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Biomolecules Jun 2024Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the...
Resveratrol, a phenylpropanoid compound, exhibits diverse pharmacological properties, making it a valuable candidate for health and disease management. However, the demand for resveratrol exceeds the capacity of plant extraction methods, necessitating alternative production strategies. Microbial synthesis offers several advantages over plant-based approaches and presents a promising alternative. stands out among microbial hosts due to its safe nature, abundant acetyl-CoA and malonyl-CoA availability, and robust pentose phosphate pathway. This study aimed to engineer for resveratrol production. The resveratrol biosynthetic pathway was integrated into by adding genes encoding tyrosine ammonia lyase from , 4-coumarate CoA ligase from , and stilbene synthase from . This resulted in the production of 14.3 mg/L resveratrol. A combination of endogenous and exogenous malonyl-CoA biosynthetic modules was introduced to enhance malonyl-CoA availability. This included genes encoding acetyl-CoA carboxylase 2 from , malonyl-CoA synthase, and a malonate transporter protein from . These strategies increased resveratrol production to 51.8 mg/L. The further optimization of fermentation conditions and the utilization of sucrose as an effective carbon source in YP media enhanced the resveratrol concentration to 141 mg/L in flask fermentation. By combining these strategies, we achieved a titer of 400 mg/L resveratrol in a controlled fed-batch bioreactor. These findings demonstrate the efficacy of as a platform for the de novo production of resveratrol and highlight the importance of metabolic engineering, enhancing malonyl-CoA availability, and media optimization for improved resveratrol production.
Topics: Resveratrol; Yarrowia; Metabolic Engineering; Sucrose; Acyltransferases; Vitis; Coenzyme A Ligases; Malonyl Coenzyme A; Nicotiana; Rhodotorula; Fermentation; Arabidopsis; Ammonia-Lyases; Bacterial Proteins
PubMed: 38927115
DOI: 10.3390/biom14060712 -
Journal of Fungi (Basel, Switzerland) Apr 2024Sodium butyrate (SB) is a histone deacetylase inhibitor that can induce changes in gene expression and secondary metabolite titers by inhibiting histone deacetylation....
Sodium butyrate (SB) is a histone deacetylase inhibitor that can induce changes in gene expression and secondary metabolite titers by inhibiting histone deacetylation. Our preliminary analysis also indicated that SB significantly enhanced the biosynthesis of carotenoids in the strain YM25079, although the underlying regulatory mechanisms remained unclear. Based on an integrated analysis of transcriptomics and metabolomics, this study revealed changes in cell membrane stability, DNA and protein methylation levels, amino acid metabolism, and oxidative stress in the strain YM25079 under SB exposure. Among them, the upregulation of oxidative stress may be a contributing factor for the increase in carotenoid biosynthesis, subsequently enhancing the strain resistance to oxidative stress and maintaining the membrane fluidity and function for normal cell growth. To summarize, our results showed that SB promoted carotenoid synthesis in the strain YM25079 and increased the levels of the key metabolites and regulators involved in the stress response of yeast cells. Additionally, epigenetic modifiers were applied to produce fungal carotenoid, providing a novel and promising strategy for the biosynthesis of yeast-based carotenoids.
PubMed: 38786675
DOI: 10.3390/jof10050320 -
International Journal of Biological... Jun 2024Rhodotorula spp. has been studied as one powerful source for a novel cell factory with fast growth and its high added-value biomolecules. However, its inadequate genome...
A chromosome-scale genome provides new insights into the typical carotenoid biosynthesis in the important red yeast Rhodotorula glutinis QYH-2023 with anti-inflammatory effects.
Rhodotorula spp. has been studied as one powerful source for a novel cell factory with fast growth and its high added-value biomolecules. However, its inadequate genome and genomic annotation have hindered its widespread use in cosmetics and food industries. Rhodotorula glutinis QYH-2023, was isolated from rice rhizosphere soil, and the highest quality of the genome of the strain was obtained at chromosome level (18 chromosomes) than ever before in red yeast in this study. Comparative genomics analysis revealed that there are more key gene copies of carotenoids biosynthesis in R. glutinis QYH-2023 than other species of Rhodotorula spp. Integrated transcriptome and metabolome analysis revealed that lipids and carotenoids biosynthesis was significantly enriched during fermentation. Subsequent investigation revealed that the over-expression of the strain three genes related to carotenoids biosynthesis in Komagataella phaffii significantly promoted the carotenoid production. Furthermore, in vitro tests initially confirmed that the longer the fermentation period, the synthesized metabolites controlled by R. glutinis QYH-2023 genome had the stronger anti-inflammatory properties. All of the findings revealed a high-quality reference genome which highlight the potential of R. glutinis strains to be employed as chassis cells for biosynthesizing carotenoids and other active chemicals.
Topics: Carotenoids; Rhodotorula; Genome, Fungal; Anti-Inflammatory Agents; Fermentation; Chromosomes, Fungal; Genomics; Transcriptome
PubMed: 38719011
DOI: 10.1016/j.ijbiomac.2024.132103 -
ACS Omega Apr 2024Two unprecedented isomeric secondary metabolites named vibralactones Z () and Z (), in addition to eleven known compounds (-), were isolated from solid-state rice...
Two unprecedented isomeric secondary metabolites named vibralactones Z () and Z (), in addition to eleven known compounds (-), were isolated from solid-state rice culture medium of (Bondarzewiaceae). Chemical structures of the isolated compounds were established via spectral analyses. The new lactone derivatives were weakly active against without any significant cytotoxicity, while the molecules containing an aldehyde functionality showed significant antimicrobial and cytotoxic effects. For instance, erinacine P () and (+)-isovelleral () and erinacine P () were cytotoxic against all tested cell lines at IC values in the ranges of 3.5-14.2 and 2.8-30.2 μM, respectively. In addition, they revealed moderate antimicrobial activity with the lowest minimum inhibitory concentration (MIC) values recorded against (8.3 μg/mL), , and at 16.6 μg/mL.
PubMed: 38680360
DOI: 10.1021/acsomega.4c02011 -
Frontiers in Chemistry 2024This study investigates the biological activities of essential oil (LPEO), an endemic lavender species from the Canary Islands, traditionally used in treating various...
This study investigates the biological activities of essential oil (LPEO), an endemic lavender species from the Canary Islands, traditionally used in treating various ailments. LPEO was extracted by hydrodistillation and analyzed using GC-MS. Antioxidant activity was assessed by DPPH radical scavenging and total antioxidant capacity assays. Antimicrobial activity was evaluated by disc diffusion, MIC, MBC, and MFC determination against bacterial () and fungal () strains. Antidiabetic and anti-gout potential were investigated through α-amylase, α-glucosidase, and xanthine oxidase inhibition assays. Antityrosinase activity was determined using a modified dopachrome method. Cytotoxicity was assessed by MTT assay against breast (MCF-7, MDA-MB-468), liver (HepG2), colon (HCT-15) cancer cells, and normal cells (PBMCs). LPEO exhibits potent antiradical activity (IC50 = 148.33 ± 2.48 μg/mL) and significant antioxidant capacity (TAC = 171.56 ± 2.34 μg AA/mg of EO). It demonstrates notable antibacterial activity against four strains ( and ) with inhibition zones ranging from 18.70 ± 0.30 mm to 29.20 ± 0.30 mm, along with relatively low MIC and MBC values. LPEO displays significant antifungal activity against four strains ( and ) with a fungicidal effect at 1 mg/mL, surpassing the positive control (cycloheximide), and MIC and MFC values indicating a fungicidal effect. It exhibits substantial inhibition of xanthine oxidase enzyme (IC50 = 26.48 ± 0.90 μg/mL), comparable to allopurinol, and marked inhibitory effects on α-amylase (IC50 = 31.56 ± 0.46 μg/mL) and α-glucosidase (IC50 = 58.47 ± 2.35 μg/mL) enzymes.The enzyme tyrosinase is inhibited by LPEO (IC50 = 29.11 ± 0.08 mg/mL). LPEO displays moderate cytotoxic activity against breast, liver, and colon cancer cells, with low toxicity towards normal cells (PBMC). LPEO exhibits greater selectivity than cisplatin for breast (MCF-7) and colon (HCT-15) cancer cells but lower selectivity for liver (HepG2) and metastatic breast (MDA-MB-468) cancer cells. These findings suggest the potential of LPEO as an antioxidant, antimicrobial, anti-gout, antidiabetic, and anticancer agent.
PubMed: 38660570
DOI: 10.3389/fchem.2024.1383731 -
Indian Journal of Medical Microbiology 2024Candida auris is increasingly being isolated from patients all over the world. It has five clades. In this study, it was aimed to compare the results of biochemical...
PURPOSE
Candida auris is increasingly being isolated from patients all over the world. It has five clades. In this study, it was aimed to compare the results of biochemical tests obtained using different methods and the antifungal susceptibility profiles of C. auris strains isolated from the first seven cases reported in Türkiye, and evaluate whether this information could be useful as preliminary data in determining the clade of strains in centers that lack the opportunity to apply molecular methods.
METHODS
Identification test results obtained using API ID 32 C, API 20 C AUX, VITEK-2 YST, and MALDI-TOF MS; colony color and morphology on Chromagar Candida, CHROMagar Candida Plus media, and cornmeal-Tween 80 agar; susceptibility to antifungals were tested and compared. Antifungal susceptibility test was studied using microdilution method according to the recommendations of EUCAST. Additionally, a pilot study was conducted to investigate the value of CHROMagar Candida Plus.
RESULTS
All seven strains were identified as Lachancea kluyveri with API ID 32 C, Rhodotorula glutinis; Cryptococcus neoformans with API 20 C AUX, and C. auris with both VITEK-2 YST and MALDI-TOF MS. MIC values for fluconazole were very high (≥64 mg/L) for all seven strains. It was observed that 11 (37.9%) of 29 Candida parapsilosis strains formed colonies with morphology similar to C. auris on CHROMagar Candida Plus medium, leading to false positivity.
CONCLUSIONS
Although there have been many isolations of C. auris in our country in recent years, clade distribution of only a small number of strains is known yet. In this study, when the biochemical properties and antifungal susceptibility profiles of the seven strains were evaluated, it was concluded that they exhibited some characteristics compatible with clade I. It was also observed that strains 1 and 2 may belong to a different clade.
Topics: Humans; Antifungal Agents; Microbial Sensitivity Tests; Candidiasis; Candida auris; Turkey; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; Mycological Typing Techniques; Candida; Male; Female
PubMed: 38636843
DOI: 10.1016/j.ijmmb.2024.100594 -
3 Biotech Jan 2024The present study aims to investigate the physicochemical characteristics of phenylalanine ammonia-lyase (PAL) extracted from agricultural waste and its potential use as...
The present study aims to investigate the physicochemical characteristics of phenylalanine ammonia-lyase (PAL) extracted from agricultural waste and its potential use as an anticancer agent in comparison to microbial PAL. We extracted and partially purified PAL from agricultural waste sources. We assessed the temperature and pH range of PAL and determined enzyme kinetics parameters including Michaelis constants (Km), maximum velocity (Vmax), and specificity constant values (Vmax/Km). Additionally, we examined the effects of different storage temperatures on PAL activity. In our analysis, we compared the efficacy of agricultural waste-derived PAL with PAL from . The results demonstrated that PAL extracted from agricultural waste exhibited significantly higher specific activity (Vmax/Km) compared to its microbial counterpart. The agricultural waste-derived PAL displayed a stronger affinity for phenylalanine, as indicated by a lower Km value than the microbial PAL did. Furthermore, PAL from agricultural waste maintained activity across a broader temperature and pH range (15-75 °C, pH 5-11), in contrast to microbial PAL (20-60 °C, pH 5.5-10). Importantly, the PAL derived from agricultural waste exhibited superior stability, retaining over 90% of its activity after 6 months of storage at room temperature (25 °C), whereas microbial PAL lost more than 70% of its activity under similar storage conditions. In anticancer experiments against various cancer cell lines, agricultural waste-derived PAL demonstrated greater anticancer activity compared to microbial PAL. These findings suggest that PAL sourced from agricultural waste has the potential to be a safe and effective natural anticancer agent.
PubMed: 38156037
DOI: 10.1007/s13205-023-03871-x -
Life (Basel, Switzerland) Oct 2023, acknowledged through its indigenous nomenclature "samar", is part of the Juncaceae taxonomic lineage, bearing considerable import as a botanical reservoir harboring...
, acknowledged through its indigenous nomenclature "samar", is part of the Juncaceae taxonomic lineage, bearing considerable import as a botanical reservoir harboring conceivable therapeutic attributes. Its historical precedence in traditional curative methodologies for the alleviation of infections and inflammatory conditions is notable. In the purview of Eastern traditional medicine, Juncus species seeds find application for their remedial efficacy in addressing diarrhea, while the botanical fruits are subjected to infusion processes targeting the attenuation of symptoms associated with cold manifestations. The primary objective of this study was to unravel the phytochemical composition of distinct constituents within , specifically leaves (JALE) and roots (JARE), originating from the indigenous expanse of the Nador region in northeastern Morocco. The extraction of plant constituents was executed utilizing an ethanol-based extraction protocol. The subsequent elucidation of chemical constituents embedded within the extracts was accomplished employing analytical techniques based on high-performance liquid chromatography (HPLC). For the purpose of in vitro antioxidant evaluation, a dual approach was adopted, encompassing the radical scavenging technique employing 2,2-diphenyl-1-picrylhydrazyl (DPPH) and the total antioxidant capacity (TAC) assay. The acquired empirical data showcase substantial radical scavenging efficacy and pronounced relative antioxidant activity. Specifically, the DPPH and TAC methods yielded values of 483.45 ± 4.07 µg/mL and 54.59 ± 2.44 µg of ascorbic acid (AA)/mL, respectively, for the leaf extracts. Correspondingly, the root extracts demonstrated values of 297.03 ± 43.3 µg/mL and 65.615 ± 0.54 µg of AA/mL for the DPPH and TAC methods. In the realm of antimicrobial evaluation, the assessment of effects was undertaken through the agar well diffusion technique. The minimum inhibitory concentration, minimum bactericidal concentration, and minimum fungicidal concentration were determined for each extract. The inhibitory influence of the ethanol extracts was observed across bacterial strains including , , and , with the notable exception of . However, fungal strains such as and exhibited comparatively lower resistance, whereas and exhibited heightened resistance, evincing negligible antifungal activity. An anticipatory computational assessment of pharmacokinetic parameters was conducted, complemented by the application of the Pro-tox II web tool to delineate the potential toxicity profile of compounds intrinsic to the studied extracts. The culmination of these endeavors underpins the conceivable prospects of the investigated extracts as promising candidates for oral medicinal applications.
PubMed: 38004275
DOI: 10.3390/life13112135 -
Applied Biochemistry and Biotechnology Nov 2023Β-Carotene is a red-orange pigment that serves as a precursor to important pharmaceutical molecules like vitamin A and retinol, making it highly significant in the...
Β-Carotene is a red-orange pigment that serves as a precursor to important pharmaceutical molecules like vitamin A and retinol, making it highly significant in the industrial sector. Consequently, there is an ongoing quest for more sustainable production methods. In this study, glucose and xylose, two primary sugars derived from sugarcane bagasse (SCB), were utilized as substrates for β-carotene production by Rhodotorula glutinis CCT-2186. To achieve this, SCB underwent pretreatment using NaOH, involved different concentrations of total solids (TS) (10%, 15%, and 20%) to remove lignin. Each sample was enzymatically hydrolyzed using two substrate loadings (5% and 10%). The pretreated SCB with 10%, 15%, and 20% TS exhibited glucose hydrolysis yields (%wt) of 93.10%, 91.88%, and 90.77%, respectively. The resulting hydrolysate was employed for β-carotene production under batch fermentation. After 72 h of fermentation, the SCB hydrolysate yielded a β-carotene concentration of 118.56 ± 3.01 mg/L. These findings showcase the robustness of R. glutinis as a biocatalyst for converting SCB into β-carotene.
PubMed: 37914962
DOI: 10.1007/s12010-023-04761-x -
BioRxiv : the Preprint Server For... Oct 2023Single-cell sorting is essential to explore cellular heterogeneity in biology and medicine. Recently developed Raman-activated cell sorting (RACS) circumvents the...
Single-cell sorting is essential to explore cellular heterogeneity in biology and medicine. Recently developed Raman-activated cell sorting (RACS) circumvents the limitations of fluorescence-activated cell sorting, such as the cytotoxicity of labels. However, the sorting throughputs of all forms of RACS are limited by the intrinsically small cross-section of spontaneous Raman scattering. Here, we report a stimulated Raman-activated cell ejection (S-RACE) platform that enables high-throughput single-cell sorting based on high-resolution multi-channel stimulated Raman chemical imaging, image decomposition, and laser-induced cell ejection. The performance of this platform was illustrated by sorting a mixture of 1 μm polymer beads, where 95% yield, 98% purity, and 14 events per second throughput were achieved. Notably, our platform allows live cell ejection, allowing for the growth of single colonies of bacteria and fungi after sorting. To further illustrate the chemical selectivity, lipid-rich cells were successfully sorted from a mixture with , confirmed by downstream quantitative PCR. Furthermore, by integrating a closed-loop feedback control circuit into the system, we realized real-time single-cell imaging and sorting, and applied this method to precisely eject regions of interest from a rat brain tissue section. The reported S-RACE platform opens exciting opportunities for a wide range of single-cell applications in biology and medicine.
PubMed: 37904930
DOI: 10.1101/2023.10.16.562526