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https://www.readbyqxmd.com/read/28335329/nanoparticle-based-delivery-of-anaplasma-marginale-membrane-proteins-virb9-1-and-virb10-produced-in-the-pichia-pastoris-expression-system
#1
Bing Zhang, Antonio S Cavallaro, Karishma T Mody, Jun Zhang, James R Deringer, Wendy C Brown, Timothy J Mahony, Chengzhong Yu, Neena Mitter
Bovine anaplasmosis or cattle-tick fever is a tick-borne haemolytic disease caused by the rickettsial haemoparasite Anaplasma marginale in tropical and subtropical areas of the world. While difficult to express, the proteins VirB9-1 and VirB10 are immunogenic components of the outer membrane type IV secretion system that have been identified as candidate antigens for vaccines targeting of A. marginale. Soluble VirB9-1 and VirB10 were successfully expressed using Pichia pastoris. When formulated with the self-adjuvanting silica vesicles, SV-100 (diameter: 50 nm, and pore entrance size: 6 nm), 200 µg of VirB9-1 and VirB10 were adsorbed per milligram of nanoparticle...
November 5, 2016: Nanomaterials
https://www.readbyqxmd.com/read/28334329/metabolomics-of-pichia-pastoris-impact-of-buffering-conditions-on-the-kinetics-and-nature-of-metabolite-loss-during-quenching
#2
Matthias Mattanovich, Hannes Rußmayer, Theresa Scharl-Hirsch, Verena Puxbaum, Jonas Burgard, Diethard Mattanovich, Stephan Hann
Mass spectrometry based metabolomic profiling is a powerful strategy to quantify the concentrations of numerous primary metabolites in parallel. To avoid distortion of metabolite concentrations, quenching is applied to stop the cellular metabolism instantly. For yeasts, cold methanol quenching is accepted to be the most suitable method to stop metabolism, while keeping the cells intact for separation from the supernatant. During this treatment, metabolite loss may occur while the cells are suspended in the quenching solution...
March 15, 2017: FEMS Yeast Research
https://www.readbyqxmd.com/read/28334164/transcription-factor-mxr1-promotes-the-expression-of-aox1-by-repressing-glycerol-transporter-1-in-pichia-pastoris
#3
Chunjun Zhan, Yankun Yang, Zhenyang Zhang, Xiang Li, Xiuxia Liu, Zhonghu Bai
In methylotrophic yeast Pichia pastoris (P. pastoris), the efficient promoter of alcohol oxidase (PAox1) is induced by methanol and repressed by glycerol, but the molecular mechanism is not clear. In this study, the relationship between alcohol oxidase 1 (aox1), methanol expression regulator 1 (mxr1) and glycerol transporter 1 (gt1) was studied. By RT-PCR, it was found that the overexpression of gt1 could increase the glycerol content in cells and repress the expression of mxr1 and aox1, and the deletion of gt1 reduced the glycerol content in cells and promoted the expression of aox1 ...
March 3, 2017: FEMS Yeast Research
https://www.readbyqxmd.com/read/28333108/the-biology-of-pichia-membranifaciens-killer-toxins
#4
REVIEW
Ignacio Belda, Javier Ruiz, Alejandro Alonso, Domingo Marquina, Antonio Santos
The killer phenomenon is defined as the ability of some yeast to secrete toxins that are lethal to other sensitive yeasts and filamentous fungi. Since the discovery of strains of Saccharomyces cerevisiae capable of secreting killer toxins, much information has been gained regarding killer toxins and this fact has substantially contributed knowledge on fundamental aspects of cell biology and yeast genetics. The killer phenomenon has been studied in Pichia membranifaciens for several years, during which two toxins have been described...
March 23, 2017: Toxins
https://www.readbyqxmd.com/read/28330197/xylitol-production-by-saccharomyces-cerevisiae-overexpressing-different-xylose-reductases-using-non-detoxified-hemicellulosic-hydrolysate-of-corncob
#5
Anushree Kogje, Anand Ghosalkar
Xylitol production was compared in fed batch fermentation by Saccharomyces cerevisiae strains overexpressing xylose reductase (XR) genes from Candida tropicalis, Pichia stipitis, Neurospora crassa, and an endogenous gene GRE3. The gene encoding a xylose specific transporter (SUT1) from P. stipitis was cloned to improve xylose transport and fed batch fermentation was used with glucose as a cosubstrate to regenerate NADPH. Xylitol yield was near theoretical for all the strains in fed batch fermentation. The highest volumetric (0...
December 2016: 3 Biotech
https://www.readbyqxmd.com/read/28330196/cloning-and-expression-of-saccharomyces-cerevisiae-suc2-gene-in-yeast-platform-and-characterization-of-recombinant-enzyme-biochemical-properties
#6
Nooshin Mohandesi, Seyed Omid Ranaei Siadat, Kamahldin Haghbeen, Ardeshir Hesampour
Invertase (EC.3.2.1.26) catalyzes the hydrolysis of sucrose to an equimolar mixture of D-glucose and D-fructose which is of interest for various industrial applications. In this research, Saccharomyces cerevisiae invertase gene (SUC2) was optimized based on Pichia pastoris codon preference. The synthetic gene was introduced into the methylotrophic yeast Pichia pastoris under the control of the inducible AOX1 promoter. High level of the extracellular recombinant invertase (R-inv) production was achieved via methanol induction for 4 days and purified by His-Tag affinity chromatography which appeared to be a mixture of glycosylated proteins with various sizes of 85-95 kDa on SDS-PAGE...
December 2016: 3 Biotech
https://www.readbyqxmd.com/read/28330129/the-recombinant-pea-defensin-drr230a-is-active-against-impacting-soybean-and-cotton-pathogenic-fungi-from-the-genera-fusarium-colletotrichum-and-phakopsora
#7
Ariane Ferreira Lacerda, Rafael Perseghini Del Sarto, Marilia Santos Silva, Erico Augusto Rosas de Vasconcelos, Roberta Ramos Coelho, Vanessa Olinto Dos Santos, Claudia Vieira Godoy, Claudine Dinali Santos Seixas, Maria Cristina Mattar da Silva, Maria Fatima Grossi-de-Sa
Plant defensins are antifungal peptides produced by the innate immune system plants developed to circumvent fungal infection. The defensin Drr230a, originally isolated from pea, has been previously shown to be active against various entomopathogenic and phytopathogenic fungi. In the present study, the activity of a yeast-expressed recombinant Drr230a protein (rDrr230a) was tested against impacting soybean and cotton fungi. First, the gene was subcloned into the yeast expression vector pPICZαA and expressed in Pichia pastoris...
June 2016: 3 Biotech
https://www.readbyqxmd.com/read/28328103/characterization-of-maize-chitinase-a-a-tough-allergenic-molecule
#8
Mariateresa Volpicella, Claudia Leoni, Immacolata Fanizza, Marco Distaso, Guido Leoni, Laura Farioli, Todd Naumann, Elide Pastorello, Luigi Ruggiero Ceci
BACKGROUND: Food allergies are recognized as an increasing health concern. They are caused by specific proteins called food allergens. Proteins commonly identified as food allergens tend to have one of about 30 different biochemical activities. This leads to the assumption that food allergens must have specific structural features which causes their allergenicity. But these structural features are not completely understood. Uncovering the structural basis of allergenicity would allow improved diagnosis and therapy of allergies and would provide insights for safer food production...
March 22, 2017: Allergy
https://www.readbyqxmd.com/read/28323169/isolation-and-characterization-of-human-capg-expressed-and-post-translationally-modified-in-pichia-pastoris
#9
Agnes Papala, Marc Sylvester, Nadine Dyballa-Rukes, Sabine Metzger, Jochen D'Haese
CapG is an actin-binding protein, which is overexpressed in a variety of tumors, i.e. breast, ovarian, pancreatic and lung carcinoma. We successfully expressed human CapG in the wild type strain X-33 of the methylotrophic yeast Pichia pastoris (P. pastoris), which does not express endogenous CapG, in order to characterize this protein in more detail. After mechanical cell lysis, debris was centrifuged and the soluble protein was precipitated with ammonium sulfate. This protein pellet was dialyzed and used for CapG purification...
March 18, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28315397/efficient-extracellular-expression-of-transpeptidase-sortase-a-in-pichia-pastoris
#10
Xinrui Zhao, Haofei Hong, Zhimeng Wu
In order to achieve efficient extracellular expression of Sortase A (SrtA), various strategies in Pichia pastoris system were applied in this study. Among different constructed recombinant strains, the SMD1168 strain integrated 5.7 copies of srtA gene under control of AOX1 promoter was proved to be the best strain for the extracellular SrtA expression. After the optimization of fermentation conditions (induction 72 h at 28 °C, initial pH 6.0, supplemented with 1.5% methanol), the highest yield and activity of extracellular SrtA reached 97...
March 15, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28315210/recombinant-allergens-production-in-yeast
#11
Maria Neophytou, Marcos Alcocer
The methylotropic yeast Pichia pastoris has been extensively used in large-scale production of properly folded recombinant proteins. As an eukaryotic organism P. pastoris presents a series of advantages at expression and processing of heterologous proteins such as post-translational modifications, protein processing, and a reasonably sophisticated quality control of protein folding when compared against Escherichia coli. In this chapter, we describe the modified lab procedure for cloning and expression in Pichia pastoris of common food allergens sequences from the raw fruit to the fully folded biotinylated protein product...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28303325/use-of-pervaporation-process-for-the-recovery-of-aroma-compounds-produced-by-p-fermentans-in-sugarcane-molasses
#12
Suzan Cristina Rossi, Adriane Bianchi Pedroni Medeiros, Thiago André Weschenfelder, Agnes de Paula Scheer, Carlos Ricardo Soccol
Natural fruity aroma was produced during submerged fermentation by Pichia fermentans using sugarcane molasses as a cultivation broth. The aroma compounds were recovered from the fermentation by a pervaporation process using a polydimethylsiloxane membrane (Pervap 4060-Sulzer). Isoamyl acetate, a characteristic compound associated with fruity aromas, was the major compound produced. The pervaporation module was fed at three different temperatures to test the best conditions to recover the natural fruity aroma...
March 17, 2017: Bioprocess and Biosystems Engineering
https://www.readbyqxmd.com/read/28302114/implications-of-evolutionary-engineering-for-growth-and-recombinant-protein-production-in-methanol-based-growth-media-in-the-yeast-pichia-pastoris
#13
Josef W Moser, Roland Prielhofer, Samuel M Gerner, Alexandra B Graf, Iain B H Wilson, Diethard Mattanovich, Martin Dragosits
BACKGROUND: Pichia pastoris is a widely used eukaryotic expression host for recombinant protein production. Adaptive laboratory evolution (ALE) has been applied in a wide range of studies in order to improve strains for biotechnological purposes. In this context, the impact of long-term carbon source adaptation in P. pastoris has not been addressed so far. Thus, we performed a pilot experiment in order to analyze the applicability and potential benefits of ALE towards improved growth and recombinant protein production in P...
March 17, 2017: Microbial Cell Factories
https://www.readbyqxmd.com/read/28300184/corrigendum-high-level-expression-of-improved-thermo-stable-alkaline-xylanase-variant-in-pichia-pastoris-through-codon-optimization-multiple-gene-insertion-and-high-density-fermentation
#14
Yihong Lu, Cheng Fang, Qinhong Wang, Yuling Zhou, Guimin Zhang, Yanhe Ma
No abstract text is available yet for this article.
March 16, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28299731/heterologous-expression-of-the-astacin-protease-meprin-%C3%AE-in-pichia-pastoris
#15
Dagmar Schlenzig, Stephan Schilling
Meprins are zinc-dependent proteases of the metzincin superfamily of metalloproteases. The enzymes are extracellular multi-domain proteins which are stabilized by disulfide bridges, dimerization, and glycosylation. Due to their complex structure, recombinant expression was first established in mammalian and insect cells. However, these methods have several disadvantages such as high costs and the low yields. For this reason, yeast is often considered a preferable expression system. Here, we describe the manipulation and secretory expression of human meprin β in the methylotrophic yeast P...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28297751/preparation-of-vsv-g-viral-envelope-glycoprotein-from-pichia-pastoris-enhances-transfection-of-dna-into-animal-cells
#16
Xin Liu, Ying Dong, Jingquan Wang, Long Li, Zhenmin Zhong, Yun-Pan Li, Shao-Jun Chen, Yu-Cai Fu, Wen-Can Xu, Chi-Ju Wei
Vesicular stomatitis virus G glycoprotein (VSV-G) has been widely used for pseudotyping retroviral, lentiviral and artificial viral vectors. The objective of this study was to establish a potential approach for large scale production of VSV-G. To this end, VSV-G was cloned with a N-terminal His-tag into Pichia pastoris expression vector pPIC3.5K. Three clones (Mut(s)) containing the VSV-G expression cassette were identified by PCR. All clones proliferated normally in expansion medium, while the proliferation was reduced significantly under induction conditions...
March 15, 2017: Journal of Microbiology and Biotechnology
https://www.readbyqxmd.com/read/28296967/the-combined-rapid-detection-and-species-level-identification-of-yeasts-in-simulated-blood-culture-using-a-colorimetric-sensor-array
#17
Nabin K Shrestha, Sung H Lim, Deborah A Wilson, Ana Victoria SalasVargas, Yair S Churi, Paul A Rhodes, Peter J Mazzone, Gary W Procop
BACKGROUND: A colorimetric sensor array (CSA) has been demonstrated to rapidly detect and identify bacteria growing in blood cultures by obtaining a species-specific "fingerprint" of the volatile organic compounds (VOCs) produced during growth. This capability has been demonstrated in prokaryotes, but has not been reported for eukaryotic cells growing in culture. The purpose of this study was to explore if a disposable CSA could differentially identify 7 species of pathogenic yeasts growing in blood culture...
2017: PloS One
https://www.readbyqxmd.com/read/28295011/increased-dosage-of-aox1-promoter-regulated-expression-cassettes-leads-to-transcription-attenuation-of-the-methanol-metabolism-in-pichia-pastoris
#18
Elena Cámara, Nils Landes, Joan Albiol, Brigitte Gasser, Diethard Mattanovich, Pau Ferrer
The methanol-regulated alcohol oxidase promoter (PAOX1) of Pichia pastoris is one of the strongest promoters for heterologous gene expression in this methylotrophic yeast. Although increasing gene dosage is one of the most common strategies to increase recombinant protein productivities, the increase of gene dosage of Rhizopus oryzae lipase (ROL) in P. pastoris has been previously shown to reduce cell growth, lipase production and substrate consumption in high-copy strains. To better assess that physiological response, transcriptomics analysis was performed of a subset of strains with 1 to 15 ROL copies...
March 15, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28293387/cfp10-mfc%C3%AE-2-as-a-novel-tuberculosis-vaccine-candidate-increases-immune-response-in-mouse
#19
Ali Asghar Baghani, Saman Soleimanpour, Hadi Farsiani, Arman Mosavat, Masoud Yousefi, Zahra Meshkat, Seyed Abdolrahim Rezaee, Saeid Amel Jamehdar, Mohammad Reza Akbari Eydgahi, Hamid Sadeghian, Kiarash Ghazvini
OBJECTIVES: Despite treatment with antibiotics and vaccination with BCG, tuberculosis (TB) is still considered as one of the most important public health problems in the world. Therefore, designing and producing a more effective vaccine against TB seems urgently. In this study, immunogenicity of a fusion protein which consisting or comprising CFP-10 from Mycobacterium tuberculosis and the Fc-domain of mouse IgG2a was evaluated as a novel subunit vaccine candidate against TB. MATERIALS AND METHODS: The genetic constructs were cloned in pPICZαA expression vector and recombinant vectors (pPICZαA-CFP-10: Fcγ2a and pPICZαA-CFP-10:His) were transformed into Pichia pastoris...
February 2017: Iranian Journal of Basic Medical Sciences
https://www.readbyqxmd.com/read/28285179/expanding-the-feruloyl-esterase-gene-family-of-aspergillus-niger-by-characterization-of-a-feruloyl-esterase-faec
#20
Adiphol Dilokpimol, Miia R Mäkelä, Sadegh Mansouri, Olga Belova, Martin Waterstraat, Mirko Bunzel, Ronald P de Vries, Kristiina S Hildén
A feruloyl esterase (FAE) from Aspergillus niger N402, FaeC was heterologously produced in Pichia pastoris X-33 in a yield of 10mg/L. FaeC was most active at pH 7.0 and 50°C, and showed broad substrate specificity and catalyzed the hydrolysis of methyl 3,4-dimethoxycinnamate, ethyl ferulate, methyl ferulate, methyl p-coumarate, ethyl coumarate, methyl sinapate, and methyl caffeate. The enzyme released both ferulic acid and p-coumaric acid from wheat arabinoxylan and sugar beet pectin (up to 3mg/g polysaccharide), and acted synergistically with a commercial xylanase increasing the release of ferulic acid up to six-fold...
March 8, 2017: New Biotechnology
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