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1.
Microorganisms ; 11(9)2023 Aug 22.
Article in English | MEDLINE | ID: mdl-37763976

ABSTRACT

Bisphenol A (BPA), one of the most widely used plasticizers, is an endocrine-disrupting chemical that is released from plastic products. The aim of this study was to screen and characterize bacteria with excellent BPA-degrading abilities for application in foods. BPA degradation ability was confirmed in 127 of 129 bacterial strains that were isolated from fermented soybean foods. Among the strains, B. subtilis P74, which showed the highest BPA degradation performance, degraded 97.2% of 10 mg/L of BPA within 9 h. This strain not only showed a fairly stable degradation performance (min > 88.2%) over a wide range of temperatures (30-45 °C) and pH (5.0-9.0) but also exhibited a degradation of 63% against high concentrations of BPA (80 mg/L). The metabolites generated during the degradation were analyzed using high-performance liquid chromatography-mass spectrometry, and predicted degradation pathways are tentatively proposed. Finally, the application of this strain to soybean fermentation was conducted to confirm its applicability in food.

2.
Int J Mol Sci ; 24(10)2023 May 16.
Article in English | MEDLINE | ID: mdl-37240172

ABSTRACT

Punicic acid (PuA) is a polyunsaturated fatty acid with significant medical, biological, and nutraceutical properties. The primary source of punicic acid is the pomegranate seed oil obtained from fruits of trees that are mainly cultivated in subtropical and tropical climates. To establish sustainable production of PuA, various recombinant microorganisms and plants have been explored as platforms with limited efficiencies. In this study, the oleaginous yeast Yarrowia lipolytica was employed as a host for PuA production. First, growth and lipid accumulation of Y. lipolytica were evaluated in medium supplemented with pomegranate seed oil, resulting in the accumulation of lipids up to 31.2%, consisting of 22% PuA esterified in the fraction of glycerolipids. In addition, lipid-engineered Y. lipolytica strains, transformed with the bifunctional fatty acid conjugase/desaturase from Punica granatum (PgFADX), showed the ability to accumulate PuA de novo. PuA was detected in both polar and neutral lipid fractions, especially in phosphatidylcholine and triacylglycerols. Promoter optimization for PgFADX expression resulted in improved accumulation of PuA from 0.9 to 1.8 mg/g of dry cell weight. The best-producing strain expressing PgFADX under the control of a strong erythritol-inducible promoter produced 36.6 mg/L PuA. These results demonstrate that the yeast Y. lipolytica is a promising host for PuA production.


Subject(s)
Yarrowia , Fatty Acid Desaturases/metabolism , Linolenic Acids/metabolism , Plant Oils/metabolism , Fatty Acids/metabolism
3.
Waste Manag ; 165: 70-81, 2023 Jun 15.
Article in English | MEDLINE | ID: mdl-37086658

ABSTRACT

Phenolic recovery from agricultural byproducts has been highlighted due to their health-promoting bioactivities. However, uncontrolled discard of residues after extraction process would induce environmental pollution and bioresource waste. In this study, biorefining of phenolic-rich rapeseed meal (RSM) and its defatted sample (dRSM) was attempted by holistic utilization of phenolic extract and residue separately. Phenolic removal could significantly improve residues' Cr(VI) adsorption capacities by about 21%, which presented extended physical surface and more released functional groups. Moreover, simulating raw material by remixing 3% separated phenolic extracts or main component sinapic acid therein with corresponding residues further improved about 12% adsorption efficiencies. These indicated that the different present forms of phenolics had opposite effects on Cr(VI) removal. While natural conjugational form inhibited hosts' biosorption, free form had enhanced functions for either extract or residue. Four optimal adsorption parameters (pH, adsorbent dosage, contact time and initial Cr(VI) concentration), three kinetic (pseudo-first order, pseudo-second order and intra-particle diffusion) models and two isotherms (Langmuir and Freundlich) were used to reveal the adsorption process. The maximum Cr(VI) adsorption capacity on residues could reach about 100 mg/g, which was superior to that of most biosorbents derived from agricultural byproducts, even some biochar. Together with the residues' advantages with everlasting capacity after 3 adsorption-desorption cycles and excellent abilities for adsorbing multiple co-existed metal ions (Cr(VI), Cd(II), Cu(II), Pb(II), Ni(II) and Zn(II)), phenolic recovery was first proved to be a new and sustainable strategy for modifying biosorbents from agricultural byproducts with zero waste.


Subject(s)
Brassica napus , Water Pollutants, Chemical , Hydrogen-Ion Concentration , Chromium/chemistry , Adsorption , Kinetics , Plant Extracts , Water Pollutants, Chemical/analysis
4.
Microb Cell Fact ; 22(1): 7, 2023 Jan 12.
Article in English | MEDLINE | ID: mdl-36635727

ABSTRACT

BACKGROUND: The oleaginous yeast Yarrowia lipolytica is increasingly used as a chassis strain for generating bioproducts. Several hybrid promoters with different strengths have been developed by combining multiple copies of an upstream activating sequence (UAS) associated with a TATA box and a core promoter. These promoters display either constitutive, phase-dependent, or inducible strong expression. However, there remains a lack of bidirectional inducible promoters for co-expressing genes in Y. lipolytica. RESULTS: This study built on our previous work isolating and characterizing the UAS of the erythritol-induced genes EYK1 and EYD1 (UAS-eyk1). We found an erythritol-inducible bidirectional promoter (BDP) located in the EYK1-EYL1 intergenic region. We used the BDP to co-produce YFP and RedStarII fluorescent proteins and demonstrated that the promoter's strength was 2.7 to 3.5-fold stronger in the EYL1 orientation compared to the EYK1 orientation. We developed a hybrid erythritol-inducible bidirectional promoter (HBDP) containing five copies of UAS-eyk1 in both orientations. It led to expression levels 8.6 to 19.2-fold higher than the native bidirectional promoter. While the BDP had a twofold-lower expression level than the strong constitutive TEF promoter, the HBDP had a 5.0-fold higher expression level when oriented toward EYL1 and a 2.4-fold higher expression level when oriented toward EYK1. We identified the optimal media for BDP usage by exploring yeast growth under microbioreactor conditions. Additionally, we constructed novel Golden Gate biobricks and a destination vector for general use. CONCLUSIONS: In this research, we developed novel bidirectional and hybrid bidirectional promoters of which expression can be fine-tuned, responding to the need for versatile promoters in the yeast Y. lipolytica. This study provides effective tools that can be employed to smoothly adjust the erythritol-inducible co-expression of two target genes in biotechnology applications. BDPs developed in this study have potential applications in the fields of heterologous protein production, metabolic engineering, and synthetic biology.


Subject(s)
Yarrowia , Yarrowia/metabolism , Erythritol/metabolism , Synthetic Biology , Promoter Regions, Genetic , Metabolic Engineering
5.
Trends Biotechnol ; 41(2): 242-254, 2023 02.
Article in English | MEDLINE | ID: mdl-35940976

ABSTRACT

Yarrowia lipolytica possesses natural and engineered traits that make it a good host for the industrial bioproduction of chemicals, fuels, foods, and pharmaceuticals. In recent years, academic and industrial researchers have assessed its potential, developed synthetic biology techniques, improved its features, scaled its processes, and identified its limitations. Both publications and patents related to Y. lipolytica have shown a drastic increase during the past decade. Here, we discuss the characteristics of this yeast that make it suitable for industry and the remaining challenges for its wider use at large scale. We present evidence herein that shows the importance and potential of Y. lipolytica in bioproduction such that it may soon be one of the preferred choices of industry.


Subject(s)
Yarrowia , Yarrowia/genetics , Metabolic Engineering/methods , Industry , Synthetic Biology
6.
Biomolecules ; 12(8)2022 07 25.
Article in English | MEDLINE | ID: mdl-35892336

ABSTRACT

Yarrowia lipolytica is a promising oleaginous yeast for producing unusual lipids, such as odd-chain fatty acids (OCFA). Their diverse applications and low natural production make OCFA particularly interesting. In recent studies, inhibiting the catabolic pathway of precursor, boosting precursor pools, and optimizing substrate combination greatly improved the production of OCFA in Y. lipolytica. We explored the lipid readjustment of OCFA in engineered Y. lipolytica strains. NPLC-Corona-CAD® evidenced a time-dependent overproduction of free fatty acids, diglycerides, and phosphatidylcholine (PC) in obese LP compared to obese L. Phosphatidylethanolamine (PE) and phosphatidylinositol, largely overproduced in obese LP at 72 h compared to obese L, vanished at 216 h. The fatty acyls (FAs) composition of glycero- and glycerophospholipids was determined by NPLC-APPI+-HRMS from in-source generated monoacylglycerol-like fragment ions. C18:1 and C17:1 were predominant acylglycerols in obese L and obese LP, respectively. Phosphatidic acid, PE, and PC exhibited similar FAs composition but differed in their molecular species distributions. Cardiolipin (CL) is known to contain mostly C18:2 FAs corresponding to the composition in obese L, 50% of C18:2, and 35% of C18:1. In obese LP, both FAs dropped to drop to 20%, and C17:1 were predominant, reaching 55%. We hypothesize that CL-modified composition in obese LPs may alter mitochondrial function and limit lipid production.


Subject(s)
Yarrowia , Fatty Acids/metabolism , Obesity , Yarrowia/metabolism
7.
FEMS Yeast Res ; 21(6)2021 09 11.
Article in English | MEDLINE | ID: mdl-34453534

ABSTRACT

In recent years, there has been a growing interest in the use of renewable sources for bio-based production aiming at developing sustainable and feasible approaches towards a circular economy. Among these renewable sources, organic wastes (OWs) can be anaerobically digested to generate carboxylates like volatile fatty acids (VFAs), lactic acid, and longer-chain fatty acids that are regarded as novel building blocks for the synthesis of value-added compounds by yeasts. This review discusses on the processes that can be used to create valuable molecules from OW-derived VFAs; the pathways employed by the oleaginous yeast Yarrowia lipolytica to directly metabolize such molecules; and the relationship between OW composition, anaerobic digestion, and VFA profiles. The review also summarizes the current knowledge about VFA toxicity, the pathways by which VFAs are metabolized and the metabolic engineering strategies that can be employed in Y. lipolytica to produce value-added biobased compounds from VFAs.


Subject(s)
Yarrowia , Carboxylic Acids , Fatty Acids , Fatty Acids, Volatile , Metabolic Engineering , Yarrowia/genetics
8.
Metab Eng Commun ; 12: e00158, 2021 Jun.
Article in English | MEDLINE | ID: mdl-33391990

ABSTRACT

Microbial production of lipids is one of the promising alternatives to fossil resources with increasing environmental and energy concern. Odd-chain fatty acids (OCFA), a type of unusual lipids, are recently gaining a lot of interest as target compounds in microbial production due to their diverse applications in the medical, pharmaceutical, and chemical industries. In this study, we aimed to enhance the pool of precursors with three-carbon chain (propionyl-CoA) and five-carbon chain (ß-ketovaleryl-CoA) for the production of OCFAs in Yarrowia lipolytica. We evaluated different propionate-activating enzymes and the overexpression of propionyl-CoA transferase gene from Ralstonia eutropha increased the accumulation of OCFAs by 3.8 times over control strain, indicating propionate activation is the limiting step of OCFAs synthesis. It was shown that acetate supplement was necessary to restore growth and to produce a higher OCFA contents in total lipids, suggesting the balance of the precursors between acetyl-CoA and propionyl-CoA is crucial for OCFA accumulation. To improve ß-ketovaleryl-CoA pools for further increase of OCFA production, we co-expressed the bktB encoding ß-ketothiolase in the producing strain, and the OCFA production was increased by 33% compared to control. Combining strain engineering and the optimization of the C/N ratio promoted the OCFA production up to 1.87 â€‹g/L representing 62% of total lipids, the highest recombinant OCFAs titer reported in yeast, up to date. This study provides a strong basis for the microbial production of OCFAs and its derivatives having high potentials in a wide range of applications.

9.
Microorganisms ; 8(12)2020 Dec 06.
Article in English | MEDLINE | ID: mdl-33291339

ABSTRACT

Using microorganisms as lipid-production factories holds promise as an alternative method for generating petroleum-based chemicals. The non-conventional yeast Yarrowia lipolytica is an excellent microbial chassis; for example, it can accumulate high levels of lipids and use a broad range of substrates. Furthermore, it is a species for which an array of efficient genetic engineering tools is available. To date, extensive work has been done to metabolically engineer Y. lipolytica to produce usual and unusual lipids. Unusual lipids are scarce in nature but have several useful applications. As a result, they are increasingly becoming the targets of metabolic engineering. Unusual lipids have distinct structures; they can be generated by engineering endogenous lipid synthesis or by introducing heterologous enzymes to alter the functional groups of fatty acids. In this review, we describe current metabolic engineering strategies for improving lipid production and highlight recent researches on unusual lipid production in Y. lipolytica.

10.
Medicine (Baltimore) ; 99(34): e21751, 2020 Aug 21.
Article in English | MEDLINE | ID: mdl-32846799

ABSTRACT

INTRODUCTION: Chryseobacterium arthrosphaerae is a gram-negative bacteria, known for its intrinsic multidrug resistance, which can lead to treatment difficulties. PATIENT CONCERNS: A 56-year-old male had an indwelling external ventricular drainage catheter for 6 months and had been frequently treated with antibiotics for nosocomial infections. He showed cerebrospinal fluid pleocytosis and an abrupt fever during hospitalization. DIAGNOSIS: He was diagnosed as a ventriculitis caused by Chryseobacterium arthrosphaerae (C arthrosphaerae). INTERVENTION: Initially, we used ciprofloxacin as the backbone in combination with minocycline (and rifampin). However, fever and pleocytosis persisted, and improvement was slow. We then switched the minocycline and rifampin regiment to trimethoprim/sulfamethoxazole. Following this switch of antibiotics, the patient's pleocytosis rapidly improved, allowing the replacement of his external ventricular drainage catheters. C arthrospharae was no longer growing in cerebrospinal fluid and he was recovered from ventriculitis. OUTCOMES: The patient remains alive without any incidence of C arthrosphaerae recurrence. CONCLUSION: We propose trimethoprim/sulfamethoxazole alone or in combination with ciprofloxacin to be good candidates for the treatment of ventriculitis by C arthrosphaerae.


Subject(s)
Anti-Bacterial Agents/therapeutic use , Cerebral Ventriculitis/drug therapy , Chryseobacterium , Anti-Bacterial Agents/administration & dosage , Catheters, Indwelling , Cerebral Ventriculitis/complications , Cerebral Ventriculitis/diagnosis , Drainage , Drug Therapy, Combination , Humans , Leukocytosis/etiology , Male , Middle Aged , Minocycline/therapeutic use , Rifampin/therapeutic use , Trimethoprim, Sulfamethoxazole Drug Combination/therapeutic use
11.
Foods ; 9(7)2020 Jul 11.
Article in English | MEDLINE | ID: mdl-32664514

ABSTRACT

The study evaluated the role of Enterococcus faecium in tyramine production and its response to fermentation temperature in a traditional Korean fermented soybean paste, Cheonggukjang. Tyramine content was detected in retail Cheonggukjang products at high concentrations exceeding the recommended limit up to a factor of 14. All retail Cheonggukjang products contained Enterococcus spp. at concentrations of at least 6 Log CFU/g. Upon isolation of Enterococcus strains, approximately 93% (157 strains) produced tyramine at over 100 µg/mL. The strains that produced the highest concentrations of tyramine (301.14-315.29 µg/mL) were identified as E. faecium through 16S rRNA sequencing. The results indicate that E. faecium is one of the major contributing factors to high tyramine content in Cheonggukjang. During fermentation, tyramine content in Cheonggukjang groups co-inoculated with E. faecium strains was highest at 45 °C, followed by 37 °C and 25 °C. The tyramine content of most Cheonggukjang groups continually increased as fermentation progressed, except groups fermented at 25 °C. At 45 °C, the tyramine content occasionally exceeded the recommended limit within 3 days of fermentation. The results suggest that lowering fermentation temperature and shortening duration may reduce the tyramine content of Cheonggukjang, thereby reducing the safety risks that may arise when consuming food with high tyramine concentrations.

12.
BMC Infect Dis ; 20(1): 286, 2020 Apr 17.
Article in English | MEDLINE | ID: mdl-32303196

ABSTRACT

BACKGROUND: Henoch-Schönlein purpura (HSP) may be caused by several allergens. However, to date, HSP caused by Orientia tsutsugamushi has not been reported. Here, we report an unusual rash with features of HSP caused by Orientia tsutsugamushi. CASE PRESENTATION: A man visited a tertiary hospital with bilateral symmetrical purpura and fever. He presented with an eschar in the left popliteal fossa and proteinuria. He was diagnosed with tsutsugamushi disease by indirect fluorescent antibody and positive polymerase chain reaction tests. Purpura biopsy demonstrated a feature of leukocytoclastic vasculitis and IgA deposition in dermal vessels, indicative of HSP. CONCLUSIONS: When examining patients with unique rashes, such as in this case, we suggest investigating out-door activities and evidence of mite bites. Furthermore, differential diagnosis of tsutsugamushi disease should be considered when necessary.


Subject(s)
IgA Vasculitis/diagnosis , Orientia tsutsugamushi/isolation & purification , Scrub Typhus/diagnosis , Anti-Bacterial Agents/therapeutic use , Biopsy , Diagnosis, Differential , Humans , IgA Vasculitis/drug therapy , IgA Vasculitis/microbiology , IgA Vasculitis/pathology , Male , Middle Aged , Orientia tsutsugamushi/genetics , Orientia tsutsugamushi/immunology , Scrub Typhus/drug therapy , Scrub Typhus/microbiology , Scrub Typhus/pathology , Skin/pathology , Treatment Outcome
13.
Yeast ; 37(1): 131-140, 2020 01.
Article in English | MEDLINE | ID: mdl-31293017

ABSTRACT

Microbial oils are regarded as promising alternatives to fossil fuels. For bio-oil production to be sustainable over the long term, utilizing low-cost substrates like volatile fatty acids (VFAs) is crucial. Increasing attention is being paid to one of the most common VFAs: propionate, a substrate that could be used to produce the odd-chain FAs of industrial interest. However, little is known about microbial responses to propionate-induced stress and the genes involved. Using genomic library screening, we identified two genes involved in propionate tolerance in Yarrowia lipolytica-MFS1 and RTS1. Strains containing each of the genes displayed enhanced tolerance to propionate even when the genes were expressed in truncated form via a replicative plasmid. Compared with the control strain, the strain overexpressing MFS1 under a constitutive promoter displayed greater tolerance to propionate: It had a shorter lag phase and higher growth rate in propionate medium (0.047 hr-1 versus 0.030 hr-1 for the control in 40 g/L propionate); it also accumulated more total lipids and more odd-chain lipids (10% and 3.3%, respectively) than the control. The strain overexpressing RTS1 showed less tolerance for propionate than the strains harboring the truncated form (0.057 hr-1 versus 0.065 hr-1 in 40 g/L propionate medium) but still had higher tolerance than the control strain. Furthermore, the overexpression of RTS1 seemed to confer tolerance to other weak acids such as lactate, formic acid, malic acid, and succinic acid. This work provides a basis for better understanding the response to propionate-induced stress in Y. lipolytica.


Subject(s)
Genes, Fungal , Genomic Library , Propionates/pharmacology , Stress, Physiological , Yarrowia/genetics , Culture Media/chemistry , High-Throughput Nucleotide Sequencing , Propionates/metabolism
14.
Yeast ; 37(1): 103-115, 2020 01.
Article in English | MEDLINE | ID: mdl-31119792

ABSTRACT

Glutamate dehydrogenases (GDHs) are fundamental to cellular nitrogen and energy balance. Yet little is known about these enzymes in the oleaginous yeast Yarrowia lipolytica. The YALI0F17820g and YALI0E09603g genes, encoding potential GDH enzymes in this organism, were examined. Heterologous expression in gdh-null Saccharomyces cerevisiae and examination of Y. lipolytica strains carrying gene deletions demonstrate that YALI0F17820g (ylGDH1) encodes a NADP-dependent GDH whereas YALI0E09603g (ylGDH2) encodes a NAD-dependent GDH enzyme. The activity encoded by these two genes accounts for all measurable GDH activity in Y. lipolytica. Levels of the two enzyme activities are comparable during logarithmic growth on rich medium, but the NADP-ylGDH1p enzyme activity is most highly expressed in stationary and nitrogen starved cells by threefold to 12-fold. Replacement of ammonia with glutamate causes a decrease in NADP-ylGdh1p activity, whereas NAD-ylGdh2p activity is increased. When glutamate is both carbon and nitrogen sources, the activity of NAD-ylGDH2p becomes dominant up to 18-fold compared with that of NADP-ylGDH1p. Gene deletion followed by growth on different carbon and nitrogen sources shows that NADP-ylGdh1p is required for efficient nitrogen assimilation whereas NAD-ylGdh2p plays a role in nitrogen and carbon utilization from glutamate. Overexpression experiments demonstrate that ylGDH1 and ylGDH2 are not interchangeable. These studies provide a vital basis for future consideration of how these enzymes function to facilitate energy and nitrogen homeostasis in Y. lipolytica.


Subject(s)
Glutamate Dehydrogenase/metabolism , Glutamates/metabolism , Yarrowia/enzymology , Yarrowia/growth & development , Culture Media , Gene Deletion , Glutamate Dehydrogenase/genetics , Homeostasis , NAD/metabolism , NADP/metabolism , Nitrogen/metabolism , Saccharomyces cerevisiae/genetics , Yarrowia/genetics
15.
Infect Dis (Lond) ; 52(3): 177-185, 2020 03.
Article in English | MEDLINE | ID: mdl-31718355

ABSTRACT

Introduction: Antinuclear antibody (ANA) tests are widely used for the diagnosis of autoimmune diseases, but ANAs are also commonly found in patients with various infections. This retrospective study aimed to investigate the relationship between infections and ANA status.Methods: Patients that visited the Department of Infectious Diseases at Inha University Hospital between January 2007 and July 2018 were investigated. We analysed their ANA test results and reviewed rheumatic and infectious diagnoses of patients with positive ANA findings.Results: Of the 9,320 patients during the study period, 1,111 underwent ANA testing and 110 tested positive. Seven of the 110 patients were previously diagnosed with ANA-positive disease, and 21 were diagnosed with autoimmune disease during the present study. Of the remaining 82 patients, 43 were confirmed with infectious disease. The most common pathogen was Mycobacterium tuberculosis (n = 10), followed by Treponema pallidum (n = 5), Orientia tsutsugamushi (n = 5), Escherichia coli (n = 5), Bartonella henselae (n = 3), and human immunodeficiency virus (n = 3). Of the 39 patients without a confirmed pathogen, 7 were seropositive for O. tsutsugamushi, B. henselae, or Rickettsia spp. Patients were observed at an average of 24 weeks in our hospital. One patient developed systemic lupus erythematosus after being diagnosed with Epstein-Barr virus-induced infectious mononucleosis, and another patient developed adult-onset Still's disease after being diagnosed with scrub typhus.Conclusion: This study showed that various relationships exist between infections and rheumatic diseases. In particular, several patients with a positive ANA test result were found to have intracellular infections such as mycobacterial infections, syphilis, or scrub typhus.


Subject(s)
Antibodies, Antinuclear/blood , Autoimmune Diseases/blood , Infections/blood , Infections/microbiology , Rheumatic Diseases/blood , Adolescent , Adult , Aged , Aged, 80 and over , Autoimmune Diseases/diagnosis , Bartonella henselae , Escherichia coli , Escherichia coli Infections/blood , Female , HIV Infections/blood , Humans , Male , Middle Aged , Mycobacterium tuberculosis , Orientia tsutsugamushi , Retrospective Studies , Rheumatic Diseases/diagnosis , Rickettsia , Scrub Typhus/blood , Syphilis/blood , Treponema pallidum , Tuberculosis, Pulmonary/blood , Young Adult
16.
Foods ; 8(11)2019 Nov 04.
Article in English | MEDLINE | ID: mdl-31689884

ABSTRACT

Biogenic amines produced during fermentation may be harmful when ingested in high concentrations. As current regulations remain insufficient to ensure the safety of fermented vegetable products, the current study determined the risks associated with the consumption of kimchi by evaluating the biogenic amine concentrations reported by various studies. Upon evaluation, some kimchi products were found to contain histamine and tyramine at potentially hazardous concentrations exceeding the recommended limit of 100 mg/kg for both histamine and tyramine. The biogenic amines may have originated primarily from metabolic activity by microorganisms during fermentation, as well as from Jeotgal (Korean fermented seafood) and Aekjeot (Korean fermented fish sauce) products commonly used as ingredients for kimchi production. Many studies have suggested that Jeotgal and Aekjeot may contribute to the histamine and tyramine content in kimchi. Microorganisms isolated from kimchi and Jeotgal have been reported to produce both histamine and tyramine. Despite the potential toxicological risks, limited research has been conducted on reducing the biogenic amine content of kimchi and Jeotgal products. The regulation and active monitoring of biogenic amine content during kimchi production appear to be necessary to ensure the safety of the fermented vegetable products.

17.
Microb Cell Fact ; 18(1): 167, 2019 Oct 10.
Article in English | MEDLINE | ID: mdl-31601223

ABSTRACT

BACKGROUND: The oleaginous yeast Yarrowia lipolytica is increasingly used as an alternative cell factory for the production of recombinant proteins. Recently, regulated promoters from genes EYK1 and EYD1, encoding an erythrulose kinase and an erythritol dehydrogenase, respectively, have been identified and characterized in this yeast. Hybrid promoters up-regulated by polyols such as erythritol and erythrulose have been developed based on tandem copies of upstream activating sequences from EYK1 (UAS1EYK1) and XPR2 (encoding extracellular protease, UAS1XPR2) promoters. RESULTS: The strength of native (pEYD1) and engineered promoters (pEYK1-3AB and pHU8EYK) was compared using the extracellular lipase CalB from Candida antarctica as a model protein and a novel dedicated host strain. This latter is engineered in polyol metabolism and allows targeted chromosomal integration. In process conditions, engineered promoters pEYK1-3AB and pHU8EYK yielded 2.8 and 2.5-fold higher protein productivity, respectively, as compared to the reference pTEF promoter. We also demonstrated the possibility of multicopy integration in the newly developed host strain. In batch bioreactor, the CalB multi-copy strain RIY406 led to a 1.6 fold increased lipase productivity (45,125 U mL-1) within 24 h as compared to the mono-copy strain. CONCLUSIONS: The expression system described herein appears promising for recombinant extracellular protein production in Y. lipolytica.


Subject(s)
Fungal Proteins , Lipase , Microorganisms, Genetically-Modified , Recombinant Proteins , Yarrowia , Fungal Proteins/genetics , Fungal Proteins/metabolism , Gene Expression/genetics , Lipase/genetics , Lipase/metabolism , Microorganisms, Genetically-Modified/genetics , Microorganisms, Genetically-Modified/metabolism , Promoter Regions, Genetic , Recombinant Proteins/genetics , Recombinant Proteins/metabolism , Yarrowia/genetics , Yarrowia/metabolism
18.
Microb Biotechnol ; 12(6): 1249-1259, 2019 11.
Article in English | MEDLINE | ID: mdl-31148366

ABSTRACT

The oleaginous yeast Yarrowia lipolytica is an established host for the bio-based production of valuable compounds and an organism for which many genetic tools have been developed. However, to properly engineer Y. lipolytica and take full advantage of its potential, we need efficient, versatile, standardized and modular cloning tools. Here, we present a new modular Golden Gate toolkit for the one-step assembly of three transcription units that includes a selective marker and sequences for genome integration. Perfectly suited to a combinatorial approach, it contains nine different validated promoters, including inducible promoters, which allows expression to be fine-tuned. Moreover, this toolbox incorporates six different markers (three auxotrophic markers, two antibiotic-resistance markers and one metabolic marker), which allows the fast sequential construction and transformation of multiple elements. In total, the toolbox contains 64 bricks, and it has been validated and characterized using three different fluorescent reporter proteins. Additionally, it was successfully used to assemble and integrate a three-gene pathway allowing xylose utilization by Y. lipolytica. This toolbox provides a powerful new tool for rapidly engineering Y. lipolytica strains and is available to the community through Addgene.


Subject(s)
Genetics, Microbial/methods , Metabolic Engineering/methods , Synthetic Biology/methods , Yarrowia/genetics , Industrial Microbiology/methods
19.
Foods ; 8(4)2019 Apr 07.
Article in English | MEDLINE | ID: mdl-30959957

ABSTRACT

Spores are resistant against many extreme conditions including the disinfection and sterilization methods used in the food industry. Selective prevention of sporulation of Bacillus species is an ongoing challenge for food scientists and fermentation technologists. This study was conducted to evaluate the effects of single and combined supplementation of calcium and manganese on sporulation of common pathogenic and food spoilage Bacillus species: B. cereus, B. licheniformis, B. subtilis and B. coagulans. Sporulation of Bacillus vegetative cells was induced on sporulation media supplemented with diverse concentrations of the minerals. Under the various mineral supplementation conditions, the degree of sporulation was quantified with colonies formed by the Bacillus spores. The results revealed that B. licheniformis and B. cereus displayed the weakest sporulation capabilities on media with minimal supplementation levels of calcium and manganese. The lowest sporulation of B. subtilis and B. coagulans was observed on media supplemented with the highest level of calcium and low levels of manganese. Depending on effect of supplementation on sporulation, the Bacillus species were divided into two distinct groups: B. licheniformis and B. cereus; and B. subtilis and B. coagulans. The information provides valuable insight to selectively reduce sporulation of Bacillus species undesirable in the food industry.

20.
Foods ; 8(3)2019 Mar 24.
Article in English | MEDLINE | ID: mdl-30909649

ABSTRACT

In this study, biogenic amine content in Pa (green onion) kimchi and Gat (mustard leaf) kimchi, Korean specialty kimchi types, was determined by high-performance liquid chromatography (HPLC). Many kimchi samples contained low levels of biogenic amines, but some samples had histamine and tyramine content over the safe levels. Based on the comparative analysis between the ingredient information on food labels and biogenic amine content of kimchi samples, Myeolchi-aekjeot appeared to be an important source of biogenic amines in both kimchi. Besides, through the 16s rRNA sequence analysis, Lactobacillus brevis appeared to be responsible for the formation of biogenic amines (tyramine, ß-phenylethylamine, putrescine, and cadaverine) in both kimchi, in a strain-dependent manner. During fermentation, a higher accumulation of tyramine, ß-phenylethylamine, and putrescine was observed in both or one (for putrescine) of kimchi types when L. brevis strains served as inocula. The addition of Myeolchi-aekjeot affected the initial concentrations of most biogenic amines (except for spermidine in Gat kimchi) in both kimchi. Therefore, this study suggests that using appropriately salted and fermented seafood products for kimchi preparation and using biogenic amine-negative and/or biogenic amine-degrading starter cultures would be effective in reducing biogenic amine content in Pa kimchi and Gat kimchi.

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