scholarly journals Optimisation of Recombinant Myrosinase Production in Pichia pastoris

2021 ◽  
Vol 22 (7) ◽  
pp. 3677
Author(s):  
Zuzana Rosenbergová ◽  
Kristína Kántorová ◽  
Martin Šimkovič ◽  
Albert Breier ◽  
Martin Rebroš
Keyword(s):  
Pichia Pastoris ◽  
Plant Defence ◽  
Plant Secondary Metabolites ◽  
Specific Productivity ◽  
Fermentation Conditions ◽  
Threefold Increase ◽  
Dry Cell Weight ◽  
First Time ◽  
Hydrolysis Of ◽  
Dry Cell

Myrosinase is a plant defence enzyme catalysing the hydrolysis of glucosinolates, a group of plant secondary metabolites, to a range of volatile compounds. One of the products, isothiocyanates, proved to have neuroprotective and chemo-preventive properties, making myrosinase a pharmaceutically interesting enzyme. In this work, extracellular expression of TGG1 myrosinase from Arabidopsis thaliana in the Pichia pastoris KM71H (MutS) strain was upscaled to a 3 L laboratory fermenter for the first time. Fermentation conditions (temperature and pH) were optimised, which resulted in a threefold increase in myrosinase productivity compared to unoptimised fermentation conditions. Dry cell weight increased 1.5-fold, reaching 100.5 g/L without additional glycerol feeding. Overall, a specific productivity of 4.1 U/Lmedium/h was achieved, which was 102.5-fold higher compared to flask cultivations.

Optimization of fermentation conditions for the production of recombinant hpdgf-bb in Pichia pastoris

2015 ◽  
Vol 37 (1se) ◽  
Author(s):  
Duong Long Duy ◽  
Pham Minh Vu ◽  
Nguyen Tri Nhan ◽  
Tran Linh Thuoc ◽  
Dang Thi Phuong Thao
Keyword(s):  
Pichia Pastoris ◽  
Fermentation Conditions ◽  
Optimization Of Fermentation

scholarly journals Integrating metabolomics and targeted gene expression to uncover potential biomarkers of fungal/oomycetes-associated disease susceptibility in grapevine

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Marisa Maia ◽  
António E. N. Ferreira ◽  
Rui Nascimento ◽  
Filipa Monteiro ◽  
Francisco Traquete ◽  
...  
Keyword(s):  
Gene Expression ◽  
Plant Defence ◽  
Breeding Programs ◽  
Analysis Group ◽  
Expression Of Genes ◽  
Leucoanthocyanidin Reductase ◽  
Defence Responses ◽  
Targeted Gene Expression ◽  
Plant Defence Responses ◽  
First Time

Abstract Vitis vinifera, one of the most cultivated fruit crops, is susceptible to several diseases particularly caused by fungus and oomycete pathogens. In contrast, other Vitis species (American, Asian) display different degrees of tolerance/resistance to these pathogens, being widely used in breeding programs to introgress resistance traits in elite V. vinifera cultivars. Secondary metabolites are important players in plant defence responses. Therefore, the characterization of the metabolic profiles associated with disease resistance and susceptibility traits in grapevine is a promising approach to identify trait-related biomarkers. In this work, the leaf metabolic composition of eleven Vitis genotypes was analysed using an untargeted metabolomics approach. A total of 190 putative metabolites were found to discriminate resistant/partial resistant from susceptible genotypes. The biological relevance of discriminative compounds was assessed by pathway analysis. Several compounds were selected as promising biomarkers and the expression of genes coding for enzymes associated with their metabolic pathways was analysed. Reference genes for these grapevine genotypes were established for normalisation of candidate gene expression. The leucoanthocyanidin reductase 2 gene (LAR2) presented a significant increase of expression in susceptible genotypes, in accordance with catechin accumulation in this analysis group. Up to our knowledge this is the first time that metabolic constitutive biomarkers are proposed, opening new insights into plant selection on breeding programs.

scholarly journals Complete biosynthesis of a sulfated chondroitin in Escherichia coli

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Abinaya Badri ◽  
Asher Williams ◽  
Adeola Awofiranye ◽  
Payel Datta ◽  
Ke Xia ◽  
...  
Keyword(s):  
Chondroitin Sulfate ◽  
Scale Up ◽  
Production Methods ◽  
E Coli ◽  
Microbial Cell Factories ◽  
Cell Factories ◽  
Microbial Product ◽  
Dry Cell Weight ◽  
One Step ◽  
Dry Cell

AbstractSulfated glycosaminoglycans (GAGs) are a class of important biologics that are currently manufactured by extraction from animal tissues. Although such methods are unsustainable and prone to contamination, animal-free production methods have not emerged as competitive alternatives due to complexities in scale-up, requirement for multiple stages and cost of co-factors and purification. Here, we demonstrate the development of single microbial cell factories capable of complete, one-step biosynthesis of chondroitin sulfate (CS), a type of GAG. We engineer E. coli to produce all three required components for CS production–chondroitin, sulfate donor and sulfotransferase. In this way, we achieve intracellular CS production of ~27 μg/g dry-cell-weight with about 96% of the disaccharides sulfated. We further explore four different factors that can affect the sulfation levels of this microbial product. Overall, this is a demonstration of simple, one-step microbial production of a sulfated GAG and marks an important step in the animal-free production of these molecules.

scholarly journals Biosynthesis of anandamide and N-palmitoylethanolamine by sequential actions of phospholipase A2 and lysophospholipase D

2004 ◽  
Vol 380 (3) ◽  
pp. 749-756 ◽  
Author(s):  
Yong-Xin SUN ◽  
Kazuhito TSUBOI ◽  
Yasuo OKAMOTO ◽  
Takeharu TONAI ◽  
Makoto MURAKAMI ◽  
...  
Keyword(s):  
Biosynthetic Pathway ◽  
Brain Homogenate ◽  
Wide Distribution ◽  
Rat Tissues ◽  
Lysophospholipase D ◽  
Pla2 Enzyme ◽  
First Time ◽  
Hydrolysis Of ◽  
The Brain

Anandamide (an endocannabinoid) and other bioactive long-chain NAEs (N-acylethanolamines) are formed by direct release from N-acyl-PE (N-acyl-phosphatidylethanolamine) by a PLD (phospholipase D). However, the possible presence of a two-step pathway from N-acyl-PE has also been suggested previously, which comprises (1) the hydrolysis of N-acyl-PE to N-acyl-lysoPE by PLA1/PLA2 enzyme(s) and (2) the release of NAEs from N-acyllysoPE by lysoPLD (lysophospholipase D) enzyme(s). In the present study we report for the first time the characterization of enzymes responsible for this pathway. The PLA1/PLA2 activity for N-palmitoyl-PE was found in various rat tissues, with the highest activity in the stomach. This stomach enzyme was identified as group IB sPLA2 (secretory PLA2), and its product was determined as N-acyl-1-acyl-lysoPE. Recombinant group IB, IIA and V of sPLA2s were also active with N-palmitoyl-PE, whereas group X sPLA2 and cytosolic PLA2α were inactive. In addition, we found wide distribution of lysoPLD activity generating N-palmitoylethanolamine from N-palmitoyl-lysoPE in rat tissues, with higher activities in the brain and testis. Based on several lines of enzymological evidence, the lysoPLD enzyme could be distinct from the known N-acyl-PE-hydrolysing PLD. sPLA2-IB dose dependently enhanced the production of N-palmitoylethanolamine from N-palmitoyl-PE in the brain homogenate showing the lysoPLD activity. N-Arachidonoyl-PE and N-arachidonoyl-lysoPE as anandamide precursors were also good substrates of sPLA2-IB and the lysoPLD respectively. These results suggest that the sequential actions of PLA2 and lysoPLD may constitute another biosynthetic pathway for NAEs, including anandamide.

Enzyme system of Clostridium stercorarium for hydrolysis of arabinoxylan: reconstitution of the in vivo system from recombinant enzymes

Microbiology ◽  
2004 ◽  
Vol 150 (7) ◽  
pp. 2257-2266 ◽  
Author(s):  
Helmuth Adelsberger ◽  
Christian Hertel ◽  
Erich Glawischnig ◽  
Vladimir V. Zverlov ◽  
Wolfgang H. Schwarz
Keyword(s):  
Extracellular Enzymes ◽  
Enzyme System ◽  
Thermophilic Bacterium ◽  
Recombinant Enzymes ◽  
Type Mode ◽  
Complete Set ◽  
First Time ◽  
Hydrolysis Of

Four extracellular enzymes of the thermophilic bacterium Clostridium stercorarium are involved in the depolymerization of de-esterified arabinoxylan: Xyn11A, Xyn10C, Bxl3B, and Arf51B. They were identified in a collection of eight clones producing enzymes hydrolysing xylan (xynA, xynB, xynC), β-xyloside (bxlA, bxlB, bglZ) and α-arabinofuranoside (arfA, arfB). The modular enzymes Xyn11A and Xyn10C represent the major xylanases in the culture supernatant of C. stercorarium. Both hydrolyse arabinoxylan in an endo-type mode, but differ in the pattern of the oligosaccharides produced. Of the glycosidases, Bxl3B degrades xylobiose and xylooligosaccharides to xylose, and Arf51B is able to release arabinose residues from de-esterified arabinoxylan and from the oligosaccharides generated. The other glycosidases either did not attack or only marginally attacked these oligosaccharides. Significantly more xylanase and xylosidase activity was produced during growth on xylose and xylan. This is believed to be the first time that, in a single thermophilic micro-organism, the complete set of enzymes (as well as the respective genes) to completely hydrolyse de-esterified arabinoxylan to its monomeric sugar constituents, xylose and arabinose, has been identified and the enzymes produced in vivo. The active enzyme system was reconstituted in vitro from recombinant enzymes.

scholarly journals NH<sub>3</sub>-promoted hydrolysis of NO<sub>2</sub> induces explosive growth in HONO

2018 ◽  
Author(s):  
Wanyun Xu ◽  
Ye Kuang ◽  
Chunsheng Zhao ◽  
Jiangchuan Tao ◽  
Gang Zhao ◽  
...  
Keyword(s):  
North China Plain ◽  
North China ◽  
Ph Value ◽  
Primary Source ◽  
Field Measurements ◽  
Oh Radicals ◽  
The North ◽  
The North China Plain ◽  
First Time ◽  
Hydrolysis Of

Abstract. The study of atmospheric nitrous acid (HONO), which is the primary source of OH radicals, is crucial to atmospheric photochemistry and heterogeneous chemical processes. The heterogeneous NO2 chemistry under haze conditions was pointed out to be one of the missing sources of HONO on the North China Plain, producing sulfate and nitrate in the process. However, controversy exists between various proposed mechanisms, mainly debating on whether SO2 directly takes part in the HONO production process and what roles NH3 and the pH value play in it. In this paper, never before seen explosive HONO production (maximum rate: 16 ppb/hour) was reported and evidence was found for the first time in field measurements during fog episodes (usually with pH > 5) and haze episodes under high relative humidity (usually with pH 

Halides and oxide halides of molybdenum and tungsten. III. Oxo-bridged species of tungsten(IV) and molybdenum(V)

1968 ◽  
Vol 21 (4) ◽  
pp. 883 ◽  
Author(s):  
R Colton ◽  
GG Rose
Keyword(s):  
Partial Hydrolysis ◽  
First Time ◽  
Hydrolysis Of ◽  
And Tungsten

The compound formulated in the literature as K2W(OH)Cl5 has been shown to be the oxo-bridged dimer K4[WT2Ocl10], and the diamagnetic dimer formed by the partial hydrolysis of the MOOCl2- ion has been isolated for the first time and shown to be the oxo-bridged [Mo2O3Cl8]4- ion.

Topotactic route to synthesis of novel hydroxylated phases: I. Trioctahedral Micas

Clay Minerals ◽  
1986 ◽  
Vol 21 (2) ◽  
pp. 125-131 ◽  
Author(s):  
S. Komarneni ◽  
R. Roy
Keyword(s):  
Ion Exchange ◽  
Alkaline Earth ◽  
Water Molecules ◽  
Hydration Energy ◽  
Hydrothermal Conditions ◽  
Potential Applications ◽  
Clay Layers ◽  
First Time ◽  
Hydrolysis Of ◽  
Alkaline Earth Cations

AbstractK-depleted phlogopite mica was used as a topotactic precursor and treated with alkali (Li+, K+, , Rb+, Cs+), alkaline-earth (Mg2+, Ca2+, Sr2+, Ba2+) and trivalent (Al3+) cations under hydrothermal conditions of 200°C and 30 MPa pressure. K-, NH4-, Rb- and Cs-aluminosilicate micas were synthesised at 200°C in one day. The synthesis of Cs-aluminosilicate mica, with potential applications in the management of nuclear wastes, has been achieved for the first time by this approach. Ion exchange by Li+, Na+ and alkaline-earth cations under hydrothermal conditions did not produce anhydrous mica phases but resulted in hydrous phases with one or two layers of water molecules between the clay layers. The formation of hydrous phases may be attributed to the high hydration energy of the above cations compared to K+, , RB+ and Cs+. Ion exchange with Al3+ produced a chlorite-like phase because of the hydrolysis of Al3+ under these hydrothermal conditions. These studies are of relevance in the immobilization of wastes where hazardous ions can be fixed in highly stable insoluble phases like mica or chlorite.

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