Bioproduction of Linalool From Paper Mill Waste.
Bioproduction of Linalool From Paper Mill Waste.
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DOI：
10.3389/fbioe.2022.892896
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发表时间：
2022
影响因子：
5.7
通讯作者：
Scrutton, Nigel S.
中科院分区：
文献类型：
--
作者：
Rinaldi, Mauro A.;Tait, Shirley;Toogood, Helen S.;Scrutton, Nigel S.
关键词：
化学品生物制造的一个关键挑战是保持稳定、高产的微生物菌株，以实现大规模的成本效益发酵。微生物工程的“一刀切”方法通常用于优化宿主稳定性和生产力。这可能涉及确定菌株特征的潜在限制，然后尝试通过靶向工程化系统地优化生产菌株。然而，这种有针对性的方法并不总是导致所需的性状。在这里，我们展示了两个'命中和错过'的结果，有针对性的方法，试图产生一个稳定的大肠杆菌菌株的生物生产的单萜芳樟醇，香料分子的工业利益。首先，我们通过消除编码芳樟醇生产途径的质粒构建体中负责切除途径组分的重复序列来稳定芳樟醇生产菌株。然后将这些优化的途径构建体整合到E.大肠杆菌中的三个部分，以消除对抗生素的需要，以维持芳樟醇生产。还采用了其他策略，包括：通过适应性实验室进化和修饰或关键瓶颈酶GPPS/林斯的同源基因替换来降低芳樟醇的细胞毒性。我们的研究表明，影响芳樟醇滴度的主要因素是E。大肠杆菌的稳定性是遗传构建体对切除或类似重组事件的稳定性。其他因素，如降低芳樟醇细胞毒性和改变途径基因，没有导致稳定性或滴度的改善。为了降低规模化发酵成本，还研究了使用含有造纸废水二次纸纤维作为唯一碳源的最小基础培养基。这涉及同时糖化和发酵使用补充纤维素酶混合物或通过共表达分泌型纤维素酶在E。含有稳定的芳樟醇生产途径的大肠杆菌。结合起来，这项研究已经证明了一种稳定的芳樟醇生产方法，使用丰富和低成本的原料和改进的生产菌株，为从造纸厂废物流中生产化学品提供了重要的概念验证。对于规模化生产，需要使用更全面的方法进行优化，这些方法涉及进一步的微生物工程和发酵工艺开发。
A key challenge in chemicals biomanufacturing is the maintenance of stable, highly productive microbial strains to enable cost-effective fermentation at scale. A “cookie-cutter” approach to microbial engineering is often used to optimize host stability and productivity. This can involve identifying potential limitations in strain characteristics followed by attempts to systematically optimize production strains by targeted engineering. Such targeted approaches however do not always lead to the desired traits. Here, we demonstrate both ‘hit and miss’ outcomes of targeted approaches in attempts to generate a stable Escherichia coli strain for the bioproduction of the monoterpenoid linalool, a fragrance molecule of industrial interest. First, we stabilized linalool production strains by eliminating repetitive sequences responsible for excision of pathway components in plasmid constructs that encode the pathway for linalool production. These optimized pathway constructs were then integrated within the genome of E. coli in three parts to eliminate a need for antibiotics to maintain linalool production. Additional strategies were also employed including: reduction in cytotoxicity of linalool by adaptive laboratory evolution and modification or homologous gene replacement of key bottleneck enzymes GPPS/LinS. Our study highlights that a major factor influencing linalool titres in E. coli is the stability of the genetic construct against excision or similar recombination events. Other factors, such as decreasing linalool cytotoxicity and changing pathway genes, did not lead to improvements in the stability or titres obtained. With the objective of reducing fermentation costs at scale, the use of minimal base medium containing paper mill wastewater secondary paper fiber as sole carbon source was also investigated. This involved simultaneous saccharification and fermentation using either supplemental cellulase blends or by co-expressing secretable cellulases in E. coli containing the stabilized linalool production pathway. Combined, this study has demonstrated a stable method for linalool production using an abundant and low-cost feedstock and improved production strains, providing an important proof-of-concept for chemicals production from paper mill waste streams. For scaled production, optimization will be required, using more holistic approaches that involve further rounds of microbial engineering and fermentation process development.
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影响因子：
3.5
作者：
CHEREPANOV, PP;WACKERNAGEL, W
通讯作者：
WACKERNAGEL, W
通过质粒重组消除和途径优化提高蒎烯的生物合成产量。
影响因子：
2.6
作者：
Bao, Shao-Heng;Zhang, Dong-Yi;Meng, Er
通讯作者：
Meng, Er
影响因子：
11.4
作者：
Elliston, Adam;Wilson, David R.;Waldron, Keith W.
通讯作者：
Waldron, Keith W.
影响因子：
3.8
作者：
Alonso-Gutierrez, Jorge;Koma, Daisuke;Lee, Taek S.
通讯作者：
Lee, Taek S.
DOI：
10.1073/pnas.120163297
发表时间：
2000-06-06
影响因子：
11.1
作者：
Datsenko, KA;Wanner, BL
通讯作者：
Wanner, BL