利用巨大芽孢桿菌將豆渣轉化為生物製造 蛋白質原料之綠色循環模組

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洪烱斌(Jiong-Bin Hong) 查詢紙本館藏

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論文名稱

利用巨大芽孢桿菌將豆渣轉化為生物製造蛋白質原料之綠色循環模組
(A green circular module for converting soybean residue into biomass protein raw material using Bacillus megaterium)

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摘要(中)

本研究利用Bacillus megaterium YYBM1，實現豆渣的有效循環利用，以符合聯合國永續發展目標（SDGs）第12項目標—負責任的消費和生產。透過建立一個將豆渣轉化為含豐富胺基酸培養基的可持續循環水解系統，進而達到農業廢棄物的有效轉化和資源回收，同時減少了廢棄物的環境影響。本研究，通過異源表達的TET aminopeptidase在B. megaterium YYBM1細胞中成功表達。確定了最佳的蛋白質表達條件，特別是7小時的最佳誘導時間。值得注意的是，即使在不添加傳統的誘導劑—木糖的情況下，豆渣培養基也能有效地誘導TETamp蛋白的表達，甚至效率高於Luria Bertani商業培養基，突顯豆渣培養基之高營養價值與可商業化應用性。此外，探討鎂濃度和糖蜜添加對蛋白質表達的影響，發現這些因素在本研究設置中對TETamp蛋白的表達量無顯著影響。研究還發現，以5 g/L的細胞濃度進行水解是最低有效的細胞使用量，足以實現豆渣的有效水解。通過該系統，每次水解後能從原始1升培養基中獲得額外53.6%的豆渣水解溶液，提高了資源的循環利用效率。綜上所述，本研究為農業廢棄物的有效處理和再利用提供了一個創新模型，也為全球面臨的廢棄物管理問題提供了實用的解決方案。這一豆渣水解製造蛋白之綠色循環模組的成功開發為微生物技術在促進綠色循環經濟和可持續發展中的應用開闢了新的選擇。

摘要(英)

This study utilizes Bacillus megaterium YYBM1 to achieve the effective recycling of soybean residue, aligning with the United Nations Sustainable Development Goal (SDG) 12 - Responsible Consumption and Production. By establishing a sustainable cyclic hydrolysis system that transforms soybean residue into a nutrient-rich amino acid medium, this approach facilitates the effective conversion and resource recovery of agricultural waste, simultaneously reducing environmental impact. The heterologous expression of TET aminopeptidase was successfully executed within B. megaterium YYBM1 cells. Optimal protein expression conditions were determined, notably a prime induction time of 7 hours. Importantly, even without the addition of the traditional inducer - xylose, the soybean residue medium efficiently induced TETamp protein expression, surpassing the efficiency seen with Luria Bertani commercial medium, thus highlighting the high nutritional value and potential commercial applicability of the soybean residue medium. Furthermore, the study explored the impact of magnesium concentration and molasses addition on protein expression, finding no significant effects on TETamp protein expression within this research setup. It was also found that hydrolyzing with a cell concentration of 5 g/L was the minimum effective dosage for efficient soybean residue hydrolysis. Through this system, each hydrolysis cycle yielded an additional 53.6% of soybean residue hydrolysate from the original 1-liter medium, enhancing resource recycling efficiency. In summary, this research provides an innovative model for the effective treatment and reuse of agricultural waste, offering practical solutions to the global waste management challenge. The successful development of this soybean residue hydrolysis and protein manufacturing green cycle module opens new avenues for the application of microbial technology in promoting a green circular economy and sustainable development.

關鍵字(中)

★ 微生物工程
★ 嗜熱性蛋白酶
★ 農業廢棄物再利用
★ 豆渣水解
★ 循環經濟
★ 資源回收

關鍵字(英)

★ Microbial Engineering
★ Thermophilic Protease
★ Agricultural Waste Reutilization
★ Soybean Residue Hydrolysis
★ Circular Economy
★ resource recovery

論文目次

摘要 i
Abstract ii
誌謝 iv
目錄 v
圖目錄 viii
表目錄 ix
符號說明 x
第一章前言 1
1.1 研究背景 1
1.2 研究目的 2
第二章文獻回顧 4
2.1 聯合國永續發展目標 4
2.2 5+2產業創新政策 6
2.3 豆渣 7
2.4 細菌細胞結構 10
2.5 巨大芽孢桿菌 (Bacillus megaterium) 11
2.6 豆渣培養基 12
2.7 微生物生長培養基 13
2.7.1 碳源 13
2.7.2 金屬離子 13
2.8 蛋白質水解 14
2.8.1 化學水解 14
2.8.2 酵素水解 15
2.9 酵素 16
2.9.1 蛋白酶 19
2.9.2 Alcalase內肽酶 22
2.9.3 TET aminopeptidase 22
第三章材料與方法 24
3.1 實驗架構 24
3.2 實驗材料 25
3.2.1 實驗藥品 25
3.2.2 實驗設備 26
3.3 菌種保存及培養 27
3.3.1菌種保存 27
3.3.2 菌種培養 27
3.4 微生物操作 28
3.4.1 DNA操作 (DNA manipulation) 29
3.4.2 分子克隆 (Molecular cloning) 30
3.4.3 轉形作用 (Transformation) 30
3.5 蛋白質實驗分析 31
3.5.1 目標蛋白表達 31
3.5.2 目標蛋白純化 31
3.5.3 目標蛋白定性 32
3.6 豆渣培養基製作 33
3.7 最佳生長條件實驗分析 34
3.7.1 不同誘導時間 34
3.7.2 不同鎂濃度 34
3.7.3 不同糖蜜濃度 34
3.7.4 不同細胞重量 35
3.8 特性分析 35
3.8.1 細胞活性分析 35
3.8.2 蛋白質定量 35
3.8.3 胺基酸定量 37
第四章結果與討論 39
4.1 異源蛋白表達 39
4.2 水解豆渣 41
4.3 最佳條件TETamp異源蛋白比較 42
4.3.1 誘導時間 42
4.3.2 鎂濃度 46
4.3.3 糖蜜 47
4.4 循環模組建立 49
4.4.1 豆渣培養基蛋白表達 49
4.4.2 細胞水解豆渣分析 50
4.4.3 細胞產量 51
4.4.4 永續水解豆渣培養基 52
結論與建議 54
5.1 結論 54
5.2 建議 55
參考文獻 56

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指導教授

王柏翔(Po-Hsiang Wang)

審核日期

2024-3-19

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