JOURNAL OF THE SCIENCE OF FOOD AND AGRICULTURE
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2024
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IF 3.3
Buqing Wang,Yasen Wang,Xiaoman Zhou,Xiao-Dong Gao,Morihisa Fujita,Zijie Li
BACKGROUND Acidic lipases with high catalytic activities under acidic conditions have important application values in the food, feed and pharmaceutical industries. However, the availability of acidic lipases is still the main obstacle to their industrial applications. Although a novel acidic lipase Rasamsonia emersonii (LIPR) was heterologously expressed in Escherichia coli , the expression level was unsatisfactory. RESULTS To achieve the high-efficiency expression and secretion of LIPR in Pichia pastoris GS115, the combinatorial optimization strategy was adopted including gene codon preference, signal peptide, molecular chaperone co-expression and disruption of vacuolar sorting receptor VPS10 . The activity of the combinatorial optimization engineered strain in a shake flask reached 1480 U mL −1 , which was 8.13 times greater than the P. pastoris GS115 parental strain. After high-density fermentation in a 5-L bioreactor, the highest enzyme activity reached as high as 11 820 U mL −1 . LIPR showed the highest activity at 40 °C and pH 4.0 in the presence of Ca 2+ ion. LIPR exhibited strong tolerance to methanol, indicating its potential application in biodiesel biosynthesis. Moreover, the gastrointestinal digestion simulation results demonstrated that LIPR was tolerant to pepsin and trypsin, but its activity was inhibited by sodium taurodeoxycholate. CONCLUSION This study provided an effective approach for the high expression of acidic lipase LIPR. LIPR was more appropriate for lipid digestion in the stomach than in intestine according to the gastrointestinal digestion simulation results. © 2024 Society of Chemical Industry.
研究领域:
生物技术
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酶工程
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食品科学
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DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
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2024
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IF 7.7
Buqing Wang,Xiaoman Zhou,Yasen Wang,Yahui Gao,Hideki Nakanishi,Morihisa Fujita,Zijie Li
The acidic lipase from Rasamsonia emersonii named LIPR has great potential for biodiesel synthesis due to its strong methanol tolerance. Nonetheless, the limited thermostability of LIPR and low expression level in Escherichia coli remain major obstacles to its use in biodiesel synthesis. To enhance the thermostability, the mutant LIPR harboring mutations A126C-P238C for the formation of a new disulfide bond and amino acid substitution D214L was obtained through rational design. To our delight, the thermostability of LIPR mutant was greatly improved. Moreover, a comprehensive optimization strategy, such as employing the Mss signal peptide, co-expressing the molecular chaperone protein disulfide isomerase (PDI), knocking out the vacuolar sorting receptor gene VPS10-01 , and overexpressing the dihydroxyacetone synthase gene DAS2 , was adopted to obtain the combination-optimized mutant Pichia pastoris strain GS54. Furthermore, the biodiesel synthetic capability with the mutant GS54-LIPR was verified and the production yield was 52.2 % after 24 h in a shake flask. Subsequently, a continuous flow system was adopted to increase the biodiesel yield to 73.6 % within 3 h, demonstrating its efficacy in enhancing enzyme biocatalysis. The engineered GS54-LIPR mutant lipase is an efficient and reusable biocatalyst for the sustained production of biodiesel in a continuous flow reaction.
研究领域:
生物化学
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分子生物学
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工业微生物学
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DOI
PubMed
高影响因子
Food Chemistry-X
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2024
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IF 6.5
Tiehan Li,Yuming Wei,Mingxia Lu,Yida Wu,Yanqun Jiang,Han Ke,Aiju Shao,Jingming Ning
Piling fermentation (PF) is crucial for Pu-erh tea aroma, yet its microbial and moist-heat impact on aroma quality is poorly understood. Solid-phase microextraction, solvent-assisted flavor evaporation, and gas chromatography–mass spectrometry were used to detected and analyses the samples of sun-green green tea, sterile PF and spontaneous PF. Microbiological action promotes the formation of stale aromas. Moist-heat action promotes the formation of plum-fragrance and sweet aroma. 20 microbial markers and 28 moist-heat markers were screened from 184 volatile components. Combining odor activity values and gas chromatography-olfactometry, 22 aroma-active compounds were screened (1,2,3-trimethoxybenzene, linalool, 1,2,4-trimethoxybenzene …), and analyzed during PF processing. Aroma omission and addition experiments verified its importance. Gallic acid addition experiments successfully verified that microorganisms are the main contributors to the synthesis of methoxybenzenes. Finally, Blastobotrys, Rasamsonia , and Thermomyces showed positive correlation with the synthesis of 1-ethyl-4-methoxybenzene, 1,2,4-trimethoxybenzene, 1,2,3-trimethoxybenzene, and 1,2-dimethoxybenzene. The formation mechanism of Pu-erh tea's aroma was clarified. Exploring microbial and moist-heat effects on Pu-erh tea volatiles and understanding the methoxybenzene formation mechanism using molecular sensory science.
研究领域:
食品科学
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微生物学
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感官科学
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化学
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产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）