scholarly journals Disruption of protease A and B orthologous genes in the basidiomycetous yeast Pseudozyma antarctica GB-4(0) yields a stable extracellular biodegradable plastic-degrading enzyme

PLoS ONE ◽  
2021 ◽  
Vol 16 (3) ◽  
pp. e0247462
Author(s):  
Natsuki Omae ◽  
Yuka Sameshima-Yamashita ◽  
Kazunori Ushimaru ◽  
Hideaki Koike ◽  
Hiroko Kitamoto ◽  
...  
Keyword(s):  
Large Scale ◽  
Amino Acid Sequences ◽  
Biodegradable Plastic ◽  
Orthologous Genes ◽  
Degrading Enzyme ◽  
Pseudozyma Antarctica ◽  
Overexpression Strain ◽  
Genes Encoding ◽  
Large Scale Cultivation ◽  
And Storage

The yeast Pseudozyma antarctica (currently designated Moesziomyces antarcticus) secretes a xylose-induced biodegradable plastic-degrading enzyme (PaE). To suppress degradation of PaE during production and storage, we targeted the inhibition of proteolytic enzyme activity in P. antarctica. Proteases A and B act as upper regulators in the proteolytic network of the model yeast, Saccharomyces cerevisiae. We searched for orthologous genes encoding proteases A and B in the genome of P. antarctica GB-4(0) based on the predicted amino acid sequences. We found two gene candidates, PaPRO1 and PaPRO2, with conserved catalytically important domains and signal peptides indicative of vacuolar protease function. We then prepared gene-deletion mutants of strain GB-4(0), ΔPaPRO1 and ΔPaPRO2, and evaluated PaE stability in culture by immunoblotting analysis. Both mutants exhibited sufficient production of PaE without degradation fragments, while the parent strain exhibited the degradation fragments. Therefore, we concluded that the protease A and B orthologous genes are related to the degradation of PaE. To produce a large quantity of PaE, we made a PaPRO2 deletion mutant of a PaE-overexpression strain named XG8 by introducing a PaE high-production cassette into the strain GB-4(0). The ΔPaPRO2 mutant of XG8 was able to produce PaE without the degradation fragments during large-scale cultivation in a 3-L jar fermenter for 3 days at 30°C. After terminating the agitation, the PaE activity in the XG8 ΔPaPRO2 mutant culture was maintained for the subsequent 48 h incubation at 25°C regardless of remaining cells, while activity in the XG8 control was reduced to 55.1%. The gene-deleted mutants will be useful for the development of industrial processes of PaE production and storage.

Biodegradable plastic-degrading enzyme from Pseudozyma antarctica: cloning, sequencing, and characterization

2012 ◽  
Vol 97 (7) ◽  
pp. 2951-2959 ◽  
Author(s):  
Yukiko Shinozaki ◽  
Tomotake Morita ◽  
Xiao-hong Cao ◽  
Shigenobu Yoshida ◽  
Motoo Koitabashi ◽  
...  
Keyword(s):  
Biodegradable Plastic ◽  
Degrading Enzyme ◽  
Pseudozyma Antarctica

scholarly journals Comparative Nutrigenomics Analysis of the Pig, Mouse and Human (P15-004-19)

2019 ◽  
Vol 3 (Supplement_1) ◽  
Author(s):  
Harry Dawson ◽  
Celine Chen ◽  
Thomas Wang ◽  
Joseph Urban
Keyword(s):  
Large Scale ◽  
Great Majority ◽  
Cell Types ◽  
Nutrient Sensing ◽  
Orthologous Genes ◽  
Funding Sources ◽  
Large Scale Analysis ◽  
Genes Encoding ◽  
Superfamily Analysis ◽  
Unique Genes

Abstract Objectives The seminal work of Miller and Ullrey examined the suitability of the pig as a model for human nutrition concluded “With the possible exception of nonhuman primates, it is apparent that the omnivorous pig is one of the best models for study of nutrition issues in the omnivorous human.” To date, a cross-species, large-scale analysis of genes related to nutrition and metabolism has not been reported. Our goal was to systematically review similarities and differences in nutrition-related physiology, and where different, to explore possible etiologies behind each phenomena using comparative genomics. Methods A broad literature and laboratory-based analysis was conducted comparing 1532 genes associated with porcine, murine, and human macro and micronutrient metabolism, including metalloproteins. Four questions were addressed. Are genes in specific pathways conserved? Are the genes/proteins structurally and functionally similar? Are the genes expressed in similar cell types? Are the genes regulated in a similar manner to stimuli? Results Pigs have roughly 4-fold fewer unique genes (66) than the mouse (240) and human (209). The great majority of these unique genes were zinc-containing members of the KRAB-A box Transcription Factor Superfamily. Analysis of 142 non-orthologous genes revealed that these genes were 10 times more likely to be present in only pigs and humans (120) than only in mice and humans (17). Genes involved in vitamin A and lipid metabolism were more highly conserved between pigs and humans. Notable, differences were found between humans and pigs in regard to genes encoding digestive enzymes and nutrient sensing genes. In some cases, mechanistic data were obtained to explain for previously described differences in physiology. For example, the lack of porcine salivary lipase and amylase activities is likely related to the absence of these genes in the pig. Analysis of 888 orthologous genes indicated a greater pig-human protein similarity for almost every gene examined. Conclusions Overall, the genomic and physiological parameters examined were more similar between pigs and humans than mice and humans. This supports the proposition that evaluating nutrition in pigs provides data that is more physiologically relevant to humans. Funding Sources Supported by USDA/ARS Project Plan 1235-51,000-055-00D.

Production of a biodegradable plastic-degrading enzyme from cheese whey by the phyllosphere yeast Pseudozyma antarctica GB-4(1)W

2014 ◽  
Vol 118 (2) ◽  
pp. 183-187 ◽  
Author(s):  
Takashi Watanabe ◽  
Yukiko Shinozaki ◽  
Ken Suzuki ◽  
Motoo Koitabashi ◽  
Shigenobu Yoshida ◽  
...  
Keyword(s):  
Cheese Whey ◽  
Biodegradable Plastic ◽  
Degrading Enzyme ◽  
Pseudozyma Antarctica

scholarly journals Flavonoid-Modifying Capabilities of the Human Gut Microbiome—An In Silico Study

Nutrients ◽  
2021 ◽  
Vol 13 (8) ◽  
pp. 2688
Author(s):  
Tobias Goris ◽  
Rafael R. C. Cuadrat ◽  
Annett Braune
Keyword(s):  
Gut Microbiome ◽  
Large Scale ◽  
Health Impact ◽  
Gut Bacteria ◽  
Human Gut ◽  
Positive Health ◽  
Human Gut Microbiome ◽  
Nutritional Recommendations ◽  
Genes Encoding ◽  
A Minor

Flavonoids are a major group of dietary plant polyphenols and have a positive health impact, but their modification and degradation in the human gut is still widely unknown. Due to the rise of metagenome data of the human gut microbiome and the assembly of hundreds of thousands of bacterial metagenome-assembled genomes (MAGs), large-scale screening for potential flavonoid-modifying enzymes of human gut bacteria is now feasible. With sequences of characterized flavonoid-transforming enzymes as queries, the Unified Human Gastrointestinal Protein catalog was analyzed and genes encoding putative flavonoid-modifying enzymes were quantified. The results revealed that flavonoid-modifying enzymes are often encoded in gut bacteria hitherto not considered to modify flavonoids. The enzymes for the physiologically important daidzein-to-equol conversion, well studied in Slackiaisoflavoniconvertens, were encoded only to a minor extent in Slackia MAGs, but were more abundant in Adlercreutzia equolifaciens and an uncharacterized Eggerthellaceae species. In addition, enzymes with a sequence identity of about 35% were encoded in highly abundant MAGs of uncultivated Collinsella species, which suggests a hitherto uncharacterized daidzein-to-equol potential in these bacteria. Of all potential flavonoid modification steps, O-deglycosylation (including derhamnosylation) was by far the most abundant in this analysis. In contrast, enzymes putatively involved in C-deglycosylation were detected less often in human gut bacteria and mainly found in Agathobacter faecis (formerly Roseburia faecis). Homologs to phloretin hydrolase, flavanonol/flavanone-cleaving reductase and flavone reductase were of intermediate abundance (several hundred MAGs) and mainly prevalent in Flavonifractor plautii. This first comprehensive insight into the black box of flavonoid modification in the human gut highlights many hitherto overlooked and uncultured bacterial genera and species as potential key organisms in flavonoid modification. This could lead to a significant contribution to future biochemical-microbiological investigations on gut bacterial flavonoid transformation. In addition, our results are important for individual nutritional recommendations and for biotechnological applications that rely on novel enzymes catalyzing potentially useful flavonoid modification reactions.

scholarly journals Arabidopsis Genes Essential for Seedling Viability: Isolation of Insertional Mutants and Molecular Cloning

Genetics ◽  
2001 ◽  
Vol 159 (4) ◽  
pp. 1765-1778
Author(s):  
Gregory J Budziszewski ◽  
Sharon Potter Lewis ◽  
Lyn Wegrich Glover ◽  
Jennifer Reineke ◽  
Gary Jones ◽  
...  
Keyword(s):  
Large Scale ◽  
Protein Translocation ◽  
Gene Families ◽  
Mutant Phenotype ◽  
Lethal Mutant ◽  
A Genome ◽  
Genes Encoding ◽  
High Level ◽  
Mutant Lines ◽  
Genome Scale

Abstract We have undertaken a large-scale genetic screen to identify genes with a seedling-lethal mutant phenotype. From screening ~38,000 insertional mutant lines, we identified >500 seedling-lethal mutants, completed cosegregation analysis of the insertion and the lethal phenotype for >200 mutants, molecularly characterized 54 mutants, and provided a detailed description for 22 of them. Most of the seedling-lethal mutants seem to affect chloroplast function because they display altered pigmentation and affect genes encoding proteins predicted to have chloroplast localization. Although a high level of functional redundancy in Arabidopsis might be expected because 65% of genes are members of gene families, we found that 41% of the essential genes found in this study are members of Arabidopsis gene families. In addition, we isolated several interesting classes of mutants and genes. We found three mutants in the recently discovered nonmevalonate isoprenoid biosynthetic pathway and mutants disrupting genes similar to Tic40 and tatC, which are likely to be involved in chloroplast protein translocation. Finally, we directly compared T-DNA and Ac/Ds transposon mutagenesis methods in Arabidopsis on a genome scale. In each population, we found only about one-third of the insertion mutations cosegregated with a mutant phenotype.

scholarly journals JA signal-mediated immunity of Dendrobium catenatum to necrotrophic Southern Blight pathogen

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Cong Li ◽  
Qiuyi Shen ◽  
Xiang Cai ◽  
Danni Lai ◽  
Lingshang Wu ◽  
...  
Keyword(s):  
Large Scale ◽  
Plant Immunity ◽  
Expression Patterns ◽  
Protein Domain ◽  
Functional Identification ◽  
Necrotrophic Pathogen ◽  
Southern Blight ◽  
Meja Treatment ◽  
Sclerotium Delphinii ◽  
Large Scale Cultivation

Abstract Background Dendrobium catenatum belongs to the Orchidaceae, and is a precious Chinese herbal medicine. In the past 20 years, D. catenatum industry has developed from an endangered medicinal plant to multi-billion dollar grade industry. The necrotrophic pathogen Sclerotium delphinii has a devastating effection on over 500 plant species, especially resulting in widespread infection and severe yield loss in the process of large-scale cultivation of D. catenatum. It has been widely reported that Jasmonate (JA) is involved in plant immunity to pathogens, but the mechanisms of JA-induced plant resistance to S. delphinii are unclear. Results In the present study, the role of JA in enhancing D. catenatum resistance to S. delphinii was investigated. We identified 2 COI1, 13 JAZ, and 12 MYC proteins in D. catenatum genome. Subsequently, systematic analyses containing phylogenetic relationship, gene structure, protein domain, and motif architecture of core JA pathway proteins were conducted in D. catenatum and the newly characterized homologs from its closely related orchid species Phalaenopsis equestris and Apostasia shenzhenica, along with the well-investigated homologs from Arabidopsis thaliana and Oryza sativa. Public RNA-seq data were investigated to analyze the expression patterns of D. catenatum core JA pathway genes in various tissues and organs. Transcriptome analysis of MeJA and S. delphinii treatment showed exogenous MeJA changed most of the expression of the above genes, and several key members, including DcJAZ1/2/5 and DcMYC2b, are involved in enhancing defense ability to S. delphinii in D. catenatum. Conclusions The findings indicate exogenous MeJA treatment affects the expression level of DcJAZ1/2/5 and DcMYC2b, thereby enhancing D. catenatum resistance to S. delphinii. This research would be helpful for future functional identification of core JA pathway genes involved in breeding for disease resistance in D. catenatum.

scholarly journals Alterations of mesenchymal stromal cells in cerebrospinal fluid: insights from transcriptomics and an ALS clinical trial

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ashley A. Krull ◽  
Deborah O. Setter ◽  
Tania F. Gendron ◽  
Sybil C. L. Hrstka ◽  
Michael J. Polzin ◽  
...  
Keyword(s):  
Gene Expression ◽  
Cerebrospinal Fluid ◽  
Growth Factors ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Large Scale ◽  
Therapeutic Benefit ◽  
Protein Levels ◽  
Genes Encoding ◽  
Intrathecal Space

Abstract Background Mesenchymal stromal cells (MSCs) have been studied with increasing intensity as clinicians and researchers strive to understand the ability of MSCs to modulate disease progression and promote tissue regeneration. As MSCs are used for diverse applications, it is important to appreciate how specific physiological environments may stimulate changes that alter the phenotype of the cells. One need for neuroregenerative applications is to characterize the spectrum of MSC responses to the cerebrospinal fluid (CSF) environment after their injection into the intrathecal space. Mechanistic understanding of cellular biology in response to the CSF environment may predict the ability of MSCs to promote injury repair or provide neuroprotection in neurodegenerative diseases. Methods In this study, we characterized changes in morphology, metabolism, and gene expression occurring in human adipose-derived MSCs cultured in human (hCSF) or artificial CSF (aCSF) as well as examined relevant protein levels in the CSF of subjects treated with MSCs for amyotrophic lateral sclerosis (ALS). Results Our results demonstrated that, under intrathecal-like conditions, MSCs retained their morphology, though they became quiescent. Large-scale transcriptomic analysis of MSCs revealed a distinct gene expression profile for cells cultured in aCSF. The aCSF culture environment induced expression of genes related to angiogenesis and immunomodulation. In addition, MSCs in aCSF expressed genes encoding nutritional growth factors to expression levels at or above those of control cells. Furthermore, we observed a dose-dependent increase in growth factors and immunomodulatory cytokines in CSF from subjects with ALS treated intrathecally with autologous MSCs. Conclusions Overall, our results suggest that MSCs injected into the intrathecal space in ongoing clinical trials remain viable and may provide a therapeutic benefit to patients.

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