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https://www.readbyqxmd.com/read/29772751/what-we-do-not-know-about-fungal-cell-adhesion-molecules
#1
REVIEW
Peter N Lipke
There has been extensive research on structure and function of fungal cell adhesion molecules, but the most of the work has been about adhesins in Candida albicans and Saccharomyces cerevisiae. These yeasts are members of a single ascomycete order, and adhesion molecules from the six other fungal phyla are only sparsely described in the literature. In these other phyla, most of the research is at the cellular level, rather than at the molecular level, so there has been little characterization of the adhesion molecules themselves...
May 17, 2018: Journal of Fungi (Basel, Switzerland)
https://www.readbyqxmd.com/read/29751659/high-level-expression-of-a-thermally-stable-alginate-lyase-using-pichia-pastoris-characterization-and-application-in-producing-brown-alginate-oligosaccharide
#2
Haifeng Li, Shuling Wang, Yunyi Zhang, Liehuan Chen
An alginate lyase encoding gene sagl from Flavobacterium sp. H63 was codon optimized and recombinantly expressed at high level in P.pastoris through high cell-density fermentation. The highest yield of recombinant enzyme of sagl (rSAGL) in yeast culture supernatant reached 226.4 μg/mL (915.5 U/mL). This was the highest yield record of recombinant expression of alginate lyase so far. The rSAGL was confirmed as a partially glycosylated protein through EndoH digestion. The optimal reaction temperature and pH of this enzyme were 45 °C and 7...
May 11, 2018: Marine Drugs
https://www.readbyqxmd.com/read/29710826/comparison-of-yeasts-as-hosts-for-recombinant-protein-production
#3
REVIEW
Antonio Milton Vieira Gomes, Talita Souza Carmo, Lucas Silva Carvalho, Frederico Mendonça Bahia, Nádia Skorupa Parachin
Recombinant protein production emerged in the early 1980s with the development of genetic engineering tools, which represented a compelling alternative to protein extraction from natural sources. Over the years, a high level of heterologous protein was made possible in a variety of hosts ranging from the bacteria Escherichia coli to mammalian cells. Recombinant protein importance is represented by its market size, which reached $1654 million in 2016 and is expected to reach $2850.5 million by 2022. Among the available hosts, yeasts have been used for producing a great variety of proteins applied to chemicals, fuels, food, and pharmaceuticals, being one of the most used hosts for recombinant production nowadays...
April 29, 2018: Microorganisms
https://www.readbyqxmd.com/read/29704654/metabolic-engineering-of-pichia-pastoris
#4
REVIEW
David A Peña, Brigitte Gasser, Jürgen Zanghellini, Matthias G Steiger, Diethard Mattanovich
Besides its use for efficient production of recombinant proteins the methylotrophic yeast Pichia pastoris (syn. Komagataella spp.) has been increasingly employed as a platform to produce metabolites of varying origin. We summarize here the impressive methodological developments of the last years to model and analyze the metabolism of P. pastoris, and to engineer its genome and metabolic pathways. Efficient methods to insert, modify or delete genes via homologous recombination and CRISPR/Cas9, supported by modular cloning techniques, have been reported...
April 25, 2018: Metabolic Engineering
https://www.readbyqxmd.com/read/29671116/mpdu1-regulates-ceacam1-and-cell-adhesion-in-vitro-and-in-vivo
#5
Daniel C Bennett, Aurelie Cazet, Jon Charest, Joseph N Contessa
N-linked glycosylation (NLG) is a co-translational modification that is essential for the folding, stability, and trafficking of transmembrane (TM) and secretory glycoproteins. Efficient NLG requires the stepwise synthesis and en bloc transfer of a 14-sugar carbohydrate known as a lipid-linked oligosaccharide (LLO). The genetics of LLO biosynthesis have been established in yeast and Chinese hamster systems, but human models of LLO biosynthesis are lacking. In this study we report that Kato III human gastric cancer cells represent a model of deficient LLO synthesis, possessing a homozygous deletion of the LLO biosynthesis factor, MPDU1...
April 18, 2018: Glycoconjugate Journal
https://www.readbyqxmd.com/read/29648454/effect-of-n-linked-glycosylation-of-recombinant-windmill-palm-tree-peroxidase-on-its-activity-and-stability
#6
Bo Fu, Margaret R Baker, Qing X Li
Plant secretory peroxidases are valuable commercial enzymes. The windmill palm tree Trachycarpus fortunei produces one of the most stable and fastest peroxidases (WPTP) characterized to date, however, an economical source is needed. Pichia pastoris has been used as an expression system for WPTP and other peroxidases. However, yeast and plants synthesize different types of N-linked glycan structures and may differ the level of glycosylation at each site. Such non-native glycosylation can have unwanted consequences...
April 12, 2018: Journal of Agricultural and Food Chemistry
https://www.readbyqxmd.com/read/29577692/-cloning-of-oryza-sativa-n-glycanase-gene-ospngase-a-and-its-expression-in-pichia-pastoris
#7
Yuan Wang, Peng Jia, Xuejun Li, Yuhong Li, Peng Chen
N-glycanase is a class of deglycosylation enzymes, widely used in the study of N-glycosylation modification of glycoprotein. In this study, an N-glycanase gene (OsPNGase A, XM_015775832) with high GC content (69.48%) was cloned from rice and then the yeast secretory expression vector pPICZ(α)A-OsPNGase A was constructed for the purpose of transformation to Pichia pastoris. After induction in Pichia pastoris SMD1168H, the target protein was purified by DEAE Sepharose and HisTrap HP chromatography, with a yield of 12...
March 25, 2018: Sheng Wu Gong Cheng Xue Bao, Chinese Journal of Biotechnology
https://www.readbyqxmd.com/read/29566240/identification-of-high-affinity-her2-binding-antibodies-using-cho-fab-surface-display
#8
Annalee W Nguyen, Kevin C Le, Jennifer A Maynard
Discovery of monoclonal antibodies is most commonly performed using phage or yeast display but mammalian cells are used for production because of the complex antibody structure, including the multiple disulfide bonds and glycosylation, required for function. As this transition between host organisms is often accompanied by impaired binding, folding or expression, development pipelines include laborious plate-based screening or engineering strategies to adapt an antibody to mammalian expression. To circumvent these problems, we developed a plasmid-based Fab screening platform on Chinese hamster ovary (CHO) cells which allows for antibody selection in the production host and in the presence of the same post-translational modifications as the manufactured product...
March 1, 2018: Protein Engineering, Design & Selection: PEDS
https://www.readbyqxmd.com/read/29547901/molecular-partners-of-hnot-alg3-the-human-counterpart-of-the-drosophila-not-and-yeast-alg3-gene-suggest-its-involvement-in-distinct-cellular-processes-relevant-to-congenital-disorders-of-glycosylation-cancer-neurodegeneration-and-a-variety-of-further-pathologies
#9
Benedikt Hacker, Christoph Schultheiß, Michael Döring, Ursula Kurzik-Dumke
This study provides first insights into the involvement of hNOT/ALG3, the human counterpart of the Drosophila NOT and yeast ALG3 gene, in various putative molecular networks. HNOT/ALG3 encodes two translated transcripts encoding precursor proteins differing in their N-terminus and showing 33% identity with the yeast asparagine-linked glycosylation 3 (ALG3) protein. Experimental evidence for the functional homology of the proteins of fly and man in the N-glycosylation has still to be provided. In this study, using the yeast two-hybrid technique we identify 17 molecular partners of hNOT-1/ALG3-1...
March 14, 2018: Human Molecular Genetics
https://www.readbyqxmd.com/read/29507322/the-apicomplexa-specific-glucosamine-6-phosphate-n-acetyltransferase-gene-family-encodes-a-key-enzyme-for-glycoconjugate-synthesis-with-potential-as-therapeutic-target
#10
Marta Cova, Borja López-Gutiérrez, Sara Artigas-Jerónimo, Aida González-Díaz, Giulia Bandini, Steven Maere, Lorenzo Carretero-Paulet, Luis Izquierdo
Apicomplexa form a phylum of obligate parasitic protozoa of great clinical and veterinary importance. These parasites synthesize glycoconjugates for their survival and infectivity, but the enzymatic steps required to generate the glycosylation precursors are not completely characterized. In particular, glucosamine-phosphate N-acetyltransferase (GNA1) activity, needed to produce the essential UDP-N-acetylglucosamine (UDP-GlcNAc) donor, has not been identified in any Apicomplexa. We scanned the genomes of Plasmodium falciparum and representatives from six additional main lineages of the phylum for proteins containing the Gcn5-related N-acetyltransferase (GNAT) domain...
March 5, 2018: Scientific Reports
https://www.readbyqxmd.com/read/29505764/heligmosomoides-polygyrus-venom-allergen-like-protein-4-hpval-4-is-a-sterol-binding-protein
#11
Oluwatoyin A Asojo, Rabih Darwiche, Selam Gebremedhin, Geert Smant, Jose L Lozano-Torres, Claire Drurey, Jeroen Pollet, Rick M Maizels, Roger Schneiter, Ruud H P Wilbers
Heligmosomoides polygyrus bakeri is a model parasitic hookworm used to study animal and human helminth diseases. During infection, the parasite releases excretory/secretory products that modulate the immune system of the host. The most abundant protein family in excretory/secretory products comprises the venom allergen-like proteins (VALs), which are members of the SCP/TAPS (sperm-coating protein/Tpx/antigen 5/pathogenesis related-1/Sc7) superfamily. There are >30 secreted Heligmosomoides polygyrus VAL proteins (HpVALs) and these proteins are characterised by having either one or two 15 kDa CAP (cysteine-rich secretory protein (CRISP)/antigen 5/pathogenesis related-1) domains...
April 2018: International Journal for Parasitology
https://www.readbyqxmd.com/read/29501266/crystal-structure-of-brugia-malayi-venom-allergen-like-protein-1-bmval-1-a-vaccine-candidate-for-lymphatic-filariasis
#12
Rabih Darwiche, Fernanda Lugo, Claire Drurey, Koen Varossieau, Geert Smant, Ruud H P Wilbers, Rick M Maizels, Roger Schneiter, Oluwatoyin A Asojo
Brugia malayi is a causative agent of lymphatic filariasis, a major tropical disease. The infective L3 parasite stage releases immunomodulatory proteins including the venom allergen-like proteins (VALs), which are members of the SCP/TAPS (Sperm-coating protein/Tpx/antigen 5/pathogenesis related-1/Sc7) superfamily. BmVAL-1 is a major target of host immunity with >90% of infected B. malayi microfilaraemic cases being seropositive for antibodies to BmVAL-1. This study is part of ongoing efforts to characterize the structures and functions of important B...
April 2018: International Journal for Parasitology
https://www.readbyqxmd.com/read/29487134/the-n-terminal-domain-of-a-tick-evasin-is-critical-for-chemokine-binding-and-neutralization-and-confers-specific-binding-activity-to-other-evasins
#13
James R O Eaton, Yara Alenazi, Kamayani Singh, Graham Davies, Lucia Geis-Asteggiante, Benedikt Kessler, Carol V Robinson, Akane Kawamura, Shoumo Bhattacharya
Tick chemokine-binding proteins (evasins) are an emerging class of biologicals that target multiple chemokines and show anti-inflammatory activities in preclinical disease models. Using yeast surface display, we identified a CCL8-binding evasin, P672, from the tick Rhipicephalus pulchellus We found that P672 binds CCL8 and eight other CC-class chemokines with a Kd < 10 nm and four other CC chemokines with a Kd between 10 and 100 nm and neutralizes CCL3, CCL3L1, and CCL8 with an IC50 < 10 nm The CC chemokine-binding profile was distinct from that of evasin 1 (EVA1), which does not bind CCL8...
April 20, 2018: Journal of Biological Chemistry
https://www.readbyqxmd.com/read/29458553/streptomyces-coelicolor-strains-lacking-polyprenol-phosphate-mannose-synthase-and-protein-o-mannosyl-transferase-are-hyper-susceptible-to-multiple-antibiotics
#14
Robert Howlett, Nicholas Read, Anpu Varghese, Charles Kershaw, Y Hancock, Margaret C M Smith
Polyprenol phosphate mannose (PPM) is a lipid-linked sugar donor used by extra-cytoplasmic glycosyl tranferases in bacteria. PPM is synthesiszed by polyprenol phosphate mannose synthase, Ppm1, and in most Actinobacteria is used as the sugar donor for protein O-mannosyl transferase, Pmt, in protein glycosylation. Ppm1 and Pmt have homologues in yeasts and humans, where they are required for protein O-mannosylation. Actinobacteria also use PPM for lipoglycan biosynthesis. Here we show that ppm1 mutants of Streptomyces coelicolor have increased susceptibility to a number of antibiotics that target cell wall biosynthesis...
March 2018: Microbiology
https://www.readbyqxmd.com/read/29450673/recent-advances-in-enhanced-enzyme-activity-thermostability-and-secretion-by-n-glycosylation-regulation-in-yeast
#15
REVIEW
Fei Ge, Longbao Zhu, Anna Aang, Ping Song, Wanzhen Li, Yugui Tao, Guocheng Du
Yeast has been increasingly used as a host for the expression of enzymes. Compared to other expression systems, the yeast expression system has many advantages including its suitability for large-scale fermentation and its ability to modify enzymes. When expressed in yeast, many recombinant enzymes are N-glycosylated, and this may play an important role in their activity, thermostability and secretion. Although the mechanism underlying this process is not clear, the regulation of N-glycosylation by introducing or eliminating N-glycosylation at specific sites has developed into an important strategy for improving the production or catalytic properties of recombinant enzymes...
May 2018: Biotechnology Letters
https://www.readbyqxmd.com/read/29428518/efemp2-mediates-galnt14-dependent-breast-cancer-cell-invasion
#16
Tao Zuo, Jinshuai Shan, Yang Liu, Rong Xie, Xiaochun Yu, Chen Wu
N-Acetylgalactosaminyltransferase-14 (GALNT14) is a member of acetylgalactosaminyltransferases family. We have shown that GALNT14 could promote breast cancer cell invasion. However, the underlying molecular mechanism is unclear. Here, using yeast two hybrid, we find that EGF-containing fibulin-like extracellular matrix protein 2 (EFEMP2) interacts with GALNT14. Both in vitro and in vivo binding assays show that EFEMP2 is associated with GALNT14. Moreover, we find that GALNT14 mediates glycosylation of EFEMP2...
April 2018: Translational Oncology
https://www.readbyqxmd.com/read/29408294/efficient-expression-and-isolation-of-recombinant-human-interleukin-11-rhil-11-in-pichia-pastoris
#17
Kuo-Ming Yu, Johnson Yiu-Nam Lau, Manson Fok, Yuk-Keung Yeung, Siu-Ping Fok, Felix Shek, Wing-Tak Wong, Qui-Lim Choo
Current source of recombinant human interleukin-11 (rhIL-11) is isolated from a fusion protein expressed by E. coli that requires additional enterokinase to remove linked protein, resulting in product heterogeneity of N-terminal sequence. Due to lack of glycosylation, rhIL-11 is suitable to be expressed by yeast cells. However, the only available yeast-derived rhIL-11 presents an obstacle in low production yield, as well as an unamiable process, such as the use of reverse-phase chromatography employing plenty of toxic organic solvents...
June 2018: Protein Expression and Purification
https://www.readbyqxmd.com/read/29386126/unconventional-trafficking-of-mammalian-phospholipase-d3-to-lysosomes
#18
Adriana Carolina Gonzalez, Michaela Schweizer, Sebastian Jagdmann, Christian Bernreuther, Thomas Reinheckel, Paul Saftig, Markus Damme
Variants in the phospholipase D3 (PLD3) gene have genetically been linked to late-onset Alzheimer's disease. We present a detailed biochemical analysis of PLD3 and reveal its endogenous localization in endosomes and lysosomes. PLD3 reaches lysosomes as a type II transmembrane protein via a (for mammalian cells) uncommon intracellular biosynthetic route that depends on the ESCRT (endosomal sorting complex required for transport) machinery. PLD3 is sorted into intraluminal vesicles of multivesicular endosomes, and ESCRT-dependent sorting correlates with ubiquitination...
January 23, 2018: Cell Reports
https://www.readbyqxmd.com/read/29339117/expression-activation-and-processing-of-a-novel-plant-milk-clotting-aspartic-protease-in-pichia-pastoris
#19
Lucía Feijoo-Siota, José Luis R Rama, Angeles Sánchez-Pérez, Tomás G Villa
Galium verum, also known as Lady's Bedstraw or Cheese Rennet, is an herbaceous perennial plant traditionally used in cheese-making. We used RACE PCR to isolate novel enzymes from Galium verum with the ability to clot milk. This approach generated two cDNA sequences (named preprogaline A and B) encoding proteins displaying the typical plant aspartic protease primary structure. Preprogaline B was expressed in the yeast Pichia pastoris, after deleting and replacing its original signal peptide with the yeast α-factor signal peptide from Saccharomyces cerevisiae...
February 20, 2018: Journal of Biotechnology
https://www.readbyqxmd.com/read/29331014/biochemical-characterization-of-recombinant-thermostable-cohnella-sp-a01-%C3%AE-glucanase
#20
Meysam Rezaie, Saeed Aminzadeh, Farid Heidari, Masoud Mashhadi Akbar Boojar, Ali Asghar Karkhane
Background: Typically, non-cellulytic glucanase, including fungi and yeast cell wall hydrolyzing enzymes, are release by some symbiotic fungi and plants during the mycoparasitic fungi attack on plants. These enzymes are known as the defense mechanisms of plants. This study intends to investigate the biochemical properties of β-1,6-glucanase (bg16M) from native thermophilic bacteria, Cohnella A01. Methods: bg16M gene was cloned and expressed in E. coli BL21 (DE3)...
January 13, 2018: Iranian Biomedical Journal
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