細菌血紅素影響聚羥基烷酯產量之研究

The effect of Vitreoscilla stercoraria hemoglobin expression on polyhydroxyalkanoates (PHA) production

楊名翔(Ming-Hsiang Yang)

指導教授 ： 蘇伯琦

元智大學/工程學院/生物科技暨生物資訊研究所/碩士(2008年)

摘要

聚羥基烷酯(PHAs)屬於天然的酯類，當細菌在環境中生長過程中，因碳源過剩且其他營養源（如氮源、磷源等）缺乏時，細菌將會利用多餘的碳源代謝累積而成PHAs。PHAs的性質與傳統塑膠相似並具有生物可分解性和生物相容性。本研究中利用基因重組技術將Ralstonia eutropha的 PHAs合成基因phbCAB與Vitreoscilla stercoraria的細菌血紅素合成基因vgb (Vitreoscilla globin gene)作起動子之改造與表現，以提高在大腸桿菌之細胞生長量與PHB產量。根據實驗結果顯示所構築之表現質體pSY02含有phbCAB與vgb基因且重組於可受arabinose調控之PBAD啟動子之後，所篩選到可大量表現PHB之菌株，其PHB 含量和PHB產量分別可達47 % 和2.12 g/L 。此外，為了將細菌血紅素應用於其他代謝工程，本研究中成功地建構了數個可在格蘭氏陰性或陽性菌株內表現細菌血紅素的質體，其細菌血紅素表現能力均經由專一性抗體與西方墨點法確認，確實可以在相關細菌中表現。

關鍵字

細菌血紅素 ； 聚羥基烷酯

並列摘要

Polyhydroxyalkanoates (PHAs) are natural polyesters that could be accumulated in many bacteria when they are grown in the medium containing excess carbon source and other limited nutrient (such as nitrogen or phosphate source). The properties of PHA are not only similar to traditional plastics but also biodegradable and biocompatible. It has been reported that PHAs synthesis genes (phbCAB) from Ralstonia eutropha produce PHB in Escherichia coli. In this study, phbCAB and vgb, hemoglobin gene from Vitreoscilla stercoraria, were coexpressed and controlled by PBAD promoter to improve cell growth and enhance PHB production. The results showed that the PHB content and PHB yield were 47 % and 2.12 g/L respectively in the recombinant strain harboring the phbCAB and vgb. The second topic of this study is that construction of vgb in expressed plasmids which could express VHb in Gram negative or Gram positive strains. The expressions of VHb were confirmed by Western blotting through the specific antibody. These constructions could be applied in broad metabolic engineering.

並列關鍵字

Vitreoscilla stercoraria hemoglobin(VHb) ； polyhydroxyalkanoates (PHA)

參考文獻

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