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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 潘子明 | |
dc.contributor.author | Ya-Yun Tan | en |
dc.contributor.author | 譚雅芸 | zh_TW |
dc.date.accessioned | 2021-06-12T18:09:01Z | - |
dc.date.available | 2011-08-26 | |
dc.date.copyright | 2011-08-26 | |
dc.date.issued | 2011 | |
dc.date.submitted | 2011-08-08 | |
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dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/27541 | - |
dc.description.abstract | 紅麴菌屬 (Monascus species) 能經由聚酮 (polyketide) 生成路徑產生具有生理活性的聚酮類二次代謝物;由於紅麴二次代謝相當複雜,目前對於聚酮生合成路徑之調控機制尚未明瞭。M. purpureus BCRC 31615、M. ruber BCRC 31534、M. pilosus BCRC 31526 以及 M. purpureus NTU 568 四種 Monascus 菌株可生產的二次代謝物種類及產量皆不相同,由本研究室所分離出的 M. purpureus NTU 568,在相同培養條件下,其黃色素 (monacin 與 ankaflavin) 產量高於其他菌株,而橘黴素 (citrinin) 產量則是低於 M. purpureus BCRC 31615;M. ruber BCRC 31534 和 M. pilosus BCRC 31526 雖然在 dextrin-MSG-mineral 培養基中不生成黃色素及橘黴素,但 M. ruber BCRC 31534 之紅色素產量顯著高於 M. pilosus BCRC 31526。各菌株 polyketide 產物量之差異,應與代謝途徑酵素調控有關。因此本研究藉由蛋白質體學方法,比較菌株間之蛋白質基礎表現量,藉以了解可能參與調控 polyketides 生成路徑之機制。和 M. purpureus BCRC 31615 相比,M. purpureus NTU 568 細胞中可能與 polyketide synthases (PKSs) 有關之蛋白質 monooxygenase 表現量相對較高,且同為二次代謝路徑之一的 mevalonate 生成路徑較不活絡;分支胺基酸代謝之相關蛋白質以及 aldehyde dehydrogenase (ALDH) 皆在 M. purpureus NTU 568 細胞中有較高之基礎表現量。分支胺基酸降解可產生 malonyl-CoA 或是 butanoyl-CoA,而 ALDH 表現量的上升則可增加 acetate 的產量,malonyl-CoA、butanoyl-CoA 以及 acetate 皆可作為合成 polyketides 的前趨物。當 NTU 568 培養於含有 4% 酒精之培養基時,其分支胺基酸代謝以及 ALDH 之蛋白表現量皆下降,且橘黴素以及黃色素產量被抑制,同時誘導熱衝擊反應 (heat shock response)。另外酒精也會抑制與 polyketides 生成相關之蛋白質 fatty acid synthases (FASs) 及 epoxide hydroxylase (EH) 之表現量,且 shikimate pathway 此二次代謝途徑亦受到抑制。紅色素產量較高之菌株 M. ruber BCRC 31534 和 M. pilosus BCRC 31526 相比,其 pyruvate dehydrogenase complex 表現量相對較高,可能使細胞產生較多的 acetyl-CoA;另外參與甘油代謝之蛋白質 dihydroxyacetone kinase 表現量則是較低,甘油代謝若較不活絡可能使細胞中累積較多甘油,進而增加 propionyl-CoA 的生成,acetyl-CoA 與 propionyl-CoA 皆可做為合成 polyketides 之前趨物。透過上述結果得知紅麴生成 polyketides 產量的高低,除了與本身參與二次代謝路徑之蛋白質表現量有關外,亦可透過改變一次代謝路徑,增加polyketides 之生成前趨物使 polyketides 產量上升。此研究結果對 Monascus 如何調控 polyketides 生成路徑有更進一步瞭解,並且提供增加紅麴 polyketides 之二次代謝物產量有用的資訊。 | zh_TW |
dc.description.abstract | Monascus species produce several potent bioactive metabolites through polyketide secondary metabolic pathways. However, there is little known about the metabolic regulation processes of Monascus. We observed that four strains, M. purpureus BCRC 31615, M. ruber BCRC 31534, M. pilosus BCRC 31526 and M. purpureus NTU 568, have different expression pattern of secondary metabolites inherently. M. purpureus NTU 568, isolated by our research group, could produce higher amount of yellow pigments than other Monascus strains but lower citrinin compared with M. purpureus BCRC 31615. On the other hand, M. ruber BCRC 31534 and M. pilosus BCRC 31526 produced red pigment instead of yellow pigment and citrinin in dextrin-MSG-mineral (DMM) medium cultivation. Furthermore, the production level of red pigment was higher in M. ruber BCRC 31534 compared with M. pilosus BCRC 31526. Assuming the different expression pattern of secondary metabolites might reflect on protein expression level of Monascus species identically, in order to understand the regulatory processes of polyketide pathway in Monascus, we used a comparative proteomic approach to identify proteins with significant expression change among different strains. The protein expression level of monooxygenase, the enzyme that might correlated with polyketide synthases, is higher in M. purpureus NTU 568 compared with M. purpureus BCRC 31615; meanwhile, mevalonate secondary metabolic pathway might be inactive. The proteins involved in branched-chain amino acids degradation, together with aldehyde dehydrogenase (ALDH), were expressed higher level basically in M. purpureus NTU 568. Malonyl-CoA and butanoyl-CoA, synthesized through BCAA degradation, and acetate, produced by ALDH, could be starter units of polyketides. However, after cultured in DMM medium containing 4% EtOH, M. purpureus NTU 568’s BCAA degradation and ALDH expression level were inversely down-regulated, the ability of producing yellow pigment and citrinin was inhibited, and heat shock response was induced as well. Furthermore, both polyketide synthesis related proteins fatty acid synthase and epoxide hydrolase and shikimate secondary metabolic pathway were inhibited by EtOH treatment. M. ruber BCRC 31534, higher red pigment producing strain, has higher expression level of pyruvate dehydrogenase complex compared with M. pilosus BCRC 31526, might resulted in increasing acetyl-CoA pool in cell. Moreover, the decreasing protein level of dihydroxyacetone kinase which participated in glycerol metabolism might result in propionyl-CoA synthesis. These results suggested that higher production of polyketide metabolites result from not only protein expression level of polyketide synthesis pathway, but also elevating starter units from primary metabolism as substrates for polyketide synthesis. This study provides further understanding of the polyketide secondary metabolism of Monascus, together with useful information to improve the production of bioactive secondary metabolites in Monascus species. | en |
dc.description.provenance | Made available in DSpace on 2021-06-12T18:09:01Z (GMT). No. of bitstreams: 1 ntu-100-R98b47104-1.pdf: 4608081 bytes, checksum: bcf1fc35886f6e3e210ac1fbe63cec60 (MD5) Previous issue date: 2011 | en |
dc.description.tableofcontents | 中文摘要 i
Abstract iii 中英對照表 v 目錄 vii 圖目錄 x 表目錄 xii 第一章 緒論 01 第一節 二次代謝 (Secondary metabolism) 01 (一) 二次代謝之定義 01 (二) 二次代謝生成路徑 03 第二節 二次代謝物 06 (一) 真菌之二次代謝系統 06 (二) 植物之二次代謝物 09 第三節 紅麴菌 11 (一) 紅麴菌之菌種特性及分類 11 (二) 紅麴菌的二次代謝產物 13 1. 紅麴色素 13 1.1 色素之代謝生成 13 1.2 紅麴色素之應用 16 1.3 液態培養生成紅麴色素之研究 17 2. 橘黴素 (citrinin) 18 2.1 Citrinin之特性及研究 18 2.2 Citrinin 之代謝合成 21 2.3 Citrinin 於紅麴產品中所產生的問題 21 第四節 氧化壓力 25 (一) 活性氧 25 (二) 抗氧化系統 25 第二章 研究目的及大綱 27 第三章 材料與方法 30 第一節 儀器與設備 30 第二節 藥品 31 第三節 紅麴菌株來源與培養方法 32 第四節 紅麴代謝產物之分析方法 33 第五節 蛋白質抽取方法 34 第六節 等電集焦電泳 36 第七節 第二維電泳分析 36 第八節 蛋白質質譜分析鑑定 37 第四章 結果 39 第一節 紅麴菌種之碳源選擇 39 第二節 不同 Monascus 菌株二次代謝物生成量比較 43 第三節 酒精對 M. purpureus NTU 568 二次代謝物之影響 44 第四節 紅麴菌株蛋白質抽取方法最佳化 47 第五節 不同紅麴菌株之二維蛋白質電泳圖譜比較 49 (一) M. purpureus NTU568 和 M. purpureus BCRC 31615 基礎表現量差異之蛋白質點身分鑑定 54 (二) M. pilosus BCRC 31526 和 M. ruber BCRC 31534 基礎表現量差異之蛋白質點身分鑑定 67 第六節 酒精對 M.purpureus NTU568 之蛋白質影響 72 第五章 討論 90 第一節 M. pupureus NTU 568 代謝路徑之調控 90 第二節 比較 M. pilosus BCRC 31526 及 M. ruber BCRC 31534 代謝之差異 96 第三節 酒精對 M. pupureus NTU 568蛋白質之影響 97 第六章 參考文獻 102 | |
dc.language.iso | zh-TW | |
dc.title | 紅麴菌屬 (Monascus) 聚酮類生成路徑之蛋白質體學研究 | zh_TW |
dc.title | Proteomic insights into the polyketide pathways of Monascus | en |
dc.type | Thesis | |
dc.date.schoolyear | 99-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 蘇遠志,陳水田,黃健雄,李俊霖 | |
dc.subject.keyword | 聚酮類代謝,紅麴屬,橘黴素,黃色素, | zh_TW |
dc.subject.keyword | polyketide pathway,Monascus,citrinin,monascin,ankaflavin, | en |
dc.relation.page | 111 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2011-08-09 | |
dc.contributor.author-college | 生命科學院 | zh_TW |
dc.contributor.author-dept | 生化科技學系 | zh_TW |
顯示於系所單位: | 生化科技學系 |
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