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Purification and immobilization of engineered glucose dehydrogenase: a new approach to producing gluconic acid from breadwaste.
Biotechnology for Biofuels ( IF 6.3 ) Pub Date : 2020-06-03 , DOI: 10.1186/s13068-020-01735-7 Pinar Karagoz 1 , Ravneet Mandair 1 , Jinesh Cherukkattu Manayil 2 , Jai Lad 2 , Katie Chong 2 , Georgios Kyriakou 3 , Adam F Lee 4 , Karen Wilson 4 , Roslyn M Bill 1
Biotechnology for Biofuels ( IF 6.3 ) Pub Date : 2020-06-03 , DOI: 10.1186/s13068-020-01735-7 Pinar Karagoz 1 , Ravneet Mandair 1 , Jinesh Cherukkattu Manayil 2 , Jai Lad 2 , Katie Chong 2 , Georgios Kyriakou 3 , Adam F Lee 4 , Karen Wilson 4 , Roslyn M Bill 1
Affiliation
Platform chemicals are essential to industrial processes. Used as starting materials for the manufacture of diverse products, their cheap availability and efficient sourcing are an industrial requirement. Increasing concerns about the depletion of natural resources and growing environmental consciousness have led to a focus on the economics and ecological viability of bio-based platform chemical production. Contemporary approaches include the use of immobilized enzymes that can be harnessed to produce high-value chemicals from waste. In this study, an engineered glucose dehydrogenase (GDH) was optimized for gluconic acid (GA) production. Sulfolobus solfataricus GDH was expressed in Escherichia coli. The Km and Vmax values for recombinant GDH were calculated as 0.87 mM and 5.91 U/mg, respectively. Recombinant GDH was immobilized on a hierarchically porous silica support (MM-SBA-15) and its activity was compared with GDH immobilized on three commercially available supports. MM-SBA-15 showed significantly higher immobilization efficiency (> 98%) than the commercial supports. After 5 cycles, GDH activity was at least 14% greater than the remaining activity on commercial supports. Glucose in bread waste hydrolysate was converted to GA by free-state and immobilized GDH. After the 10th reuse cycle on MM-SBA-15, a 22% conversion yield was observed, generating 25 gGA/gGDH. The highest GA production efficiency was 47 gGA/gGDH using free-state GDH. This study demonstrates the feasibility of enzymatically converting BWH to GA: immobilizing GDH on MM-SBA-15 renders the enzyme more stable and permits its multiple reuse.
中文翻译:
工程葡萄糖脱氢酶的纯化和固定化:一种从面包屑中生产葡萄糖酸的新方法。
平台化学品对工业流程至关重要。用作制造各种产品的起始材料,它们的廉价可用性和高效采购是工业要求。对自然资源枯竭的日益关注和日益增强的环境意识导致人们关注基于生物的平台化学生产的经济和生态可行性。当代方法包括使用可用于从废物中生产高价值化学品的固定化酶。在这项研究中,工程葡萄糖脱氢酶 (GDH) 针对葡萄糖酸 (GA) 生产进行了优化。Sulfolobus solfataricus GDH 在大肠杆菌中表达。重组 GDH 的 Km 和 Vmax 值分别计算为 0.87 mM 和 5.91 U/mg。将重组 GDH 固定在分级多孔二氧化硅载体 (MM-SBA-15) 上,并将其活性与固定在三种市售载体上的 GDH 进行比较。MM-SBA-15 显示出比商业载体更高的固定效率 (> 98%)。5 个循环后,GDH 活性至少比商业载体上的剩余活性高 14%。面包废料水解物中的葡萄糖通过游离态和固定化的 GDH 转化为 GA。在 MM-SBA-15 的第 10 个重复使用周期后,观察到 22% 的转化率,产生 25 gGA/gGDH。使用自由态 GDH 的最高 GA 生产效率为 47 gGA/gGDH。这项研究证明了将 BWH 酶促转化为 GA 的可行性:将 GDH 固定在 MM-SBA-15 上使酶更稳定并允许其多次重复使用。
更新日期:2020-06-03
中文翻译:
工程葡萄糖脱氢酶的纯化和固定化:一种从面包屑中生产葡萄糖酸的新方法。
平台化学品对工业流程至关重要。用作制造各种产品的起始材料,它们的廉价可用性和高效采购是工业要求。对自然资源枯竭的日益关注和日益增强的环境意识导致人们关注基于生物的平台化学生产的经济和生态可行性。当代方法包括使用可用于从废物中生产高价值化学品的固定化酶。在这项研究中,工程葡萄糖脱氢酶 (GDH) 针对葡萄糖酸 (GA) 生产进行了优化。Sulfolobus solfataricus GDH 在大肠杆菌中表达。重组 GDH 的 Km 和 Vmax 值分别计算为 0.87 mM 和 5.91 U/mg。将重组 GDH 固定在分级多孔二氧化硅载体 (MM-SBA-15) 上,并将其活性与固定在三种市售载体上的 GDH 进行比较。MM-SBA-15 显示出比商业载体更高的固定效率 (> 98%)。5 个循环后,GDH 活性至少比商业载体上的剩余活性高 14%。面包废料水解物中的葡萄糖通过游离态和固定化的 GDH 转化为 GA。在 MM-SBA-15 的第 10 个重复使用周期后,观察到 22% 的转化率,产生 25 gGA/gGDH。使用自由态 GDH 的最高 GA 生产效率为 47 gGA/gGDH。这项研究证明了将 BWH 酶促转化为 GA 的可行性:将 GDH 固定在 MM-SBA-15 上使酶更稳定并允许其多次重复使用。