroxyacids. Depending on the monoers compositions, the properties and application of the PHAs vary.
In this study, recombination technology and feeding strategies with different carbon source were used to change the composition for more application. Bacillus sp. can use glucose to produce polyhydroxybutyrate (PHB) when use sodium succinat dibasic only produce polyhydroxyvalarate (PHV) assay by GC and NMR. PHA synthase genes which cloned in B. megaterium transferred to E. coli. Recombinant E. coli use glucose or sodium succinate dibasic as carbon source can produce PHV. The recombinant mcl-PHAs producing bacteria can produce scl-PHAs and mcl-PHAs co-polymers with better properties, but can not use sodium succinate dibasic to produce PHV. Furthermore, some PHAs with brand-new compositions are identified.

並列關鍵字

polyhydroxyalkanoic acids (PHAs), polypropylene (PP)

參考文獻

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