Astaxanthin Production Research Articles

Production and excretion of astaxanthin by engineered Yarrowia lipolytica using plant oil as both the carbon source and the biocompatible extractant.

This study aimed to develop a bioprocess using plant oil as the carbon source for lipid-assimilating yeast to produce high-value astaxanthin. Using high-oleic safflower oil as a model, efficient cell growth and astaxanthin production by the engineered Yarrowia lipolytica strain ST7403 was demonstrated, and a considerable portion of astaxanthin was found excreted into the spent oil. Astaxanthin was the predominant carotenoid in the extracellular oil phase that allowed facile in situ recovery of astaxanthin without cell lysis. Autoclaving the safflower oil medium elevated the peroxide level but it declined quickly during fermentation (reduced by 84% by day 3) and did not inhibit cell growth or astaxanthin production. In a 1.5-L fed-batch bioreactor culture with a YnB-based medium containing 20% safflower oil, and with the feeding of casamino acids, astaxanthin production reached 54mg/L (53% excreted) in 28days. Further improvement in astaxanthin titer and productivity was achieved by restoring leucine biosynthesis in the host, and running fed-batch fermentation using a high carbon-to-nitrogen ratio yeast extract/peptone medium containing 70% safflower oil, with feeding of additional yeast extract/peptone, to attain 167mg/L astaxanthin (48% excreted) in 9.5days of culture. These findings facilitate industrial microbial biorefinery development that utilizes renewable lipids as feedstocks to not only produce high-value products but also effectively extract and recover the products, including non-native ones.Key Points• Yarrowia lipolytica can use plant oil as a C-source for astaxanthin production.• Astaxanthin is excreted and accumulated in the extracellular oil phase.• Astaxanthin is the predominant carotenoid in the extracellular oil phase.• Plant oil serves as a biocompatible solvent for in situ astaxanthin extraction. Graphical abstract.

Enhanced Biomass and Astaxanthin Production of Haematococcus pluvialis by a Cell Transformation Strategy with Optimized Initial Biomass Density.

Astaxanthin from H. pluvialis is an antioxidant and presents a promising application in medicine for human health. The two-stage strategy has been widely adopted to produce astaxanthin by the Haematococcus industry and research community. However, cell death and low astaxanthin productivity have seriously affected the stability of astaxanthin production. This study aims to test the effect of cell transformation strategies on the production of astaxanthin from H. pluvialis and determine the optimal initial biomass density (IBD) in the red stage. The experimental design is divided into two parts, one is the vegetative growth experiment and the other is the stress experiment. The results indicated that: (1) the cell transformation strategy of H. pluvialis can effectively reduce cell death occurred in the red stage and significantly increase the biomass and astaxanthin production. (2) Compared with the control group, the cell mortality rate of the red stage in the treatment group was reduced by up to 81.6%, and the biomass and astaxanthin production was increased by 1.63 times and 2.1 times, respectively. (3) The optimal IBD was determined to be 0.5, and the highest astaxanthin content can reach 38.02 ± 2.40 mg·g−1. Thus, this work sought to give useful information that will lead to an improved understanding of the cost-effective method of cultivation of H. pluvialis for natural astaxanthin. This will be profitable for algal and medicine industry players.

Preparation of astaxanthin by lipase-catalyzed hydrolysis from its esters in a slug-flow microchannel reactor

Natural astaxanthin is widely used as a food and cosmetics additive because of its multiple biological activities. However, astaxanthin produced by Haematococcus pluvialis is generally esterified, and its activity is far less than that of free astaxanthin. Hydrolysis of astaxanthin esters to free astaxanthin by enzymes can overcome the drawbacks of chemical saponification methods. In this paper, a slug-flow microchannel reactor was constructed and tested in enzymatic hydrolysis of astaxanthin esters. The reactor consists of a “T” slug-flow generator, a stainless-steel microchannel, two constant-flow pumps, and a temperature controller. The reactor has the advantages of simple configuration and easy scale-up, and is suitable for two-phase biochemical reactions. Using the microchannel reactor, astaxanthin esters in H. pluvialis oil were efficiently hydrolyzed to free astaxanthin by lipase from Aspergillus niger. After hydrolysis, the content of free astaxanthin in H. pluvialis oil was 18.8 mg/L, 7.83-times higher than that before hydrolysis (2.13 mg/L). The hydrolysis rate reached 75.4 %. These results indicate that the microchannel reactor can be useful for the production of free astaxanthin from its esters.

High light boosts salinity stress-induced biosynthesis of astaxanthin and lipids in the green alga Chromochloris zofingiensis

Chromochloris zofingiensis, an astaxanthin-producing oleaginous green alga, is emerging as a model for studying carotenogenesis and lipogenesis. While the stimulated biosynthesis of secondary carotenoids and storage lipids by abiotic stresses has long been perceived in C. zofingiensis, the underlying mechanisms particularly under combinatorial stresses remain to be explored. In the present study, the synthesis of carotenoids and lipids was comparatively investigated in C. zofingiensis under high light (HL), salinity stress (SS) and the combinatorial HL and SS (HL + SS) conditions. While HL or SS alone enhanced astaxanthin and lipids moderately, HL + SS had synergistic effect and promoted these compounds considerably. HL + SS also gave rise to the highest productivities. Carotenoid profiling and transcriptional analysis supported the diversion of carotenoid flux from primary carotenoids to secondary carotenoids particularly astaxanthin and violaxanthin cycle that produces zeaxanthin likely contributed to astaxanthin synthesis. Lipid profiling and transcriptional analysis, on the other hand, suggested that both de novo fatty acid synthesis and membrane lipid turnover contributed fatty acyls for supporting triacylglycerol assembly via the acyl CoA-dependent and acyl CoA-independent routes. Astaxanthin and triacylglycerol that share common carbon precursors were induced to synthesize in a coordinated way regulated at the transcriptional level, yet the carbon flux was allocated predominantly to the latter. Taken together, our results demonstrated the beneficial effect of combinatorial stresses on both astaxanthin and triacylglycerol production, identified critical genes involved in carotenogenesis and lipogenesis, and provided new insights into algal astaxanthin biosynthesis.

Enhancing astaxanthin accumulation in Xanthophyllomyces dendrorhous by a phytohormone: metabolomic and gene expression profiles.

Summary Xanthophyllomyces dendrorhous is a promising source of natural astaxanthin due to its ability to accumulate high amounts of astaxanthin. This study showed that 6‐benzylaminopurine (6‐BAP) is an effective substrate that enhances cell biomass and astaxanthin accumulation in X. dendrorhous. In the current study, the biomass and astaxanthin content in X. dendrorhous were determined to be improved by 21.98% and 24.20%, respectively, induced by 6‐BAP treatments. To further understand the metabolic responses of X. dendrorhous to 6‐BAP, time‐course metabolomics and gene expression levels of X. dendrorhous cultures with and without 6‐BAP feeding were investigated. Metabolome analysis revealed that 6‐BAP facilitated glucose consumption, promoted the glycolysis, suppressed the TCA cycle, drove carbon flux of acetyl‐CoA into fatty acid and mevalonate biosynthesis, and finally facilitated the formation of astaxanthin. ROS analysis suggested that the antioxidant mechanism in X. dendrorhous can be induced by 6‐BAP. Additionally, the process of 6‐BAP significantly upregulated the expression of six key genes involved in pathways related to astaxanthin biosynthesis. This research demonstrates the metabolomic mechanism of phytohormone stimulation of astaxanthin production iNn X. dendrorhous and presents a new strategy to improve astaxanthin production to prevent the dilemma of choosing between accumulation of astaxanthin and cell biomass.

In vitro and in vivo recombination of heterologous modules for improving biosynthesis of astaxanthin in yeast

BackgroundAstaxanthin is a kind of tetraterpene and has strong antioxygenic property. The biosynthesis of astaxanthin in engineered microbial chassis has greater potential than its chemical synthesis and extraction from natural producers in an environmental-friendly way. However, the cost-offsetting production of astaxanthin in engineered microbes is still constrained by the poor efficiency of astaxanthin synthesis pathway as a heterologous pathway.ResultsTo address the bottleneck of limited production of astaxanthin in microbes, we developed in vitro and in vivo recombination methods respectively in engineered yeast chassis to optimize the combination of heterologous β-carotene ketolase (crtW) and hydroxylase (crtZ) modules that were selected from different species. As a result, the in vitro and in vivo recombination methods enhanced the astaxanthin yield respectively to 2.11–8.51 folds and 3.0–9.71 folds compared to the initial astaxanthin pathway, according to the different combination of particular genes. The highest astaxanthin producing strain yQDD022 was constructed by in vivo method and produced 6.05 mg g−1 DCW of astaxanthin. Moreover, it was proved that the in vivo recombination method showed higher DNA-assembling efficiency than the in vitro method and contributed to higher stability to the engineered yeast strains.ConclusionsThe in vitro and in vivo recombination methods of heterologous modules provide simple and efficient ways to improve the astaxanthin yield in yeast. Both the two methods enable high-throughput screening of heterologous pathways through recombination of certain crtW and crtZ derived from different species. This study not only exploited the underlying optimal combination of crtZ and crtW for astaxanthin synthesis, but also provided a general approach to evolve a heterologous pathway for the enhanced accumulation of desired biochemical products.

Enhancing Astaxanthin Biosynthesis by Rhodosporidium toruloides Mutants and Optimization of Medium Compositions Using Response Surface Methodology

Astaxanthin is a valuable carotenoid, which has been approved as a food coloring by the US Food and Drug Administration and is considered as a food dye by the European Union (European Commission). This work aimed to attain Rhodosporidium toruloides mutants for enhanced astaxanthin accumulation using ultraviolet (UV) and gamma irradiation mutagenesis. Gamma irradiation was shown to be more efficient than UV for producing astaxanthin-overproducer. Among the screened mutants, G17, a gamma-induced mutant, exhibited the highest astaxanthin production, which was significantly higher than that of the wild strain. Response surface methodology was then applied to optimize the medium compositions for maximizing astaxanthin production by the mutant G17. The optimal medium compositions for the cultivation of G17 were determined as a peptone concentration of 19.75 g/L, malt extract concentration of 13.56 g/L, and glucose concentration of 19.92 g/L, with the maximum astaxanthin yield of 3021.34 µg/L ± 16.49 µg/L. This study suggests that the R. toruloides mutant (G17) is a potential candidate for astaxanthin production.

Novel insights into salinity-induced lipogenesis and carotenogenesis in the oleaginous astaxanthin-producing alga Chromochloris zofingiensis: a multi-omics study

BackgroundChromochloris zofingiensis, a freshwater alga capable of synthesizing both triacylglycerol (TAG) and astaxanthin, has been receiving increasing attention as a leading candidate producer. While the mechanism of oleaginousness and/or carotenogenesis has been studied under such induction conditions as nitrogen deprivation, high light and glucose feeding, it remains to be elucidated in response to salt stress, a condition critical for reducing freshwater footprint during algal production processes.ResultsFirstly, the effect of salt concentrations on growth, lipids and carotenoids was examined for C. zofingiensis, and 0.2 M NaCl demonstrated to be the optimal salt concentration for maximizing both TAG and astaxanthin production. Then, the time-resolved lipid and carotenoid profiles and comparative transcriptomes and metabolomes were generated in response to the optimized salt concentration for congruent analysis. A global response was triggered in C. zofingiensis allowing acclimation to salt stress, including photosynthesis impairment, ROS build-up, protein turnover, starch degradation, and TAG and astaxanthin accumulation. The lipid metabolism involved a set of stimulated biological pathways that contributed to carbon precursors, energy and reductant molecules, pushing and pulling power, and storage sink for TAG accumulation. On the other hand, salt stress suppressed lutein biosynthesis, stimulated astaxanthin biosynthesis (mainly via ketolation), yet had little effect on total carotenoid flux, leading to astaxanthin accumulation at the expense of lutein. Astaxanthin was predominantly esterified and accumulated in a well-coordinated manner with TAG, pointing to the presence of common regulators and potential communication for the two compounds. Furthermore, the comparison between salt stress and nitrogen deprivation conditions revealed distinctions in TAG and astaxanthin biosynthesis as well as critical genes with engineering potential.ConclusionsOur multi-omics data and integrated analysis shed light on the salt acclimation of C. zofingiensis and underlying mechanisms of TAG and astaxanthin biosynthesis, provide engineering implications into future trait improvements, and will benefit the development of this alga for production uses under saline environment, thus reducing the footprint of freshwater.

A novel fed-batch strategy enhances lipid and astaxanthin productivity without compromising biomass of Chromochloris zofingiensis

To improve lipid and astaxanthin productivity without compromising biomass during the whole cultivation period, carbon-dependent kinetics involving nitrogen stress was applied under excess light to elevate intracellular carbon availability and metabolic activity of Chromochloris zofingiensis. Results suggested that fed-batch strategy proposed could increase lipid and astaxanthin productivity to 457.1 and 2.0 mg L−1 d−1, respectively. Biomass productivity at 1084.3 mg L−1 d−1 was comparable with that under suitable condition. Then 13C tracer-based metabolic flux analysis (13C-MFA) demonstrated that central carbon metabolism provided ATP, NADPH and carbon availability for lipid biosynthesis during the strategy. In combination with targeted metabolite analysis, 13C-MFA revealed that the strategy improved precursor content for lipid biosynthesis and elevated path rate to synthesize C16:0 and C18:0. The enhanced lipid content potentially accounted for the high biomass productivity. Therefore, comprehensively understanding relationships between carbon availability and carbon conversion could precisely design strategy for productivity improvements during cultivation.

About the Cover

Previous articleNext article FreeAbout the CoverPDFPDF PLUSFull Text Add to favoritesDownload CitationTrack CitationsPermissionsReprints Share onFacebookTwitterLinked InRedditEmailQR Code SectionsMoreCoverThe cover features the Hawaiian endemic shrimp, Halocaridina rubra, known by native Hawaiians as ‘ōpae ‘ula (literally, tiny red shrimp). These shrimp live in anchialine pools, which are small, shallow surface bodies of water with extensive subterranean connections to the sea and underground aquifers that can experience widely fluctuating gradients in temperature, salinity, and light. True to their Hawaiian name, they reach a maximum length of around one centimeter and typically display a red coloration with substantial variation. Halocaridina rubra is distributed across the islands of Oahu, Maui, and Hawaii, and at least eight unique genetic lineages have been identified. In the cover photo, each shrimp represents a different genetic lineage and displays the “typical” coloration observed for that lineage, highlighting the variation.In this issue (pp. 119–130), R. J. Weaver, B. K. Gonzalez, S. R. Santos, and J. C. Havird identify and quantify the pigment responsible for red coloration in H. rubra. Using chromatography and spectrophotometry, they found that the red carotenoid, astaxanthin, was more abundant in lineages that appeared redder than in lineages that were less colorful. Carotenoid supplementation experiments revealed that less colorful lineages were not limited by dietary access to pigments, suggesting a possible genetic basis for red coloration. By using a bioinformatic approach to explore the genetic basis for production of astaxanthin, they found that the gene responsible for astaxanthin production in H. rubra and other crustaceans is more closely related to a carotenoid metabolism gene found in fungi than to an astaxanthin gene found in birds and turtles. Despite the prevalence of red carotenoid coloration in animals, the genetic basis for this phenotype has only recently been explored and then mostly in vertebrate animals. Identifying the genetic basis for carotenoid coloration in non-vertebrate animals may prove useful in understanding the selective forces that drive the evolution of conspicuous coloration in this group.Credits: Photo, Ryan J. Weaver, University of Texas at Austin. Cover design, Jeannie Harrell, University of Chicago Press, and Ryan J. Weaver. Previous articleNext article DetailsFiguresReferencesCited by The Biological Bulletin Volume 238, Number 2April 2020 Published in association with the Marine Biological Laboratory Article DOIhttps://doi.org/10.1086/709957 © 2020 by The University of Chicago. All rights reserved.PDF download Crossref reports no articles citing this article.

Red Coloration in an Anchialine Shrimp: Carotenoids, Genetic Variation, and Candidate Genes.

Red coloration is a widely distributed phenotype among animals, yet the pigmentary and genetic bases for this phenotype have been described in relatively few taxa. Here we show that the Hawaiian endemic anchialine shrimp Halocaridina rubra is red because of the accumulation of astaxanthin. Laboratory colonies of phylogenetically distinct lineages of H. rubra have colony-specific amounts of astaxanthin that are developmentally, and likely genetically, fixed. Carotenoid supplementation and restriction experiments failed to change astaxanthin content from the within-colony baseline levels, suggesting that dietary limitation is not a major factor driving coloration differences. A possible candidate gene product predicted to be responsible for the production of astaxanthin in H. rubra and other crustaceans is closely related to the bifunctional cytochrome P450 family 3 enzyme CrtS found in fungi. However, homologs to the enzyme thought to catalyze ketolation reactions in birds and turtles, CYP2J19, were not found. This work is one of the first steps in linking phenotypic variation in red coloration of H. rubra to genotypic variation. Future work should focus on (1) pinpointing the genes that function in the bioconversion of dietary carotenoids to astaxanthin, (2) examining what genomic variants might drive variation in coloration among discrete lineages, and (3) testing more explicitly for condition-dependent carotenoid coloration in crustaceans.

Turning a green alga red: engineering astaxanthin biosynthesis by intragenic pseudogene revival in Chlamydomonas reinhardtii.

SummaryThe green alga Chlamydomonas reinhardtii does not synthesize high‐value ketocarotenoids like canthaxanthin and astaxanthin; however, a β‐carotene ketolase (CrBKT) can be found in its genome. CrBKT is poorly expressed, contains a long C‐terminal extension not found in homologues and likely represents a pseudogene in this alga. Here, we used synthetic redesign of this gene to enable its constitutive overexpression from the nuclear genome of C. reinhardtii. Overexpression of the optimized CrBKT extended native carotenoid biosynthesis to generate ketocarotenoids in the algal host causing noticeable changes the green algal colour to reddish‐brown. We found that up to 50% of native carotenoids could be converted into astaxanthin and more than 70% into other ketocarotenoids by robust CrBKT overexpression. Modification of the carotenoid metabolism did not impair growth or biomass productivity of C. reinhardtii, even at high light intensities. Under different growth conditions, the best performing CrBKT overexpression strain was found to reach ketocarotenoid productivities up to 4.3 mg/L/day. Astaxanthin productivity in engineered C. reinhardtii shown here might be competitive with that reported for Haematococcus lacustris (formerly pluvialis) which is currently the main organism cultivated for industrial astaxanthin production. In addition, the extractability and bio‐accessibility of these pigments were much higher in cell wall‐deficient C. reinhardtii than the resting cysts of H. lacustris. Engineered C. reinhardtii strains could thus be a promising alternative to natural astaxanthin producing algal strains and may open the possibility of other tailor‐made pigments from this host.

Enhancement of astaxanthin production by Haematococcus pluvialis using magnesium aminoclay nanoparticles

Improving the content and production of high-value ketocarotenoid pigments is critical for the commercialization of microalgal biorefineries. This study reported the use of magnesium aminoclay (MgAC) nanoparticles for enhancement of astaxanthin production by Haematococcus pluvialis in photoautotrophic cultures. Addition of 1.0 g/L MgAC significantly promoted cellular astaxanthin biosynthesis (302 ± 69 pg/cell), presumably by inducing tolerable oxidative stress, corresponding to a 13.7-fold higher production compared to that in the MgAC-untreated control (22 ± 2 pg/cell). The lipid content and cell size of H. pluvialis improved by 13.6- and 2.1-fold, respectively, compared to that of the control. Despite reduced cell numbers, the overall astaxanthin production (10.3 ± 0.4 mg/L) improved by 40% compared to the control (7.3 ± 0.6 mg/L), owing to improved biomass production. However, an MgAC dosage above 1.0 g/L inhibited biomass production by inducing electrostatic cell wall destabilization and aggregation. Therefore, MgAC-induced stimulation of algae varies widely based on their morphological and physiological characteristics.

Production of Biogas and Astaxanthin from Fruit and Vegetable Wastes Using an Integrated System

The use of fruit and vegetable wastes in biogas production is an attractive option, as it provides simultaneous waste treatment and energy production. The use of the CO2 in biogas for algae cultivation with a zero waste approach would make this process even more attractive. In this way, biogas enrichment, which is traditionally done using economically costly and non-environmentally friendly methods, would be improved, and algae cultivation would become more economical. In the first part of this study, the operation conditions for the biogas reactor and the algae reactor for astaxanthin production were optimized separately. Rates of up to 1.2 L CH4/day and yields up to 0.5 L CH4/g volatile solids were obtained with a 2.5 g dry matter/L day organic loading rate with an anaerobic bioreactor, and 5.1 mg/g astaxanthin was produced by air feeding. When it was decided that sufficient astaxanthin was produced, astaxanthin was obtained using vegetable oils (olive and nut), an environmentally friendly extraction method. In the second part of this study, the anaerobic bioreactor and the algae reactor were integrated, and 6 mg/g astaxanthin production was observed using fruit and vegetable wastes as the substrate for biogas production and the CO2 in biogas for cultivation of Haematococcus pluvialis and therefore astaxanthin production. The integrated system resulted in higher astaxhantin production with a zero waste approach. Moreover, the residual biomass remaining after extraction was fed back into the biogas reactor as a substrate, adopting a zero waste biorefinery approach.

Optimised Production and Extraction of Astaxanthin from the Yeast Xanthophyllomyces dendrorhous.

Currently, astaxanthin demand is fulfilled by chemical synthesis using petroleum-based feedstocks. As such, alternative pathways of natural astaxanthin production attracts much research interest. This study aimed at optimising bioreactor operation parameters for astaxanthin production and evaluating strategies for its subsequent extraction. The effect of pH and agitation was evident, as a significant reduction in both biomass and astaxanthin production was observed when the culture pH was not controlled and a low agitation speed was applied. At controlled pH conditions and a high agitation speed, a significant increase in biomass (16.4 g/L) and astaxanthin production (3.6 mg/L) was obtained. Enzymatic yeast cell lysis using two commercial enzymes (Accellerase 1500 and Glucanex) was optimised using the central composite design of experiment (DoE). Accellerase 1500 led to mild cell disruption and only 9% (w/w) astaxanthin extraction. However, Glucanex treatment resulted in complete astaxanthin extractability, compared to standard extraction method (DMSO/acetone). When supercritical CO2 was employed as an extraction solvent in Accellerase-pre-treated Xanthophyllomyces dendrorhous cells, astaxanthin extraction increased 2.5-fold. Overall, the study showed that extraction conditions can be tailored towards targeted pigments present in complex mixtures, such as in microbial cells.

High irradiance compensated with CO2 enhances the efficiency of Haematococcus lacustris growth

Haematococcus lacustris (H. lacustris), a promising source for astaxanthin production, is a light-sensitive microalga that is prone to sluggish growth when subjected to high levels of irradiance. A challenge in H. lacustris culture is to find a way to efficiently use illumination to maintain vigorous growth and harvest dense biomass, which is essential for further exploiting the potential for astaxanthin production. Previous studies have shown that in addition to illumination, carbon supply in culture is a key limitation for algae growth. Here, we investigated a combined culture approach involving high light intensity (110 μmol m−2s-1) and injection of a 1% (v/v) CO2 air-gas mixture which provided an effective method for H. lacustris culture to achieve both a high growth rate and high cell density. The cell number in the group with high light exposure combined with CO2 enrichment was increased almost four-fold compared with a high light group (110 μmol m−2s-1 without CO2 injection). Additional experiments suggested a possible mechanism in which elevated CO2 increases the electron sink capacity, thus alleviating photoinhibition and oxidative damage. The scaled-up photobioreactor demonstrated much better performance, with growth rates improved by 50–350 %, providing further evidence that this new method can improve algal cell production. Overall, our work provides an efficient way for H. lacustris culture and manufacture, with potential industrial applications.

Initial study of polymer-based nanoparticles effect on carotenogenesis of Haematococcus lacustris

The carotenogenesis in Haematococcus lacustris, accompanying by encystment starting from green motile cells to aplanospores, is suggested mediating by reactive oxygen species (ROS). Our previous study showed that isobutyl-cyanoacrylate nanoparticles (IBCA- NPs) induced the cell mortality, preceding by intracellular ROS accumulation, in most of Volvocales species but not H. lacustris. This study aimed to investigate the effect of IBCA- NPs exposure to H. lacustris on carotenogenesis. The cells were cultured on C medium then exposed to 100 mg · L−1 -1 of 180 nm of IBCA-NPs for the 14 days. The accumulation of astaxanthin was started after four days of IBCA-NPs exposure. The ratio of intermediate cells reached to 46% after 14 days of IBCA-NPs exposure, while the ratio of intermediate cells of untreated culture was only 11%. The results suggested that the addition of nanoparticles could enhance the production of astaxanthin. Further studies are required to investigate the correlation between nanoparticle exposure and ROS generation that leads to the biosynthesis of astaxanthin. The condition of cell growth and the addition of nanoparticles shall be optimized to increase the production of ketocarotenoid and to minimalize the negative effect of longer exposure time of IBCA-NP that could lead the cell mortality.

Metallopeptidase Stp1 activates the transcription factor Sre1 in the carotenogenic yeast Xanthophyllomyces dendrorhous

Xanthophyllomyces dendrorhous is a basidiomycete yeast known as a natural producer of astaxanthin, a carotenoid of commercial interest because of its antioxidant properties. Recent studies indicated that X. dendrorhous has a functional SREBP pathway involved in the regulation of isoprenoid compound biosynthesis, which includes ergosterol and carotenoids. SREBP is a major regulator of sterol metabolism and homeostasis in mammals; characterization in fungi also provides information about its role in the hypoxia adaptation response and virulence. SREBP protease processing is required to activate SREBP pathway functions in fungi. Here, we identified and described the STP1 gene, which encodes a metallopeptidase of the M50 family involved in the proteolytic activation of the transcription factor Sre1 of the SREBP pathway, in X. dendrorhous We assessed STP1 function in Δstp1 strains derived from the wild-type and a mutant of ergosterol biosynthesis that overproduces carotenoids and sterols. Bioinformatic analysis of the deduced protein predicted the presence of characteristic features identified in homologs from mammals and fungi. The Δstp1 mutation decreased yeast growth in the presence of azole drugs and reduced transcript levels of Sre1-dependent genes. This mutation also negatively affected the carotenoid- and sterol-overproducing phenotype. Western blot analysis demonstrated that Sre1 was activated in the yeast ergosterol biosynthesis mutant and that the Δstp1 mutation introduced in this strain prevented Sre1 proteolytic activation. Overall, our results demonstrate that STP1 encodes a metallopeptidase involved in proteolytic activation of Sre1 in X. dendrorhous, contributing to our understanding of fungal SREBP pathways.

Enhanced astaxanthin production in yeast via combined mutagenesis and evolution

Astaxanthin exhibiting strong antioxidant activity, is widely used as food and animal feed additives as well as in cosmetics, pharmaceutical and nutraceuticals industries. Microbial production of astaxanthin has received increasing concerns in recent years. A combined strategy consisting of physical mutagenesis by atmospheric and room temperature plasma (ARTP) and adaptive evolution driven by H2O2, was established to expand disturbance in genome of the engineered Saccharomyces cerevisiae. A titer of 65.9 mg/L astaxanthin at flask level was obtained via multiple rounds of crossed ARTP and H2O2 treatment, which was increased nearly 4-fold compared with the starting strain. Moreover, a trend of decreasing intracellular reactive oxygen species (ROS) and increasing chronological lifespan (CLS) was observed in cells with improved titer of astaxanthin. Eventually, a highest reported titer of 404.78 mg/L astaxanthin in S. cerevisiae was achieved in 5-L fermenter by further optimization of fermentation conditions. Our study not only highlights the iterative cycling of the integrated strategy combined by ARTP and adaptive evolution tightly based on the natural properties of the targets provides an efficient route to boost product accumulation, but also demonstrates it can serve as a general strategy for other microbial cell factories.

Astaxanthin accumulation difference between non-motile cells and akinetes of Haematococcus pluvialis was affected by pyruvate metabolism

BackgroundHaematococcus pluvialis is the best source of natural astaxanthin, known as the king of antioxidants. H. pluvialis have four cell forms: spore, motile cell, non-motile cell and akinete. Spores and motile cells are susceptible to photoinhibition and would die under photoinduction conditions. Photoinduction using non-motile cells as seeds could result in a higher astaxanthin production than that using akinetes. However, the mechanism of this phenomenon has not been clarified.ResultsTranscriptome was sequenced and annotated to illustrate the mechanism of this phenomenon. All differentially expressed genes involved in astaxanthin biosynthesis were up-regulated. Particularly, chyb gene was up-regulated by 16-fold, improving the conversion of β-carotene into astaxanthin. Pyruvate was the precursor of carotenoids biosynthesis. Pyruvate kinase gene expression level was increased by 2.0-fold at the early stage of akinetes formation. More changes of gene transcription occurred at the early stage of akinetes formation, 52.7% and 51.9% of total DEGs in control group and treatment group, respectively.ConclusionsGenes transcription network was constructed and the synthesis mechanism of astaxanthin was clarified. The results are expected to further guide the in-depth optimization of the astaxanthin production process in H. pluvialis by improving pyruvate metabolism.