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https://www.readbyqxmd.com/read/28546903/comprehensive-reconstruction-and-evaluation-of-pichia-pastoris-genome-scale-metabolic-model-that-accounts-for-1243-orfs
#1
Rui Ye, Mingzhi Huang, Hongzhong Lu, Jiangchao Qian, Weilu Lin, Ju Chu, Yingping Zhuang, Siliang Zhang
BACKGROUND: Pichia pastoris is one of the most important cell factories for production of industrial enzymes and heterogenous proteins. The genome-scale metabolic model of high quality is crucial for comprehensive understanding of the P. pastoris metabolism. METHODS: In this paper, we upgraded P. pastoris genome-scale metabolic model based on the combination of latest genome annotations and literatures. Then the performance of the new model was evaluated using the Cobra Toolbox v2...
2017: Bioresources and Bioprocessing
https://www.readbyqxmd.com/read/28538674/biosynthesis-of-zno-nanoparticles-by-a-new-pichia-kudriavzevii-yeast-strain-and-evaluation-of-their-antimicrobial-and-antioxidant-activities
#2
Amin Boroumand Moghaddam, Mona Moniri, Susan Azizi, Raha Abdul Rahim, Arbakariya Bin Ariff, Wan Zuhainis Saad, Farideh Namvar, Mohammad Navaderi
The potential ability of a new yeast strain, Pichia kudriavzevii, in the synthesis of zinc oxide nanoparticles (ZnO-NPs) through a green method was explored in this study. The effect of reaction time (12, 24 and 36 h) on the structure of the resulting ZnO nanoparticles was investigated. From the XRD and TEM results, the ZnO-NPs with a hexagonal wurtzite structure and a particle crystal size of ~10-61 nm was formed at different reaction times. Combing XRD, TEM, and PL results, it was revealed that the sample prepared at intermediate duration (24 h) has the most favorable nanosized structure with the lowest defect concentration...
May 24, 2017: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
https://www.readbyqxmd.com/read/28534062/insights-into-the-prevalence-and-underlying-causes-of-clonal-variation-through-transcriptomic-analysis-in-pichia-pastoris
#3
Rochelle Aw, Geraint R Barton, David J Leak
Clonal variation, wherein a range of specific productivities of secreted proteins are observed from supposedly identical transformants, is an accepted aspect of working with Pichia pastoris. It means that a significant number of transformants need to be tested to obtain a representative sample, and in commercial protein production, companies regularly screen thousands of transformants to select for the highest secretor. Here, we have undertaken a detailed investigation of this phenomenon by characterising clones transformed with the human serum albumin gene...
May 22, 2017: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28531457/new-structural-insights-into-the-oligosaccharide-phosphate-fraction-of-pichia-hansenula-holstii-nrrl-y2448-phosphomannan
#4
Paul N Handley, Anthony Carroll, Vito Ferro
The oligosaccharide phosphate fraction (OPF) obtained from mild acid hydrolysis of P. holstii NRRL Y-2448 phosphomannan is the starting material for the preparation of the Phase III anticancer drug candidate PI-88. The OPF was for the first time successfully separated by preparative ion exchange chromatography and the major oligosaccharides isolated and characterized by NMR spectroscopy. The components were also acetylated and subjected to LC-MS analysis. These studies revealed that the OPF also contained all-α(1 → 3)-linked oligosaccharides in addition to the known α(1 → 3)/(1 → 2)-linked species, most likely formed by hydrolysis of the latter...
May 13, 2017: Carbohydrate Research
https://www.readbyqxmd.com/read/28528790/characterization-of-a-novel-thermostable-gh45-endoglucanase-from-chaetomium-thermophilum-and-its-biodegradation-of-pectin
#5
Qinzheng Zhou, Peng Ji, Jianye Zhang, Xue Li, Chao Han
A novel thermostable endoglucanase (CTendo45) encoding gene was cloned from Chaetomium thermophilum and heterologously expressed in Pichia pastoris. Sequence alignment indicated that the CTendo45 enzyme belonged to glycoside hydrolase family 45. The recombinant enzyme was purified by Ni(2+) affinity chromatography, and its apparent molecular mass was estimated to be 32 kDa by SDS-PAGE. The purified enzyme displayed maximum activity at 70°C and pH 4. CTendo45 was stable at 60°C for 1 h, and residual activities of 78...
May 19, 2017: Journal of Bioscience and Bioengineering
https://www.readbyqxmd.com/read/28526747/functional-regions-of-the-peroxin-pex19-necessary-for-peroxisome-biogenesis
#6
Gaurav Agrawal, Helen H Shang, Zhi-Jie Xia, Suresh Subramani
The peroxins Pex19 and Pex3 play an indispensable role in peroxisomal membrane protein (PMP) biogenesis, peroxisome division and inheritance. Pex19 plays multiple roles in these processes, but how these functions relate to the structural organization of the Pex19 domains is unresolved. To this end, using deletion mutants, we mapped the Pex19 regions required for peroxisome biogenesis in the yeast Pichia pastoris. Surprisingly, import-competent peroxisomes still formed when Pex19 domains previously believed to be required for biogenesis were deleted, although the peroxisome size was larger than that in wild-type cells...
May 19, 2017: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/28526017/the-effect-of-hypoxia-on-the-lipidome-of-recombinant-pichia-pastoris
#7
Núria Adelantado, Pablo Tarazona, Karlheinz Grillitsch, Xavier García-Ortega, Sergi Monforte, Francisco Valero, Ivo Feussner, Günther Daum, Pau Ferrer
BACKGROUND: Cultivation of recombinant Pichia pastoris (Komagataella sp.) under hypoxic conditions has a strong positive effect on specific productivity when the glycolytic GAP promoter is used for recombinant protein expression, mainly due to upregulation of glycolytic conditions. In addition, transcriptomic analyses of hypoxic P. pastoris pointed out important regulation of lipid metabolism and unfolded protein response (UPR). Notably, UPR that plays a role in the regulation of lipid metabolism, amino acid metabolism and protein secretion, was found to be upregulated under hypoxia...
May 19, 2017: Microbial Cell Factories
https://www.readbyqxmd.com/read/28523913/expression-and-characterization-of-windmill-palm-tree-trachycarpus-fortunei-peroxidase-by-pichia-pastoris
#8
Boting Wen, Margaret R Baker, Hongwei Zhao, Zongjun Cui, Qing X Li
Currently, commercial plant peroxidases are all native and are isolated from plants such as horseradish and soybean. No recombinant plant peroxidase products have been available on the commercial market. The gene encoding peroxidase was cloned from windmill palm tree leaves. The codon-optimized gene was transformed into Pichia pastoris for expression. The recombinant windmill palm tree peroxidase (rWPTP) expressed by P. pastoris showed high stability under pH 2-10 and temperatures up to 70 °C to many metallic salts and organic solvents...
May 26, 2017: Journal of Agricultural and Food Chemistry
https://www.readbyqxmd.com/read/28523730/investigating-the-effect-of-carbon-source-on-rabies-virus-glycoprotein-production-in-pichia-pastoris-by-a-transcriptomic-approach
#9
Safa Ben Azoun, Héla Kallel
Several factors affect protein expression in Pichia pastoris, one among them is the carbon source. In this work, we studied the effect of this factor on the expression level of rabies virus glycoprotein (RABV-G) in two recombinant clones harboring seven copies of the gene of interest. The expression was driven either by the constitutive glyceraldehyde-3-phosphate dehydrogenase (GAP) promoter or the inducible alcohol oxidase1 (AOX1) promoter. Clones were compared in terms of cell physiology and carbon source metabolism...
May 18, 2017: MicrobiologyOpen
https://www.readbyqxmd.com/read/28515785/recombinant-expression-of-thermostable-processive-mteg5-endoglucanase-and-its-synergism-with-mtlpmo-from-myceliophthora-thermophila-during-the-hydrolysis-of-lignocellulosic-substrates
#10
Anthi Karnaouri, Madhu Nair Muraleedharan, Maria Dimarogona, Evangelos Topakas, Ulrika Rova, Mats Sandgren, Paul Christakopoulos
BACKGROUND: Filamentous fungi are among the most powerful cellulolytic organisms in terrestrial ecosystems. To perform the degradation of lignocellulosic substrates, these microorganisms employ both hydrolytic and oxidative mechanisms that involve the secretion and synergism of a wide variety of enzymes. Interactions between these enzymes occur on the level of saccharification, i.e., the release of neutral and oxidized products, but sometimes also reflected in the substrate liquefaction...
2017: Biotechnology for Biofuels
https://www.readbyqxmd.com/read/28513915/impact-of-a-determinant-mutations-on-detection-of-hepatitis-b-surface-antigen-hbsag-in-hbv-strains-from-chinese-patients-with-occult-hepatitis-b
#11
Xiangyan Huang, Chenyun Ma, Qiang Zhang, Qingfen Shi, Tao Huang, Chao Liu, Jie Li, F Blaine Hollinger
This study was designed to detect mutations that occur within the "a" determinant in the S gene of the hepatitis B virus (HBV) in patients with occult hepatitis B (OHB), and to analyze the influence of these mutations on expression and reactivity of the hepatitis B surface antigen (HBsAg). Twenty-three certified OHB samples were compared to 32 HBsAg positive samples from patients with chronic hepatitis B. The median HBV DNA levels in the OHB group were significantly lower than those in the control group (p < 0...
May 17, 2017: Journal of Medical Virology
https://www.readbyqxmd.com/read/28512458/a-mitochondrial-autonomously-replicating-sequence-from-pichia-pastoris-for-uniform-high-level-recombinant-protein-production
#12
Jan-Philipp Schwarzhans, Tobias Luttermann, Daniel Wibberg, Anika Winkler, Wolfgang Hübner, Thomas Huser, Jörn Kalinowski, Karl Friehs
Pichia pastoris is a non-conventional methylotrophic yeast that is widely used for recombinant protein production, typically by stably integrating the target gene into the genome as part of an expression cassette. However, the comparatively high clonal variability associated with this approach usually necessitates a time intense screening step in order to find strains with the desired productivity. Some of the factors causing this clonal variability can be overcome using episomal vectors containing an autonomously replicating sequence (ARS)...
2017: Frontiers in Microbiology
https://www.readbyqxmd.com/read/28512317/structural-determinants-of-neosartorya-fischeri-antifungal-protein-nfap-for-folding-stability-and-antifungal-activity
#13
László Galgóczy, Attila Borics, Máté Virágh, Hargita Ficze, Györgyi Váradi, Zoltán Kele, Florentine Marx
The recent global challenges to prevent and treat fungal infections strongly demand for the development of new antifungal strategies. The structurally very similar cysteine-rich antifungal proteins from ascomycetes provide a feasible basis for designing new antifungal molecules. The main structural elements responsible for folding, stability and antifungal activity are not fully understood, although this is an essential prerequisite for rational protein design. In this study, we used the Neosartorya fischeri antifungal protein (NFAP) to investigate the role of the disulphide bridges, the hydrophobic core, and the N-terminal amino acids in the formation of a highly stable, folded, and antifungal active protein...
May 16, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28508385/improving-the-temperature-characteristics-and-catalytic-efficiency-of-a-mesophilic-xylanase-from-aspergillus-oryzae-aoxyn11a-by-iterative-mutagenesis-based-on-in-silico-design
#14
Xue-Qing Li, Qin Wu, Die Hu, Rui Wang, Yan Liu, Min-Chen Wu, Jian-Fang Li
To improve the temperature characteristics and catalytic efficiency of a glycoside hydrolase family (GHF) 11 xylanase from Aspergillus oryzae (AoXyn11A), its variants were predicted based on in silico design. Firstly, Gly(21) with the maximum B-factor value, which was confirmed by molecular dynamics (MD) simulation on the three-dimensional structure of AoXyn11A, was subjected to site-saturation mutagenesis. Thus, one variant with the highest thermostability, AoXyn11A(G21I), was selected from the mutagenesis library, E...
December 2017: AMB Express
https://www.readbyqxmd.com/read/28503710/characterization-of-a-new-multifunctional-beta-glucosidase-from-musca-domestica
#15
Shu Zhang, Jian Huang, Rong Hu, Guo Guo, Xiaoli Shang, Jianwei Wu
OBJECTIVE: To engineer Pichia pastoris for heterologous production of cellulase from Musca domestica and explore its potential for industrial applications. RESULTS: A new beta-glucosidase gene (bg), encoding 562 amino acids, was cloned from M. domestica by using rapid amplification of cDNA ends. The gene bg was linked to pPICZαA and expressed in P. pastoris with a yield of 500 mg l(-1). The enzyme has the maximum activity with 27.6 U mg(-1) towards cellulose...
May 13, 2017: Biotechnology Letters
https://www.readbyqxmd.com/read/28500590/autolysis-of-pichia-pastoris-induced-by-cold
#16
Yaneth Bartolo-Aguilar, Luc Dendooven, Cipriano Chávez-Cabrera, Luis B Flores-Cotera, María E Hidalgo-Lara, Lourdes Villa-Tanaca, Rodolfo Marsch
The production of recombinant biopharmaceutical proteins is a multi-billion dollar market. Protein recovery represents a major part of the production costs. Pichia pastoris is one of the microbial systems most used for the production of heterologous proteins. The use of a cold-induced promoter to express lytic enzymes in the yeast after the growth stage could reduce protein recovery costs. This study shows that a cold-shock can be applied to induce lysis of the yeast cells. A strain of P. pastoris was constructed in which the endogenous eng gene encoding a putative endo-β-1,3-glucanase was overexpressed using the cold-shock induced promoter of the cctα gene from Saccharomyces cerevisiae...
December 2017: AMB Express
https://www.readbyqxmd.com/read/28495510/in%C3%A2-vitro-characterization-of-grass-carp-ctenopharyngodon-idella-il-26-in-regulating-inflammatory-factors
#17
Xingyang Qiu, Mengyuan Lv, Xiaoyu Jian, Di Chen, Hong Zhou, Anying Zhang, Xinyan Wang
Interleukin 26 (IL-26) gene has been identified in human, amphibian and teleost but not in rodents. It is well accepted that IL-26 was a crucial member of IL-10 family which acts as a pro-inflammatory cytokine in human. However, the role of IL-26 in regulating inflammation in lower vertebrates including teleost has not been defined yet. In the present study, grass carp IL-26 (gcIL-26) coding sequence was isolated and identified. Its chromosomal synteny was also analyzed, showing that gcIL-26 gene is flanked by IL-22 and IFN-γ genes with the same transcriptional orientation as seen in human, amphibian and zebrafish...
May 8, 2017: Fish & Shellfish Immunology
https://www.readbyqxmd.com/read/28490329/directed-evolution-and-secretory-expression-of-a-pyrethroid-hydrolyzing-esterase-with-enhanced-catalytic-activity-and-thermostability
#18
Xiaolong Liu, Mingjun Liang, Yuhuan Liu, Xinjiong Fan
BACKGROUND: Pyrethroids are potentially harmful to human health and ecosystems. It is necessary to develop some efficient strategies to degrade pyrethroid residues. Biodegradation is generally considered as a safe, efficient, and inexpensive way to eliminate environmental contaminants. To date, although several pyrethroid-hydrolyzing esterases have been cloned, there has been no report about a pyrethroid hydrolase with high hydrolytic activity, good stability, and high productivity, indispensable enzymatic properties in practical biodegradation...
May 11, 2017: Microbial Cell Factories
https://www.readbyqxmd.com/read/28484256/improved-thermostability-of-an-acidic-xylanase-from-aspergillus-sulphureus-by-combined-disulphide-bridge-introduction-and-proline-residue-substitution
#19
Wenhan Yang, Yongzhi Yang, Lingdi Zhang, Hang Xu, Xiaojing Guo, Xu Yang, Bing Dong, Yunhe Cao
As a feed additive, xylanase has been widely applied in the feed of monogastric animals, which contains multiple plant polysaccharides. However, during feed manufacture, the high pelleting temperatures challenge wild-type xylanases. The aim of this study was to improve the thermostability of Aspergillus sulphureus acidic xylanase. According to the predicted protein structure, a series of disulphide bridges and proline substitutions were created in the xylanase by PCR, and the mutants were expressed in Pichia pastoris...
May 8, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28482284/expression-of-a-novel-recombinant-cyanate-hydratase-rtl-cyn-in-pichia-pastoris-characteristics-and-applicability-in-the-detoxification-of-cyanate
#20
Bibhuti Ranjan, Santhosh Pillai, Kugenthiren Permaul, Suren Singh
A recombinant Pichia pastoris harbouring the cyanate hydratase gene (rTl-Cyn) from the thermophilic fungus Thermomyces lanuginosus SSBP yielded a high titre of extracellular cyanate hydratase (100±13UmL(-1)) which was ∼10-fold higher than the native fungal strain. The purified rTl-Cyn had a molecular mass of ∼20kDa on SDS-PAGE, with Km, Vmax, kcat and kcat/Km values of 0.34mM, 2857.14µmolesmg(-1)min(-1), 2.14×10(4)s(-1) and 6.3 ×10(7)M(-1)s(-1), respectively. Its properties of thermostability, pH stability, and heavy metals insensitivity, make it a suitable candidate for bioremediation in extreme environments...
April 27, 2017: Bioresource Technology
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