scholarly journals Vanillic acid and methoxyhydroquinone production from guaiacyl units and related aromatic compounds using Aspergillus niger cell factories

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Ronnie J. M. Lubbers ◽  
Adiphol Dilokpimol ◽  
Paula A. Nousiainen ◽  
Răzvan C. Cioc ◽  
Jaap Visser ◽  
...  
Keyword(s):  
Aspergillus Niger ◽  
Metabolic Pathway ◽  
Vanillic Acid ◽  
Aromatic Compounds ◽  
Metabolic Pathways ◽  
Fungal Cell ◽  
Cell Factories ◽  
Genes Encoding ◽  
Pharmaceutical Industries ◽  
Guaiacyl Lignin

Abstract Background The aromatic compounds vanillin and vanillic acid are important fragrances used in the food, beverage, cosmetic and pharmaceutical industries. Currently, most aromatic compounds used in products are chemically synthesized, while only a small percentage is extracted from natural sources. The metabolism of vanillin and vanillic acid has been studied for decades in microorganisms and many studies have been conducted that showed that both can be produced from ferulic acid using bacteria. In contrast, the degradation of vanillin and vanillic acid by fungi is poorly studied and no genes involved in this metabolic pathway have been identified. In this study, we aimed to clarify this metabolic pathway in Aspergillus niger and identify the genes involved. Results Using whole-genome transcriptome data, four genes involved in vanillin and vanillic acid metabolism were identified. These include vanillin dehydrogenase (vdhA), vanillic acid hydroxylase (vhyA), and two genes encoding novel enzymes, which function as methoxyhydroquinone 1,2-dioxygenase (mhdA) and 4-oxo-monomethyl adipate esterase (omeA). Deletion of these genes in A. niger confirmed their role in aromatic metabolism and the enzymatic activities of these enzymes were verified. In addition, we demonstrated that mhdA and vhyA deletion mutants can be used as fungal cell factories for the accumulation of vanillic acid and methoxyhydroquinone from guaiacyl lignin units and related aromatic compounds. Conclusions This study provides new insights into the fungal aromatic metabolic pathways involved in the degradation of guaiacyl units and related aromatic compounds. The identification of the involved genes unlocks new potential for engineering aromatic compound-producing fungal cell factories.

scholarly journals A Library of Aspergillus niger Chassis Strains for Morphology Engineering Connects Strain Fitness and Filamentous Growth With Submerged Macromorphology

2022 ◽  
Vol 9 ◽  
Author(s):  
Timothy C. Cairns ◽  
Xiaomei Zheng ◽  
Claudia Feurstein ◽  
Ping Zheng ◽  
Jibin Sun ◽  
...  
Keyword(s):  
Aspergillus Niger ◽  
Dry Weight ◽  
Filamentous Growth ◽  
Strain Engineering ◽  
Culture Conditions ◽  
Cell Factory ◽  
Fungal Cell ◽  
Cell Factories ◽  
Pellet Formation ◽  
Conditional Expression

Submerged fermentation using filamentous fungal cell factories is used to produce a diverse portfolio of useful molecules, including food, medicines, enzymes, and platform chemicals. Depending on strain background and abiotic culture conditions, different macromorphologies are formed during fermentation, ranging from dispersed hyphal fragments to approximately spherical pellets several millimetres in diameter. These macromorphologies are known to have a critical impact on product titres and rheological performance of the bioreactor. Pilot productivity screens in different macromorphological contexts is technically challenging, time consuming, and thus a significant limitation to achieving maximum product titres. To address this bottleneck, we developed a library of conditional expression mutants in the organic, protein, and secondary metabolite cell factory Aspergillus niger. Thirteen morphology-associated genes transcribed during fermentation were placed via CRISPR-Cas9 under control of a synthetic Tet-on gene switch. Quantitative analysis of submerged growth reveals that these strains have distinct and titratable macromorphologies for use as chassis during strain engineering programs. We also used this library as a tool to quantify how pellet formation is connected with strain fitness and filamentous growth. Using multiple linear regression modelling, we predict that pellet formation is dependent largely on strain fitness, whereas pellet Euclidian parameters depend on fitness and hyphal branching. Finally, we have shown that conditional expression of the putative kinase encoding gene pkh2 can decouple fitness, dry weight, pellet macromorphology, and culture heterogeneity. We hypothesize that further analysis of this gene product and the cell wall integrity pathway in which it is embedded will enable more precise engineering of A. niger macromorphology in future.

scholarly journals Draft Genome Sequence of Phenol-Degrading Variovorax boronicumulans Strain HAB-30

2020 ◽  
Vol 9 (7) ◽  
Author(s):  
Fatma Azwani Abdul Aziz ◽  
Kenshi Suzuki ◽  
Ryota Moriuchi ◽  
Hideo Dohra ◽  
Yosuke Tashiro ◽  
...  
Keyword(s):  
Genome Sequence ◽  
Aromatic Compounds ◽  
Metabolic Pathways ◽  
Chemostat Culture ◽  
Phenol Degradation ◽  
Draft Genome ◽  
Draft Genome Sequence ◽  
Degradation Pathways ◽  
Content Type ◽  
Genes Encoding

We report the draft genome sequence of Variovorax boronicumulans strain HAB-30, which was isolated from a phenol-degrading chemostat culture. This strain contains genes encoding a multicomponent type of phenol hydroxylase, with degradation pathways for catechol and other aromatic compounds. The genome sequence will be useful for understanding the metabolic pathways involved in phenol degradation.

scholarly journals Enzymatic Hydrolysate of Cinnamon Waste Material as Feedstock for the Microbial Production of Carotenoids

2021 ◽  
Vol 18 (3) ◽  
pp. 1146
Author(s):  
Stefano Bertacchi ◽  
Stefania Pagliari ◽  
Chiara Cantù ◽  
Ilaria Bruni ◽  
Massimo Labra ◽  
...  
Keyword(s):  
Industrial Sector ◽  
Waste Material ◽  
Fungal Cell ◽  
Enzymatic Hydrolysate ◽  
Carbon And Nitrogen ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Separate Hydrolysis And Fermentation ◽  
Additional Processing ◽  
Microbial Cultivation

In the context of the global need to move towards circular economies, microbial cell factories can be employed thanks to their ability to use side-stream biomasses from the agro-industrial sector to obtain additional products. The valorization of residues allows for better and complete use of natural resources and, at the same time, for the avoidance of waste management to address our needs. In this work, we focused our attention on the microbial valorization of cinnamon waste material after polyphenol extraction (C-PEW) (Cinnamomum verum J.Presl), generally discarded without any additional processing. The sugars embedded in C-PEW were released by enzymatic hydrolysis, more compatible than acid hydrolysis with the subsequent microbial cultivation. We demonstrated that the yeast Rhodosporidium toruloides was able to grow and produce up to 2.00 (±0.23) mg/L of carotenoids in the resulting hydrolysate as a sole carbon and nitrogen source despite the presence of antimicrobial compounds typical of cinnamon. To further extend the potential of our finding, we tested other fungal cell factories for growth on the same media. Overall, these results are opening the possibility to develop separate hydrolysis and fermentation (SHF) bioprocesses based on C-PEW and microbial biotransformation to obtain high-value molecules.

scholarly journals A Transcriptomic Approach to the Metabolism of Tetrapyrrolic Photosensitizers in a Marine Annelid

Molecules ◽  
2021 ◽  
Vol 26 (13) ◽  
pp. 3924
Author(s):  
Maria Leonor Santos ◽  
Mariaelena D’Ambrosio ◽  
Ana P. Rodrigo ◽  
A. Jorge Parola ◽  
Pedro M. Costa
Keyword(s):  
Metabolic Pathways ◽  
Molecular Networks ◽  
Bile Pigments ◽  
Rna Seq ◽  
The Past ◽  
Wide Range ◽  
Genes Encoding ◽  
Organ Specific ◽  
Bright Green ◽  
Green Coloration

The past decade has seen growing interest in marine natural pigments for biotechnological applications. One of the most abundant classes of biological pigments is the tetrapyrroles, which are prized targets due their photodynamic properties; porphyrins are the best known examples of this group. Many animal porphyrinoids and other tetrapyrroles are produced through heme metabolic pathways, the best known of which are the bile pigments biliverdin and bilirubin. Eulalia is a marine Polychaeta characterized by its bright green coloration resulting from a remarkably wide range of greenish and yellowish tetrapyrroles, some of which have promising photodynamic properties. The present study combined metabolomics based on HPLC-DAD with RNA-seq transcriptomics to investigate the molecular pathways of porphyrinoid metabolism by comparing the worm’s proboscis and epidermis, which display distinct pigmentation patterns. The results showed that pigments are endogenous and seemingly heme-derived. The worm possesses homologs in both organs for genes encoding enzymes involved in heme metabolism such as ALAD, FECH, UROS, and PPOX. However, the findings also indicate that variants of the canonical enzymes of the heme biosynthesis pathway can be species- and organ-specific. These differences between molecular networks contribute to explain not only the differential pigmentation patterns between organs, but also the worm’s variety of novel endogenous tetrapyrrolic compounds.

scholarly journals A squalene–hopene cyclase in Schizosaccharomyces japonicus represents a eukaryotic adaptation to sterol-limited anaerobic environments

2021 ◽  
Vol 118 (32) ◽  
pp. e2105225118
Author(s):  
Jonna Bouwknegt ◽  
Sanne J. Wiersma ◽  
Raúl A. Ortiz-Merino ◽  
Eline S. R. Doornenbal ◽  
Petrik Buitenhuis ◽  
...  
Keyword(s):  
Unsaturated Fatty Acids ◽  
Yeast Species ◽  
Early Report ◽  
Anaerobic Growth ◽  
Fungal Cell ◽  
Putative Gene ◽  
Cell Factories ◽  
Lipid Fractions ◽  
Synthetic Media ◽  
Free Media

Biosynthesis of sterols, which are key constituents of canonical eukaryotic membranes, requires molecular oxygen. Anaerobic protists and deep-branching anaerobic fungi are the only eukaryotes in which a mechanism for sterol-independent growth has been elucidated. In these organisms, tetrahymanol, formed through oxygen-independent cyclization of squalene by a squalene–tetrahymanol cyclase, acts as a sterol surrogate. This study confirms an early report [C. J. E. A. Bulder, Antonie Van Leeuwenhoek, 37, 353–358 (1971)] that Schizosaccharomyces japonicus is exceptional among yeasts in growing anaerobically on synthetic media lacking sterols and unsaturated fatty acids. Mass spectrometry of lipid fractions of anaerobically grown Sch. japonicus showed the presence of hopanoids, a class of cyclic triterpenoids not previously detected in yeasts, including hop-22(29)-ene, hop-17(21)-ene, hop-21(22)-ene, and hopan-22-ol. A putative gene in Sch. japonicus showed high similarity to bacterial squalene–hopene cyclase (SHC) genes and in particular to those of Acetobacter species. No orthologs of the putative Sch. japonicus SHC were found in other yeast species. Expression of the Sch. japonicus SHC gene (Sjshc1) in Saccharomyces cerevisiae enabled hopanoid synthesis and stimulated anaerobic growth in sterol-free media, thus indicating that one or more of the hopanoids produced by SjShc1 could at least partially replace sterols. Use of hopanoids as sterol surrogates represents a previously unknown adaptation of eukaryotic cells to anaerobic growth. The fast anaerobic growth of Sch. japonicus in sterol-free media is an interesting trait for developing robust fungal cell factories for application in anaerobic industrial processes.

IDENTIFYING RELEVANT PATHWAYS AND BIOMARKERS IN CROHN’S DISEASE USING CONTEXTUALIZED METABOLIC NETWORK MODEL

2021 ◽  
Vol 27 (Supplement_1) ◽  
pp. S9-S10
Author(s):  
Brooklyn McGrew ◽  
Aman Shrivastava ◽  
Philip Fernandes ◽  
Lubaina Ehsan ◽  
Yash Sharma ◽  
...  
Keyword(s):  
Gene Expression ◽  
Crohn’S Disease ◽  
Fatty Acid ◽  
Crohn's Disease ◽  
Fatty Acid Synthesis ◽  
Metabolic Pathway ◽  
Metabolic Pathways ◽  
Acid Synthesis ◽  
Transcriptomic Data ◽  
Context Specific

Abstract Background Candidate markers for Crohn’s Disease (CD) may be identified via gene expression-based construction of metabolic networks (MN). These can computationally describe gene-protein-reaction associations for entire tissues and also predict the flux of reactions (rate of turnover of specific molecules via a metabolic pathway). Recon3D is the most comprehensive human MN to date. We used publicly available CD transcriptomic data along with Recon3D to identify metabolites as potential diagnostic and prognostic biomarkers. Methods Terminal ileal gene expression profiles (36,372 genes; 218 CD. 42 controls) from the RISK cohort (Risk Stratification and Identification of Immunogenetic and Microbial Markers of Rapid Disease Progression in Children with Crohn’s Disease) and their transcriptomic abundances were used. Recon3D was pruned to only include RISK dataset transcripts which determined metabolic reaction linkage with transcriptionally active genes. Flux balance analysis (FBA) was then run using RiPTiDe with context specific transcriptomic data to further constrain genes (Figure 1). RiPTiDe was independently run on transcriptomic data from both CD and controls. From the pruned and constricted MN obtained, reactions were extracted for further analysis. Results After applying the necessary constraints to modify Recon3D, 527 CD and 537 control reactions were obtained. Reaction comparison with a publicly available list of healthy small intestinal epithelial reactions (n=1282) showed an overlap of 80 CD and 84 control reactions. These were then further grouped based on their metabolic pathways. RiPTiDe identified context specific metabolic pathway activity without supervision and the percentage of forward, backward, and balanced reactions for each metabolic pathway (Figure 2). The metabolite concentrations in the small intestine was altered among CD patients. Notably, the citric acid cycle and malate-aspartate shuttle were affected, highlighting changes in mitochondrial metabolic pathways. This is illustrated by changes in the number of reactions at equilibrium between CD and control. Conclusions The results are relevant as cytosolic acetyl-CoA is needed for fatty acid synthesis and is obtained by removing citrate from the citric acid cycle. An intermediate removal from the cycle has significant cataplerotic effects. The malate-aspartate shuttle also allows electrons to move across the impermeable membrane in the mitochondria (fatty acid synthesis location). These findings are reported by previously published studies where gene expression for fatty acid synthesis is altered in CD patients along with mitochondrial metabolic pathway changes, resulting in altered cell homeostasis. In-depth analysis is currently underway with our work supporting the utility of potential metabolic biomarkers for CD diagnosis, management and improved care.

scholarly journals Nonribosomal peptides in fungal cell factories: from genome mining to optimized heterologous production

2019 ◽  
Vol 37 (8) ◽  
pp. 107449 ◽  
Author(s):  
Antoine Vassaux ◽  
Loïc Meunier ◽  
Micheline Vandenbol ◽  
Denis Baurain ◽  
Patrick Fickers ◽  
...  
Keyword(s):  
Genome Mining ◽  
Nonribosomal Peptides ◽  
Heterologous Production ◽  
Fungal Cell ◽  
Cell Factories

scholarly journals Comparative Transcriptome Analysis Shows Conserved Metabolic Regulation during Production of Secondary Metabolites in Filamentous Fungi

mSystems ◽  
2019 ◽  
Vol 4 (2) ◽  
Author(s):  
Jens Christian Nielsen ◽  
Sylvain Prigent ◽  
Sietske Grijseels ◽  
Mhairi Workman ◽  
Boyang Ji ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Secondary Metabolism ◽  
Secondary Metabolite ◽  
Filamentous Fungi ◽  
Metabolic Regulation ◽  
Fungal Species ◽  
Efficient Production ◽  
Fungal Cell ◽  
Fungal Metabolism ◽  
Cell Factories

ABSTRACTFilamentous fungi possess great potential as sources of medicinal bioactive compounds, such as antibiotics, but efficient production is hampered by a limited understanding of how their metabolism is regulated. We investigated the metabolism of six secondary metabolite-producing fungi of thePenicilliumgenus during nutrient depletion in the stationary phase of batch fermentations and assessed conserved metabolic responses across species using genome-wide transcriptional profiling. A coexpression analysis revealed that expression of biosynthetic genes correlates with expression of genes associated with pathways responsible for the generation of precursor metabolites for secondary metabolism. Our results highlight the main metabolic routes for the supply of precursors for secondary metabolism and suggest that the regulation of fungal metabolism is tailored to meet the demands for secondary metabolite production. These findings can aid in identifying fungal species that are optimized for the production of specific secondary metabolites and in designing metabolic engineering strategies to develop high-yielding fungal cell factories for production of secondary metabolites.IMPORTANCESecondary metabolites are a major source of pharmaceuticals, especially antibiotics. However, the development of efficient processes of production of secondary metabolites has proved troublesome due to a limited understanding of the metabolic regulations governing secondary metabolism. By analyzing the conservation in gene expression across secondary metabolite-producing fungal species, we identified a metabolic signature that links primary and secondary metabolism and that demonstrates that fungal metabolism is tailored for the efficient production of secondary metabolites. The insight that we provide can be used to develop high-yielding fungal cell factories that are optimized for the production of specific secondary metabolites of pharmaceutical interest.

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