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Food Chemistry: X Jun 2023Mixed fermentation using saccharomyces cerevisiae and non- has become one of the main research strategies to improve wine aroma. Hence, this study applied the mixed...
Mixed fermentation using saccharomyces cerevisiae and non- has become one of the main research strategies to improve wine aroma. Hence, this study applied the mixed fermentation technique using and to brew Cabernet Sauvignon wine and to investigate the effects of inoculation timing and inoculation ratio on the polyphenolics, antioxidant activity and aroma of the resulting wine. The results showed that mixed fermentation significantly improved the amounts of flavan-3-ols. In particular, S1:5 had the highest amounts of (-)-catechin and procyanidin B1 (73.23 mg/L and 46.59 mg/L), while S1:10 had the highest (-)-epicatechin content (57.95 mg/L). Meanwhile, S1:10 showed the strongest FRAP, CUPRAC and ABTS + activities (31.46 %, 25.38 % and 13.87 % higher than that of CK, respectively). In addition, mixed fermentation also increased the amounts of phenylethanol, isoamyl alcohol and ethyl esters, which enhanced the rose-like and fruity flavor of wine. This work used a friendly non- alongside appropriate inoculation strategies to provide an alternative approach for improved wine aroma and phenolic profile.
PubMed: 37131849
DOI: 10.1016/j.fochx.2023.100685 -
Microbial Cell Factories Jan 2024Chitinases are widely distributed enzymes that perform the biotransformation of chitin, one of the most abundant polysaccharides on the biosphere, into useful... (Review)
Review
BACKGROUND
Chitinases are widely distributed enzymes that perform the biotransformation of chitin, one of the most abundant polysaccharides on the biosphere, into useful value-added chitooligosaccharides (COS) with a wide variety of biotechnological applications in food, health, and agricultural fields. One of the most important group of enzymes involved in the degradation of chitin comprises the glycoside hydrolase family 18 (GH18), which harbours endo- and exo-enzymes that act synergistically to depolymerize chitin. The secretion of a chitinase activity from the ubiquitous yeast Mestchnikowia pulcherrima and their involvement in the post-harvest biological control of fungal pathogens was previously reported.
RESULTS
Three new chitinases from M. pulcherrima, MpChit35, MpChit38 and MpChit41, were molecularly characterized and extracellularly expressed in Pichia pastoris to about 91, 90 and 71 mU ml, respectively. The three enzymes hydrolysed colloidal chitin with optimal activity at 45 ºC and pH 4.0-4.5, increased 2-times their activities using 1 mM of Mn and hydrolysed different types of commercial chitosan. The partial separation and characterization of the complex COS mixtures produced from the hydrolysis of chitin and chitosan were achieved by a new anionic chromatography HPAEC-PAD method and mass spectrometry assays. An overview of the predicted structures of these proteins and their catalytic modes of action were also presented. Depicted their high sequence and structural homology, MpChit35 acted as an exo-chitinase producing di-acetyl-chitobiose from chitin while MpChit38 and MpChit41 both acted as endo-chitinases producing tri-acetyl-chitotriose as main final product.
CONCLUSIONS
Three new chitinases from the yeast M. pulcherrima were molecularly characterized and their enzymatic and structural characteristics analysed. These enzymes transformed chitinous materials to fully and partially acetylated COS through different modes of splitting, which make them interesting biocatalysts for deeper structural-function studies on the challenging enzymatic conversion of chitin.
Topics: Chitin; Chitinases; Chitosan; Proteins; Saccharomyces cerevisiae
PubMed: 38245740
DOI: 10.1186/s12934-024-02300-9 -
Life (Basel, Switzerland) Nov 2023The gut microbiome plays an important role in the metabolism, nutrient absorption and immunocompetency of animals. The dynamics of the microbiota can be influenced by...
The gut microbiome plays an important role in the metabolism, nutrient absorption and immunocompetency of animals. The dynamics of the microbiota can be influenced by modulatory factors that involve nutrition, environment, health, diseases, etc. Few reports have been documented regarding the effects of different feeds on the fungi microbiome of Suffolk crossed with Tibetan sheep. A total of 30 Suffolk crossed with Tibetan sheep (ST sheep) were selected for the study and randomly divided into five equal groups ( = 6): AZ, BZ, CZ, DZ and EZ. Group AZ was fed with alfalfa and oat grass, whereas group BZ was fed with mixture of concentrated feed, alfalfa and oat grass. Groups CZ, DZ and EZ were fed with concentrated feed #1, #2 and #3, respectively. All experimental animals were fed twice a day for four months, and rectum samples were collected for microbiota analysis. Results revealed that 2,781,461 raw reads and 2,333,239 clean reads were achieved in the ST sheep. When compared with the sheep of groups AZ and BZ (164), the shared amplicon sequence variants (ASVs) between AZ and CZ (109), AZ (113) and DZ (118) as well as AZ along with EZ were fewer. Conspicuous different phyla (8) and genera (56) were examined and compared with free-range sheep in AZ. Genera including , , , , , etc. were found higher in animals in the CZ, DZ and EZ groups. The results of this study provide new insights regarding the effects of different feeds on the fungi microbiome of sheep farmed on the plateau. We concluded that the differences in feed in Suffolk crossed with Tibetan sheep altered their gut microbiota.
PubMed: 38004350
DOI: 10.3390/life13112210 -
Applied and Environmental Microbiology Oct 2023Lytic polysaccharide monooxygenases (LPMOs) can perform oxidative cleavage of glycosidic bonds in carbohydrate polymers (e.g., cellulose, chitin), making them more...
Lytic polysaccharide monooxygenases (LPMOs) can perform oxidative cleavage of glycosidic bonds in carbohydrate polymers (e.g., cellulose, chitin), making them more accessible to hydrolytic enzymes. While most studies have so far mainly explored the role of LPMOs in a (plant) biomass conversion context, alternative roles and paradigms begin to emerge. The AA10 LPMOs are active on chitin and/or cellulose and mostly found in bacteria and in some viruses and archaea. Interestingly, AA10-encoding genes are also encountered in some pathogenic fungi of the Ustilaginomycetes class, such as , responsible for corn smut disease. Transcriptomic studies have shown the overexpression of the AA10 gene during the infectious cycle of . In fact, has a unique AA10 gene that codes for a catalytic domain appended with a C-terminal disordered region. To date, there is no public report on fungal AA10 LPMOs. In this study, we successfully produced the catalytic domain of this LPMO (AA10_cd) in and carried out its biochemical characterization. Our results show that AA10_cd oxidatively cleaves α- and β-chitin with C1 regioselectivity and boosts chitin hydrolysis by a GH18 chitinase from (GH18A). Using a biologically relevant substrate, we show that AA10_cd exhibits enzymatic activity on fungal cell wall chitin and promotes its hydrolysis by GH18A. These results represent an important step toward the understanding of the role of LPMOs in the fungal cell wall remodeling process during the fungal life cycle.IMPORTANCELytic polysaccharide monooxygenases (LPMOs) have been mainly studied in a biotechnological context for the efficient degradation of recalcitrant polysaccharides. Only recently, alternative roles and paradigms begin to emerge. In this study, we provide evidence that the AA10 LPMO from the phytopathogen is active against fungal cell wall chitin. Given that chitin-active LPMOs are commonly found in microbes, it is important to consider fungal cell wall as a potential target for this enigmatic class of enzymes.
Topics: Chitin; Polysaccharides; Mixed Function Oxygenases; Cellulose; Cell Wall
PubMed: 37702503
DOI: 10.1128/aem.00573-23 -
Heliyon Sep 2023Profiling the endophytic microbiome of different tissues and varieties of agricultural crops can help to understand i) the tissue specific and varietal specific microbes...
Profiling the endophytic microbiome of different tissues and varieties of agricultural crops can help to understand i) the tissue specific and varietal specific microbes associated with the plants ii) their potential role in growth, stress tolerance, disease resistance, and yield of the plants. Comparative microbiome profiling across various varieties and hybrids will also be useful to identify the plant's core microbiome. The main objective of the work is to profile and study the microbiome of traditional varieties in comparison with hybrids of cucumber, which would help to understand the microbiome structure in developing consortia to engineer the microbiome of modern hybrids, for useful phenotypes. Metataxonomic sequencing of bacteria and fungi using 16S rRNA gene and ITS regions respectively were carried out in seed and leaf samples of cucumber traditional varieties and modern hybrids. Among bacteria, and and among fungal genera, and belonged to the core microbiome of cucumber. Modern hybrids were rich in antibiotic producing and toxic pollutant degrading bacteria. Many of the fungi and bacteria observed in the study are well known plant growth promoting microorganisms and play role in offering disease resistance. Some of the bacteria and fungi have beneficial roles in human gut thus revealing the dietary importance of cucumber. The microbes identified in the current study will be useful starting point to develop a consortia to engineer the cucumber microbiome for growth, yield and stress tolerance traits.
PubMed: 37809962
DOI: 10.1016/j.heliyon.2023.e20216 -
Virulence Dec 2024(APP) is an important pathogen of the porcine respiratory disease complex, which leads to huge economic losses worldwide. We previously demonstrated that -producing...
(APP) is an important pathogen of the porcine respiratory disease complex, which leads to huge economic losses worldwide. We previously demonstrated that -producing bovine neutrophil β-defensin-5 (B5) could resist the infection by the bovine intracellular pathogen . In this study, the roles of synthetic B5 in regulating mucosal innate immune response and protecting against extracellular APP infection were further investigated using a mouse model. Results showed that B5 promoted the production of tumour necrosis factor (TNF)-α, interleukin (IL)-1β, and interferon (IFN)-β in macrophages as well as dendritic cells (DC) and enhanced DC maturation . Importantly, intranasal B5 was safe and conferred effective protection against APP via reducing the bacterial load in lungs and alleviating pulmonary inflammatory damage. Furthermore, in the early stage of APP infection, we found that intranasal B5 up-regulated the secretion of TNF-α, IL-1β, IL-17, and IL-22; enhanced the rapid recruitment of macrophages, neutrophils, and DC; and facilitated the generation of group 3 innate lymphoid cells in lungs. In addition, B5 activated signalling pathways associated with cellular response to IFN-β and activation of innate immune response in APP-challenged lungs. Collectively, B5 via the intranasal route can effectively ameliorate the immune suppression caused by early APP infection and provide protection against APP. The immunization strategy may be applied to animals or human respiratory bacterial infectious diseases. Our findings highlight the potential importance of B5, enhancing mucosal defence against intracellular bacteria like APP which causes early-phase immune suppression.
Topics: Humans; Swine; Animals; Cattle; Immunity, Innate; Actinobacillus pleuropneumoniae; Lymphocytes; Lung; Tumor Necrosis Factor-alpha; Immunosuppression Therapy
PubMed: 38378464
DOI: 10.1080/21505594.2024.2316459 -
Journal of Trace Elements in Medicine... May 2024Yeasts have the remarkable capability to transform and integrate inorganic selenium into their cellular structures, thereby enhancing its bioavailability and reducing...
BACKGROUND AND OBJECTIVE
Yeasts have the remarkable capability to transform and integrate inorganic selenium into their cellular structures, thereby enhancing its bioavailability and reducing its toxicity. In recent years, yeasts have attracted attention as potential alternative sources of protein.
METHODS
This study explores the selenium accumulation potential of two less explored yeast strains, namely the probiotic Saccharomyces boulardii CCDM 2020 and Pichia fermentas CCDM 2012, in comparison to the extensively studied Saccharomyces cerevisiae CCDM 272. Our investigation encompassed diverse stress conditions. Subsequently, the selenized yeasts were subjected to an INFOGEST gastrointestinal model. The adherence and hydrophobicity were determined with undigested cells RESULTS: Stress conditions had an important role in influencing the quantity and size of selenium nanoparticles (SeNPs) generated by the tested yeasts. Remarkably, SeMet synthesis was limited to Pichia fermentas CCDM 2012 and S. boulardii CCDM 2020, with S. cerevisiae CCDM 272 not displaying SeMet production at all. Throughout the simulated gastrointestinal digestion, the most substantial release of SeCys2, SeMet, and SeNPs from the selenized yeasts occurred during the intestinal phase. Notably, exception was found in strain CCDM 272, where the majority of particles were released during the oral phase.
CONCLUSION
The utilization of both traditional and non-traditional selenized yeast types, harnessed for their noted functional attributes, holds potential for expanding the range of products available while enhancing their nutritional value and health benefits.
Topics: Saccharomyces cerevisiae; Saccharomyces boulardii; Pichia; Selenium; Probiotics; Digestion
PubMed: 38310829
DOI: 10.1016/j.jtemb.2024.127402 -
Synthetic and Systems Biotechnology Sep 2023Methanol has recently gained significant attention as a potential carbon substrate for the production of fuels and chemicals, owing to its high degree of reduction,...
Methanol has recently gained significant attention as a potential carbon substrate for the production of fuels and chemicals, owing to its high degree of reduction, abundance, and low price. Native methylotrophic yeasts and bacteria have been investigated for the production of fuels and chemicals. Alternatively, synthetic methylotrophic strains are also being developed by reconstructing methanol utilization pathways in model microorganisms, such as Owing to the complex metabolic pathways, limited availability of genetic tools, and methanol/formaldehyde toxicity, the high-level production of target products for industrial applications are still under development to satisfy commercial feasibility. This article reviews the production of biofuels and chemicals by native and synthetic methylotrophic microorganisms. It also highlights the advantages and limitations of both types of methylotrophs and provides an overview of ways to improve their efficiency for the production of fuels and chemicals from methanol.
PubMed: 37384124
DOI: 10.1016/j.synbio.2023.06.001 -
Molecules (Basel, Switzerland) Aug 2023Glycosylation is an important post-translational modification of proteins, contributing to protein function, stability and subcellular localization. Fungal...
Glycosylation is an important post-translational modification of proteins, contributing to protein function, stability and subcellular localization. Fungal immunomodulatory proteins (FIPs) are a group of small proteins with notable immunomodulatory activity, some of which are glycoproteins. In this study, the impact of glycosylation on the bioactivity and biochemical characteristics of FIP-nha (from ) is described. Three rFIP-nha glycan mutants (N5A, N39A, N5+39A) were constructed and expressed in to study the functionality of the specific N-glycosylation on amino acid N5 and N39. Their protein characteristics, structure, stability and activity were tested. WT and mutants all formed tetramers, with no obvious difference in crystal structures. Their melting temperatures were 82.2 °C (WT), 81.4 °C (N5A), 80.7 °C (N39A) and 80.1 °C (N5+39A), indicating that glycosylation improves thermostability of rFIP-nha. Digestion assays showed that glycosylation on either site improved pepsin resistance, while 39N-glycosylation was important for trypsin resistance. Based on the 3D structure and analysis of enzyme cleavage sites, we conclude that glycosylation might interfere with hydrolysis via increasing steric hindrance. WT and mutants exerted similar bioactivity on tumor cell metabolism and red blood cells hemagglutination. Taken together, these findings indicate that glycosylation of FIP-nha impacts its thermostability and digestion resistance.
Topics: Glycosylation; Proteolysis; Peptide Hydrolases; Fusarium; Fungal Proteins
PubMed: 37687215
DOI: 10.3390/molecules28176386 -
Microbial Cell Factories Aug 2023Recently, researchers have focused on the search for alternatives to conventional antibiotics. Antimicrobial peptides are small bioactive peptides that regulate immune...
BACKGROUND
Recently, researchers have focused on the search for alternatives to conventional antibiotics. Antimicrobial peptides are small bioactive peptides that regulate immune activation and have antibacterial activity with a reduced risk of bacterial resistance. Porcine myeloid antibacterial peptide 37 (PMAP-37) is a small-molecule peptide with broad-spectrum antibacterial activity isolated from pig bone marrow, and PMAP-37(F34-R) is its analogue. In this study, PMAP-37(F34-R) was recombinantly expressed in Pichia pastoris, and the recombinant peptide was further investigated for its antibacterial properties, mechanism and preservative in plums.
RESULTS
To obtain a Pichia pastoris strain expressing PMAP-37(F34-R), we constructed a plasmid expressing recombinant PMAP-37(F34-R) (pPICZα-PMAP-37(F34-R)-A) and introduced it into Pichia pastoris. Finally, we obtained a highly active recombinant peptide, PMAP-37(F34-R), which inhibited the activity of both Gram-positive and Gram-negative bacteria. The minimum inhibitory concentration is 0.12-0.24 µg/mL, and it can destroy the integrity of the cell membrane, leading to cell lysis. It has good stability and is not easily affected by the external environment. Hemolysis experiments showed that 0.06 µg/mL-0.36 µg/mL PMAP-37(F34-R) had lower hemolysis ability to mammalian cells, and the hemolysis rate was below 1.5%. Additionally, 0.36 µg/mL PMAP-37(F34-R) showed a good preservative effect in plums. The decay and weight loss rates of the treated samples were significantly lower than those of the control group, and the respiratory intensity of the fruit was delayed in the experimental group.
CONCLUSIONS
In this study, we constructed a recombinant Pichia pastoris strain, which is a promising candidate for extending the shelf life of fruits and has potential applications in the development of new preservatives.
Topics: Animals; Swine; Prunus domestica; Anti-Bacterial Agents; Hemolysis; Gram-Negative Bacteria; Gram-Positive Bacteria; Antimicrobial Cationic Peptides; Bacteria; Mammals
PubMed: 37635252
DOI: 10.1186/s12934-023-02164-5