Efficient Expression and Application of a Modified Rhizomucor miehei Lipase for Simultaneous Production of Biodiesel and Eicosapentaenoic Acid Ethyl Ester from Nannochloropsis Oil,Journal of Agricultural and Food Chemistry

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Efficient Expression and Application of a Modified Rhizomucor miehei Lipase for Simultaneous Production of Biodiesel and Eicosapentaenoic Acid Ethyl Ester from Nannochloropsis Oil
Journal of Agricultural and Food Chemistry ( IF 5.7 ) Pub Date : 2024-07-19 , DOI: 10.1021/acs.jafc.4c02360
Tong Lu ₁ , Yifan Qian ₁ , YuQing Zhu ₁ , Xiuyun Ju ₁ , Weiwei Dai ₁ , Quanbin Xu ₁ , Qianqian Yang ₁ , Shuting Li ₁ , Bo Yuan ₁ , Jinjin Huang ₁

Affiliation

Few reports exist on one-step enzymatic methods for the simultaneous production of biodiesel and eicosapentaenoic acid ethyl ester (EPA-EE), a high-value pharmaceutical compound. This study aimed to efficiently express Rhizomucor miehei lipase (pRML) in Pichia pastoris X-33 via propeptide mutation and high-copy strain screening. The mutated enzyme was then used to simultaneously catalyze the production of both biodiesel and EPA-EE. The P46N mutation in the propeptide (P46N-pRML) significantly boosted its production, with the four-copy strain increasing enzyme yield by 3.7-fold, reaching 3425 U/mL. Meanwhile, its optimal temperature increased to 45–50 °C, pH expanded to 7.0–8.0, specific activity doubled, K_m reduced to one-third, and k_cat/K_m increased 7-fold. Notably, P46N-pRML efficiently converts Nannochloropsis gaditana oil’s eicosapentaenoic acid (EPA). Under optimal conditions, it achieves up to 93% biodiesel and 92% EPA-EE yields in 9 h. Our study introduces a novel, efficient one-step green method to produce both biodiesel and EPA-EE using this advanced enzyme.

中文翻译：

改良米赫根毛霉脂肪酶的高效表达及应用，从微拟球藻油中同时生产生物柴油和二十碳五烯酸乙酯

关于同时生产生物柴油和二十碳五烯酸乙酯（EPA-EE）（一种高价值的药物化合物）的一步酶法的报道很少。本研究旨在通过前肽突变和高拷贝菌株筛选，在毕赤酵母X-33 中高效表达根毛霉脂肪酶 (pRML)。然后使用突变酶同时催化生物柴油和 EPA-EE 的生产。前肽 (P46N-pRML) 中的 P46N 突变显着提高了其产量，四拷贝菌株将酶产量提高了 3.7 倍，达到 3425 U/mL。同时，其最适温度提高到45-50℃，pH值扩大到7.0-8.0，比活性增加一倍， K _m降低到三分之一， k _cat / K _m增加7倍。值得注意的是，P46N-pRML 可有效转化微拟球藻油的二十碳五烯酸 (EPA)。在最佳条件下，9 小时内生物柴油产量高达 93%，EPA-EE 产量高达 92%。我们的研究引入了一种新颖、高效的一步绿色方法，使用这种先进的酶生产生物柴油和 EPA-EE。

更新日期：2024-07-19

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