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https://www.readbyqxmd.com/read/29150513/high-prevalence-and-genetic-diversity-of-large-phicd211-phicdif1296t-like-prophages-in-clostridioides-difficile
#1
Julian R Garneau, Ognjen Sekulovic, Bruno Dupuy, Olga Soutourina, Marc Monot, Louis-Charles Fortier
Clostridioides difficile (formerly Clostridium difficile) is a pathogenic bacterium displaying great genetic diversity. A significant proportion of this diversity is due to the presence of integrated prophages. Here, we provide an in-depth analysis of phiCD211, also known as phiCDIF1296T, the largest phage identified in C. difficile so far, with a genome of 131-kbp. It shares morphological and genomic similarity with other large siphophages like phage 949 infecting Lactococcus lactis and phage c-st infecting Clostridium botulinum...
November 17, 2017: Applied and Environmental Microbiology
https://www.readbyqxmd.com/read/29143917/lambda-phage-nanoparticles-displaying-her2-derived-e75-peptide-induce-effective-e75-cd8-t-response
#2
Atefeh Arab, Jessica Nicastro, Roderick Slavcev, Atefeh Razazan, Nastaran Barati, Amin Reza Nikpoor, Amir Abbas Momtazi Brojeni, Fatemeh Mosaffa, Ali Badiee, Mahmoud Reza Jaafari, Javad Behravan
We have investigated the in vitro immunogenicity and in vivo prophylactic and therapeutic potential of lambda (λ) phage particles displaying the E75 peptide (derived from HER2 protein) in an implantable TUBO breast tumor model of BALB/c mice. The mice were immunized with the E75-displaying phage (λF7-gpD::E75) every 2-week intervals over a 6-week period, and the generated immune responses were studied. Results showed in vitro induction of immune responses by the λF7 (gpD::E75) construct compared to the control λF7 and buffer groups...
November 16, 2017: Immunologic Research
https://www.readbyqxmd.com/read/29138389/phage-display-and-selection-of-lanthipeptides-on-the-carboxy-terminus-of-the-gene-3-minor-coat-protein
#3
Johannes H Urban, Markus A Moosmeier, Tobias Aumüller, Marcus Thein, Tjibbe Bosma, Rick Rink, Katharina Groth, Moritz Zulley, Katja Siegers, Kathrin Tissot, Gert N Moll, Josef Prassler
Ribosomally synthesized and post-translationally modified peptides (RiPPs) are an emerging class of natural products with drug-like properties. To fully exploit the potential of RiPPs as peptide drug candidates, tools for their systematic engineering are required. Here we report the engineering of lanthipeptides, a subclass of RiPPs characterized by multiple thioether cycles that are enzymatically introduced in a regio- and stereospecific manner, by phage display. This was achieved by heterologous co-expression of linear lanthipeptide precursors fused to the widely neglected C-terminus of the bacteriophage M13 minor coat protein pIII, rather than the conventionally used N-terminus, along with the modifying enzymes from distantly related bacteria...
November 15, 2017: Nature Communications
https://www.readbyqxmd.com/read/29138294/isolation-of-isoform-specific-binding-proteins-affimers-by-phage-display-using-negative-selection
#4
Anna Ah-San Tang, Christian Tiede, David J Hughes, Michael J McPherson, Darren C Tomlinson
Some 30 years after its discovery, phage display remains one of the most widely used methods of in vitro selection. Initially developed to revolutionize the generation of therapeutic antibodies, phage display is now the first choice for screening artificial binding proteins. Artificial binding proteins can be used as reagents to study protein-protein interactions, target posttranslational modifications, and distinguish between homologous proteins. They can also be used as research and affinity reagents, for diagnostic purposes, and as therapeutics...
November 14, 2017: Science Signaling
https://www.readbyqxmd.com/read/29130184/generation-of-high-sensitivity-monoclonal-antibodies-specific-for-homoserine-lactones
#5
Soumya Palliyil
A number of bacteria use a class of chemical compounds called acyl-homoserine lactones (AHLs) as quorum sensing (QS) signals to coordinate their behavior at the population level, including pathogens like Pseudomonas aeruginosa. Blocking QS using antibodies is an attractive strategy for infection control as this process takes a central role in P. aeruginosa infections. Here the methods involved in the generation of high sensitivity anti-QS monoclonal antibodies from an immunized sheep phage display antibody library are described...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29129044/anti-sema3a-antibody-a-novel-therapeutic-agent-to-suppress-gbm-tumor-growth
#6
Jaehyun Lee, Yong Jae Shin, Kyoungmin Lee, Hee Jin Cho, Jason K Sa, Sang-Yun Lee, Seok-Hyung Kim, Jeongwu Lee, Yeup Yoon, Do-Hyun Nam
Purpose: Glioblastoma (GBM) is classified as one of the most aggressive and lethal brain tumor. Great strides have been made in understanding the genomic and molecular underpinnings of GBM, which translated into development of new therapeutic approaches to combat such deadly disease. However, there are only few therapeutic agents that can effectively inhibit GBM invasion in a clinical framework. In an effort to address such challenges, we have generated anti-SEMA3A monoclonal antibody as a potential therapeutic antibody against GBM progression...
November 10, 2017: Cancer Research and Treatment: Official Journal of Korean Cancer Association
https://www.readbyqxmd.com/read/29123083/identification-of-a-peptide-recognizing-cerebrovascular-changes-in-mouse-models-of-alzheimer-s-disease
#7
Aman P Mann, Pablo Scodeller, Sazid Hussain, Gary B Braun, Tarmo Mölder, Kadri Toome, Rajesh Ambasudhan, Tambet Teesalu, Stuart A Lipton, Erkki Ruoslahti
Cerebrovascular changes occur in Alzheimer's disease (AD). Using in vivo phage display, we searched for molecular markers of the neurovascular unit, including endothelial cells and astrocytes, in mouse models of AD. We identified a cyclic peptide, CDAGRKQKC (DAG), that accumulates in the hippocampus of hAPP-J20 mice at different ages. Intravenously injected DAG peptide homes to neurovascular unit endothelial cells and to reactive astrocytes in mouse models of AD. We identified connective tissue growth factor (CTGF), a matricellular protein that is highly expressed in the brain of individuals with AD and in mouse models, as the target of the DAG peptide...
November 10, 2017: Nature Communications
https://www.readbyqxmd.com/read/29119720/phage-display-on-anti-infective-target-dxs-led-to-an-acceptor-substrate-competitive-peptidic-inhibitor
#8
Alessio Marcozzi, Tiziana Masini, Di Zhu, Diego Pesce, Boris Illarionov, Markus Fischer, Andreas Herrmann, Anna Katharina Herta Hirsch
Enzymes of the 2-C-methyl-d-erythritol-4-phosphate pathway for the biosynthesis of isoprenoid precursors are validated drug targets. By performing phage display on 1-deoxy-d-xylulose 5-phosphate synthase (DXS), which catalyzes the first step of this pathway, we discovered several peptide hits and recognized false positive hits. The enriched peptide binder P12 emerged as substrate (d-glyceraldehyde 3-phosphate) competitive against Deinococcus radiodurans DXS. The result indicates possible overlapping of cofactor- and acceptor-substrate-binding pockets, and provides inspiration for the design of inhibitors of DXS with a unique and novel mechanism...
November 8, 2017: Chembiochem: a European Journal of Chemical Biology
https://www.readbyqxmd.com/read/29119245/development-and-characterisation-of-a-novel-glucagon-like-peptide-1-receptor-antibody
#9
Emma K Biggs, Lihuan Liang, Jacqueline Naylor, Shimona Madalli, Rachel Collier, Matthew P Coghlan, David J Baker, David C Hornigold, Peter Ravn, Frank Reimann, Fiona M Gribble
AIMS/HYPOTHESIS: Glucagon like peptide-1 (GLP-1) enhances glucose-dependent insulin secretion by binding to GLP-1 receptors (GLP1Rs) on pancreatic beta cells. GLP-1 mimetics are used in the clinic for the treatment of type 2 diabetes, but despite their therapeutic success, several clinical effects of GLP-1 remain unexplained at a mechanistic level, particularly in extrapancreatic tissues. The aim of this study was to generate and characterise a monoclonal antagonistic antibody for the GLP1R for use in vivo...
November 9, 2017: Diabetologia
https://www.readbyqxmd.com/read/29118372/human-single-chain-transbodies-that-bound-to-domain-i-of-non-structural-protein-5a-ns5a-of-hepatitis-c-virus
#10
Kittirat Glab-Ampai, Monrat Chulanetra, Aijaz Ahmad Malik, Thanate Juntadech, Jeeraphong Thanongsaksrikul, Potjanee Srimanote, Kanyarat Thueng-In, Nitat Sookrung, Pongsri Tongtawe, Wanpen Chaicumpa
A safe and broadly effective direct acting anti-hepatitis C virus (HCV) agent that can withstand the viral mutation is needed. In this study, human single chain antibody variable fragments (HuscFvs) to conserved non-structural protein-5A (NS5A) of HCV were produced by phage display technology. Recombinant NS5A was used as bait for fishing-out the protein bound-phages from the HuscFv-phage display library. NS5A-bound HuscFvs produced by five phage transfected-E. coli clones were linked molecularly to nonaarginine (R9) for making them cell penetrable (become transbodies)...
November 8, 2017: Scientific Reports
https://www.readbyqxmd.com/read/29116526/phagekines-screening-binding-properties-and-biological-activity-of-functional-cytokines-displayed-on-phages
#11
Gertrudis Rojas, Tania Carmenate
The current chapter focuses on the use of filamentous phages to display, modify, and characterize cytokines, which are proteins belonging to a versatile group of essential mediators involved in cell-cell communication. Cytokines exhibit a considerable diversity, both in functions and in structural features underlying their biological effects. A broad variety of cytokines have been successfully displayed on phages, allowing the high-throughput study of their binding properties and biological activities and the discovery of novel therapeutics through directed evolution...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116525/metasecretome-phage-display
#12
Milica Ciric, Filomena Ng, Jasna Rakonjac, Dragana Gagic
Metasecretome is a collection of cell-surface and secreted proteins that mediate interactions between microbial communities and their environment. These include adhesins, enzymes, surface structures such as pili or flagella, vaccine targets or proteins responsible for immune evasion. Traditional approaches to exploring matasecretome of complex microbial communities via cultivation of microorganisms and screening of individual strains fail to sample extraordinary diversity in these communities, since only a limited fraction of microorganisms are represented by cultures...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116524/epitope-mapping-by-phage-display
#13
Gustavo Marçal Schmidt Garcia Moreira, Viola Fühner, Michael Hust
Among the molecules of the immune system, antibodies, particularly monoclonal antibodies (mAbs), have been shown to be interesting for many biological applications. Due to their ability to recognize only a unique part of their target, mAbs are usually very specific. These targets can have many different compositions, but the most common ones are proteins or peptides that are usually from outside the host, although self-proteins can also be targeted in autoimmune diseases, or in some types of cancer. The parts of a mAb that interact with its target compose the paratope, while the recognized parts of the target compose the epitope...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116523/orfeome-phage-display
#14
Jonas Zantow, Gustavo Marçal Schmidt Garcia Moreira, Stefan Dübel, Michael Hust
ORFeome phage display allows the efficient functional screening of entire proteomes or even metaproteomes to identify immunogenic proteins. For this purpose, randomly fragmented, whole genomes or metagenomes are cloned into a phage-display vector allowing positive selection for open reading frames (ORF) to improve the library quality. These libraries display all possible proteins encoded by a pathogen or a microbiome on the phage surface. Consequently, immunogenic proteins can be selected from these libraries using disease-related immunoglobulins from patient serum...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116522/monitoring-phage-biopanning-by-next-generation-sequencing
#15
Anna Vaisman-Mentesh, Yariv Wine
Phage display has enabled the rapid isolation of antigen-specific antibodies from combinatorial libraries of the variable heavy chain gene (VH) and variable light chain gene (VL). The method is based on genetic engineering of bacteriophages and repeated rounds of antigen-guided selection by phage biopanning.Next-Generation Sequencing (NGS) coupled with bioinformatics are powerful tools for analyzing the large number of DNA sequences present in an immune library.Here, we describe a method that demonstrates how NGS analysis enhances phage biopanning of complex antibody libraries as well as facilitates the antibody discovery process...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116521/high-throughput-igg-reformatting-and-expression
#16
Chao-Guang Chen, Georgina Sansome, Michael J Wilson, Con Panousis
We have recently described a one-step zero-background IgG reformatting method that enables the rapid reformatting of phage-displayed antibody fragments into a single-mammalian cell expression vector for full IgG expression (Chen et al. Nucleic Acids Res 42:e26, 2014). The strategy utilizes our unique positive selection method, referred to as insert-tagged (InTag) positive selection, where a positive selection marker (e.g. chloramphenicol-resistance gene) is cloned together with the antibody inserts into the expression vector...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116520/next-generation-dna-sequencing-of-vh-vl-repertoires-a-primer-and-guide-to-applications-in-single-domain-antibody-discovery
#17
Kevin A Henry
Immunogenetic analyses of expressed antibody repertoires are becoming increasingly common experimental investigations and are critical to furthering our understanding of autoimmunity, infectious disease, and cancer. Next-generation DNA sequencing (NGS) technologies have now made it possible to interrogate antibody repertoires to unprecedented depths, typically by sequencing of cDNAs encoding immunoglobulin variable domains. In this chapter, we describe simple, fast, and reliable methods for producing and sequencing multiplex PCR amplicons derived from the variable regions (VH, VHH or VL) of rearranged immunoglobulin heavy and light chain genes using the Illumina MiSeq platform...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116518/antibody-affinity-and-stability-maturation-by-error-prone-pcr
#18
Tobias Unkauf, Michael Hust, André Frenzel
Antibodies are the fastest growing class of pharmaceutical proteins and essential tools for research and diagnostics. Often antibodies do show a desirable specificity profile but lack sufficient affinity for the desired application. Here, we describe a method to increase the affinity of recombinant antibody fragments based on the construction of mutagenized phage display libraries.After the construction of a mutated antibody gene library by error-prone PCR, selection of high-affinity variants is either performed by panning in solution or on immobilized antigen with washing conditions optimized for off-rate-dependent selection...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116517/antibody-selection-on-ffpe-tissue-slides
#19
Andre Ten Haaf, Stefan Gattenlöhner, Mehmet Kemal Tur
Standard antibody phage-display panning uses purified proteins, antigen-transfected cells, or tumor cell lines as target structure to generate specific antibodies. Here, we describe a method for the selection of specific antibodies by phage panning against routine formalin-fixed paraffin-embedded (FFPE) tissue biopsies immobilized on glass slides. Selected antibody fragments recognize disease-associated antigens in its native conformation, suitable for the development of targeted diagnostic and therapeutic agents...
2018: Methods in Molecular Biology
https://www.readbyqxmd.com/read/29116516/screening-phage-display-antibody-libraries-using-protein-arrays
#20
Ricardo Jara-Acevedo, Paula Díez, María González-González, Rosa María Dégano, Nieves Ibarrola, Rafael Góngora, Alberto Orfao, Manuel Fuentes
Phage-display technology constitutes a powerful tool for the generation of specific antibodies against a predefined antigen. The main advantages of phage-display technology in comparison to conventional hybridoma-based techniques are: (1) rapid generation time and (2) antibody selection against an unlimited number of molecules (biological or not). However, the main bottleneck with phage-display technology is the validation strategies employed to confirm the greatest number of antibody fragments. The development of new high-throughput (HT) techniques has helped overcome this great limitation...
2018: Methods in Molecular Biology
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