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Improved production of docosahexaenoic acid in batch fermentation by newly-isolated strains of Schizochytrium sp. and Thraustochytriidae sp. through bioprocess optimization.

ABSTRACT: Thraustochytrids, rich in docosahexaenoic acid (DHA, C22:6?3), represent a potential source of dietary fatty acids. Yet, the effect of culture conditions on growth and fatty acid composition vary widely among different thraustochytrid strains. Two different thraustochytrid strains, Schizochytrium sp. PKU#Mn4 and Thraustochytriidae sp. PKU#Mn16 were studied for their growth and DHA production characteristics under various culture conditions. Although they exhibited similar fatty acid profiles, PKU#Mn4 seemed a good candidate for industrial DHA fermentation while PKU#Mn16 displayed growth tolerance to a wide range of process conditions. Relative DHA content of 48.5% and 49.2% (relative to total fatty acids), respectively, were achieved on glycerol under their optimal flask culture conditions. Maximum DHA yield (Yp/x) of 21.0% and 18.9% and productivity of 27.6 mg/L-h and 31.9 mg/L-h were obtained, respectively, in 5-L bioreactor fermentation operated with optimal conditions and dual oxygen control strategy. A 3.4- and 2.8-fold improvement of DHA production (g/L), respectively, was achieved in this study. Overall, our study provides the potential of two thraustochytrid strains and their culture conditions for efficient production of DHA-rich oil.

SUBMITTER: Wang Q

PROVIDER: S-EPMC5995480 | biostudies-literature | 2018 Jun

REPOSITORIES: biostudies-literature

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Improved production of docosahexaenoic acid in batch fermentation by newly-isolated strains of Schizochytrium sp. and Thraustochytriidae sp. through bioprocess optimization.

Wang Qiuzhen Q Ye Huike H Sen Biswarup B Xie Yunxuan Y He Yaodong Y Park Sunghoon S Wang Guangyi G

Synthetic and systems biotechnology 20180421 2

Thraustochytrids, rich in docosahexaenoic acid (DHA, C22:6ω3), represent a potential source of dietary fatty acids. Yet, the effect of culture conditions on growth and fatty acid composition vary widely among different thraustochytrid strains. Two different thraustochytrid strains, Schizochytrium sp. PKU#Mn4 and Thraustochytriidae sp. PKU#Mn16 were studied for their growth and DHA production characteristics under various culture conditions. Although they exhibited similar fatty aci ...[more]

PMID: 29900425

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Project description:Background:Schizochytrium species are known for their abundant production of docosahexaenoic acid (DHA). Low temperatures can promote the biosynthesis of polyunsaturated fatty acids (PUFAs) in many species. This study investigates low-temperature effects on DHA biosynthesis in Schizochytrium sp. TIO01 and its underlying mechanism. Results:The Schizochytrium fatty acid biosynthesis pathway was evaluated based on de novo genome assembly (contig N50?=?2.86 Mb) and iTRAQ-based protein identification. Our findings revealed that desaturases, involved in DHA synthesis via the fatty acid synthase (FAS) pathway, were completely absent. The polyketide synthase (PKS) pathway and the FAS pathway are, respectively, responsible for DHA and saturated fatty acid synthesis in Schizochytrium. Analysis of fatty acid composition profiles indicates that low temperature has a significant impact on the production of DHA in Schizochytrium, increasing the DHA content from 43 to 65% of total fatty acids. However, the expression levels of PKS pathway genes were not significantly regulated as the DHA content increased. Further, gene expression analysis showed that pathways related to the production of substrates (acetyl-CoA and NADPH) for fatty acid synthesis (the branched-chain amino acid degradation pathway and the pentose phosphate pathway) and genes related to saturated fatty acid biosynthesis (the FAS pathway genes and malic enzyme) were, respectively, upregulated and downregulated. These results indicate that low temperatures increase the DHA content by likely promoting the entry of relatively large amounts of substrates into the PKS pathway. Conclusions:In this study, we provide genomic, proteomic, and transcriptomic evidence for the fatty acid synthesis pathway in Schizochytrium and propose a mechanism by which low temperatures promote the accumulation of DHA in Schizochytrium. The high-quality and nearly complete genome sequence of Schizochytrium provides a valuable reference for investigating the regulation of polyunsaturated fatty acid biosynthesis and the evolutionary characteristics of Thraustochytriidae species.

Project description:Background:Schizochytrium sp. is a promising strain for the production of docosahexaenoic acid (DHA)-rich oil and biodiesel, and has been widely used in the food additive and bioenergy industries. Oxygen is a particularly important environmental factor for cell growth and DHA synthesis. In general, higher oxygen supply favors lipid accumulation, but could lead to a reduction of the DHA percentage in total fatty acids in Schizochytrium sp. To tackle this problem, it is essential to understand the mechanisms regulating the response of Schizochytrium sp. to oxygen. In this study, we aimed to explore the acclimatization of this DHA producer to different oxygen supply conditions by examining the transcriptome changes. Results:Two different fermentation processes, namely normal oxygen supply condition (shift agitation speeds from 400 rpm to 300 rpm) and high oxygen supply condition (constant agitation speeds: 400 rpm), were designed to study how the fermentation characteristics of Schizochytrium sp. HX-308 were affected by different oxygen supply conditions. The results indicated that high oxygen supply condition resulted in 49% and 37.5% improvement in the maximum cell dry weight (CDW) and total lipid concentration, respectively. However, the DHA percentage in total fatty acids decreased to 35%, which was 31.4% lower than that produced by normal oxygen supply condition. Moreover, transcriptome analysis was performed to explore the effect of the oxygen supply condition on genetic expression and metabolism. The results showed that glycolysis and pentose phosphate pathway metabolism-associated genes (hexokinase, phosphofructokinase, fructose-bisphosphate aldolase, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase) were substantially upregulated in response to high oxygen supply, resulting in more NADPH was available for Schizochytrium. Specially, high oxygen supply condition also led to genes (?6 desaturase, ?12 desaturase, FAS, ORFA, ORFB, and ORFC) involved in fatty acid biosynthesis upregulation. In addition, a transcriptional upregulation of catalase (CAT) became apparent under high oxygen supply condition, while superoxide dismutase (SOD) and ascorbate peroxidase (APX) were found to be down-regulated. Conclusions:This study is the first to investigate the differences of gene expression at different levels of oxygen availability in the DHA producer Schizochytrium. The results of transcriptome analyses indicated that high oxygen supply condition resulting in more NADPH and acetyl-CoA production for cell growth and lipid synthesis in Schizochytrium. ?12 desaturase and ORFC showed higher expression levels at high oxygen supply condition, which might be the key regulators for enhancing fatty acid biosynthesis in the future. These results enrich the current knowledge regarding genetic expression and provide important information to enhance DHA production in Schizochytrium sp.

Project description:BackgroundDocosahexaenoic acid (DHA, C22:6) and odd-chain fatty acids (OCFAs, C15:0 and C17:0) have attracted great interest, since they have been widely used in food and therapeutic industries, as well as chemical industry, such as biodiesel production and improvement. The oil-producing heterotrophic microalgae Schizochytrium sp. 31 is one of main DHA-producing strains. Recently, it was found that Schizochytrium can also synthesize OCFAs; however, contents and titers of DHA and OCFAs in Schizochytrium are still low, which limit its practical application.ResultsIn this study, we found that acetyl-CoA carboxylase suffered from a feedback inhibition by C16-CoA in Schizochytrium, and relief of the inhibition resulted in improved both lipid content and the ratio of OCFAs in total fatty acids. Based on this finding, a novel strategy for elevating both DHA and OCFAs contents was established. First, the total lipid accumulation was increased by overexpressing a malic enzyme from Crypthecodinium cohnii to elevate NADPH supply. Second, the inhibition effect on acetyl-CoA carboxylase was relieved by overexpressing a codon-optimized ELO3 gene from Mortierella alpina, which encodes an elongase enzyme responsible for converting C16 into C18 fatty acids. After the above two-step engineering, contents of DHA and OCFAs were increased by 1.39- and 3.30-fold, reaching a level of 26.70 and 25.08% of dry cell weight, respectively, which are the highest contents reported so far for Schizochytrium. The titers of DHA and OCFAs were elevated by 1.08- and 2.57-fold, reaching a level of 3.54 and 3.32 g/L, respectively. Notably, the OCFAs titer achieved was 2.66-fold higher than the highest reported in Escherichia coli (1.25 g/L), implying potential value for industry application. To reveal the potential metabolic mechanism for the enhanced biosynthesis of both DHA and OCFAs, LC-MS metabolomic analysis was employed and the results showed that the pentose phosphate pathway and the glycolysis pathway were strengthened and intracellular propionyl-CoA concentration were also significantly increased in the engineered Schizochytrium, suggesting an increased supply of NADPH, acetyl-CoA, and propionyl-CoA for DHA and OCFAs accumulation.ConclusionsThe discovery provides a new source of OCFAs production, and proposes a new strategy to improve contents and titers of both DHA and OCFAs in Schizochytrium. These will be valuable for improving commercial potential of Schizochytrium and guiding the engineering strategy in other fatty acids producing heterotrophic microalga.

Dataset Information

Improved production of docosahexaenoic acid in batch fermentation by newly-isolated strains of Schizochytrium sp. and Thraustochytriidae sp. through bioprocess optimization.

Publications

Improved production of docosahexaenoic acid in batch fermentation by newly-isolated strains of <i>Schizochytrium</i> sp. and <i>Thraustochytriidae</i> sp. through bioprocess optimization.

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