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https://www.readbyqxmd.com/read/28334940/aav9-delivered-bispecific-nanobody-attenuates-amyloid-burden-in-the-gelsolin-amyloidosis-mouse-model
#1
Adriaan Verhelle, Nisha Nair, Inge Everaert, Wouter Van Overbeke, Lynn Supply, Olivier Zwaenepoel, Cindy Peleman, Jo Van Dorpe, Tony Lahoutte, Nick Devoogdt, Wim Derave, Marinee K Chuah, Thierry Vanden Driessche, Jan Gettemans
Gelsolin amyloidosis is a dominantly inherited, incurable type of amyloidosis. A single point mutation in the gelsolin gene (G654A is most common) results in the loss of a Ca2+  binding site in the second gelsolin domain. Consequently, this domain partly unfolds and exposes an otherwise buried furin cleavage site at the surface. During secretion of mutant plasma gelsolin consecutive cleavage by furin and MT1-MMP results in the production of 8 and 5 kDa amyloidogenic peptides. Nanobodies that are able to (partly) inhibit furin or MT1-MMP proteolysis have previously been reported...
February 21, 2017: Human Molecular Genetics
https://www.readbyqxmd.com/read/28332258/n-terminal-chemical-protein-labeling-using-the-naturally-split-gos-terl-intein
#2
Anne-Lena Bachmann, Henning D Mootz
Chemoselective and regioselective chemical protein labeling is of great importance, yet no current technique is sufficiently general and simple to perform. Protein trans-splicing by split inteins can be used to ligate short tags with chemical labels to either the N or the C terminus of a protein. The CysTag approach exploits split intein fragments without a cysteine fused with such a short tag containing a single cysteine that is easily amenable to selective modification using classical cysteine bioconjugation...
March 23, 2017: Journal of Peptide Science: An Official Publication of the European Peptide Society
https://www.readbyqxmd.com/read/28331319/humanized-cd7-nanobody-based-immunotoxins-exhibit-promising-anti-t-cell-acute-lymphoblastic-leukemia-potential
#3
Yuan Yu, Jialu Li, Xuejun Zhu, Xiaowen Tang, Yangyi Bao, Xiang Sun, Yuhui Huang, Fang Tian, Xiaomei Liu, Lin Yang
BACKGROUND: Nanobodies, named as VHHs (variable domain of heavy chain of HCAb [heavy-chain antibodies]), are derived from heavy-chain-only antibodies that circulate in sera of camelids. Their exceptional physicochemical properties, possibility of humanization, and unique antigen recognition properties make them excellent candidates for targeted delivery of biologically active components, including immunotoxins. In our previous efforts, we have successfully generated the monovalent and bivalent CD7 nanobody-based immunotoxins, which can effectively trigger the apoptosis of CD7-positive malignant cells...
2017: International Journal of Nanomedicine
https://www.readbyqxmd.com/read/28322225/probing-cytoskeletal-modulation-of-passive-and-active-intracellular-dynamics-using-nanobody-functionalized-quantum-dots
#4
Eugene A Katrukha, Marina Mikhaylova, Hugo X van Brakel, Paul M van Bergen En Henegouwen, Anna Akhmanova, Casper C Hoogenraad, Lukas C Kapitein
The cytoplasm is a highly complex and heterogeneous medium that is structured by the cytoskeleton. How local transport depends on the heterogeneous organization and dynamics of F-actin and microtubules is poorly understood. Here we use a novel delivery and functionalization strategy to utilize quantum dots (QDs) as probes for active and passive intracellular transport. Rapid imaging of non-functionalized QDs reveals two populations with a 100-fold difference in diffusion constant, with the faster fraction increasing upon actin depolymerization...
March 21, 2017: Nature Communications
https://www.readbyqxmd.com/read/28316235/evaluation-of-nanobody-conjugates-and-protein-fusions-as-bioanalytical-reagents
#5
Virginia J Bruce, Brian R McNaughton
Enzyme-linked immunosorbent assay (ELISA), flow cytometry, and Western blot are common bioanalytical techniques. Successful execution traditionally requires the use of one or more commercially available antibody-small-molecule dye, or antibody-reporter protein conjugates that recognize relatively short peptide tags (<15 amino acids). However, the size of antibodies, and their molecular complexity (by virtue of post-translational disulfide formation and glycosylation) typically requires either expression in mammalian cells or purification from immunized mammals...
March 20, 2017: Analytical Chemistry
https://www.readbyqxmd.com/read/28301262/-213-bi-labeled-prostate-specific-membrane-antigen-targeting-agents-induce-dna-double-strand-breaks-in-prostate-cancer-xenografts
#6
Julie Nonnekens, Kristell L S Chatalic, Janneke D M Molkenboer-Kuenen, Cecile E M T Beerens, Frank Bruchertseifer, Alfred Morgenstern, Joke Veldhoven-Zweistra, Margret Schottelius, Hans-Jürgen Wester, Dik C van Gent, Wytske M van Weerden, Otto C Boerman, Marion de Jong, Sandra Heskamp
BACKGROUND: Up to now, prostate-specific membrane antigen (PSMA)-targeted radionuclide therapy mainly focused on β-emitting radionuclides. Herein, two new (213)Bi-labeled agents for PSMA-targeted α therapy of prostate cancer (PCa) are reported. METHODS: The biodistribution of (213)Bi-labeled small-molecule inhibitor PSMA I&T and nanobody JVZ-008 was evaluated in mice bearing PSMA-positive LNCaP xenografts. DNA damage response was followed using LNCaP cells and LNCaP xenografts...
March 2017: Cancer Biotherapy & Radiopharmaceuticals
https://www.readbyqxmd.com/read/28300394/an-asymmetric-conformational-change-in-lacy
#7
Irina Smirnova, Vladimir Kasho, Xiaoxu Jiang, H Ronald Kaback
Galactoside/H(+) symport by the lactose permease of Escherichia coli (LacY) involves reciprocal opening and closing of periplasmic and cytoplasmic cavities so that sugar- and H(+)-binding sites become alternatively accessible to either side of the membrane. After reconstitution into proteoliposomes, LacY with the periplasmic cavity sealed by cross-linking paired-Cys residues does not bind sugar from the periplasmic side. However, reduction of the S-S bond restores opening of the periplasmic cavity and galactoside binding...
March 23, 2017: Biochemistry
https://www.readbyqxmd.com/read/28283558/gene-expression-profiling-with-cre-conditional-pseudorabies-virus-reveals-a-subset-of-midbrain-neurons-that-participate-in-reward-circuitry
#8
Lisa E Pomeranz, Mats I Ekstrand, Kaamashri N Latcha, Gregory A Smith, Lynn W Enquist, Jeffrey M Friedman
The mesolimbic dopamine pathway receives inputs from numerous regions of the brain as part of a neural system that detects rewarding stimuli and coordinates a behavioral response. The capacity to simultaneously map and molecularly define the components of this complex multisynaptic circuit would thus advance our understanding of the determinants of motivated behavior. To accomplish this, we have constructed pseudorabies virus (PRV) strains in which viral propagation and fluorophore expression are activated only after exposure to Cre recombinase...
March 10, 2017: Journal of Neuroscience: the Official Journal of the Society for Neuroscience
https://www.readbyqxmd.com/read/28280600/structural-basis-of-a-novel-pd-l1-nanobody-for-immune-checkpoint-blockade
#9
Fei Zhang, Hudie Wei, Xiaoxiao Wang, Yu Bai, Pilin Wang, Jiawei Wu, Xiaoyong Jiang, Yugang Wang, Haiyan Cai, Ting Xu, Aiwu Zhou
The use of antibodies to target immune checkpoints, particularly PD-1/PD-L1, has made a profound impact in the field of cancer immunotherapy. Here, we identified KN035, an anti-PD-L1 nanobody that can strongly induce T-cell responses and inhibit tumor growth. The crystal structures of KN035 complexed with PD-L1 and free PD-L1, solved here at 1.7 and 2.7 Å resolution, respectively, show that KN035 competes with PD-1 (programmed death protein 1) for the same flat surface on PD-L1, mainly through a single surface loop of 21 amino acids...
2017: Cell Discovery
https://www.readbyqxmd.com/read/28277548/optimized-labeling-of-membrane-proteins-for-applications-to-super-resolution-imaging-in-confined-cellular-environments-using-monomeric-streptavidin
#10
Ingrid Chamma, Olivier Rossier, Grégory Giannone, Olivier Thoumine, Matthieu Sainlos
Recent progress in super-resolution imaging (SRI) has created a strong need to improve protein labeling with probes of small size that minimize the target-to-label distance, increase labeling density, and efficiently penetrate thick biological tissues. This protocol describes a method for labeling genetically modified proteins incorporating a small biotin acceptor peptide with a 3-nm fluorescent probe, monomeric streptavidin. We show how to express, purify, and conjugate the probe to organic dyes with different fluorescent properties, and how to label selectively biotinylated membrane proteins for SRI techniques (point accumulation in nanoscale topography (PAINT), stimulated emission depletion (STED), stochastic optical reconstruction microscopy (STORM))...
April 2017: Nature Protocols
https://www.readbyqxmd.com/read/28275529/enhanced-mrna-protein-fusion-efficiency-of-a-single-domain-antibody-by-selection-of-mrna-display-with-additional-random-sequences-in-the-terminal-translated-regions
#11
Kazuki Takahashi, Masato Sunohara, Takuya Terai, Shigefumi Kumachi, Naoto Nemoto
In vitro display technologies such as mRNA and cDNA display are powerful tools to create and select functional peptides. However, in some cases, efficiency of mRNA-protein fusion is very low, which results in decreased library size and lower chance of successful selection. In this study, to improve mRNA-protein fusion efficiency, we prepared an mRNA display library of a protein with random N- and C-terminal coding regions consisting of 12 nucleotides (i.e. four amino acids), and performed an electrophoresis mobility shift assay (EMSA)-based selection of successfully formed mRNA display molecules...
2017: Biophysics and Physicobiology
https://www.readbyqxmd.com/read/28274145/emerging-monoclonal-antibodies-against-clostridium-difficile-infection
#12
Séverine Péchiné, Claire Janoir, Anne Collignon
Clostridium difficile infections are characterized by a high recurrence rate despite antibiotic treatments and there is an urgent need to develop new treatments such as fecal transplantation and immonotherapy. Besides active immunotherapy with vaccines, passive immunotherapy has shown promise, especially with monoclonal antibodies. Areas covered: Herein, the authors review the different assays performed with monoclonal antibodies against C. difficile toxins and surface proteins to treat or prevent primary or recurrent episodes of C...
April 2017: Expert Opinion on Biological Therapy
https://www.readbyqxmd.com/read/28245129/effect-of-dye-and-conjugation-chemistry-on-the-biodistribution-profile-of-near-infrared-labeled-nanobodies-as-tracers-for-image-guided-surgery
#13
Pieterjan Debie, Jannah Van Quathem, Inge Hansen, Gezim Bala, Sam Massa, Nick Devoogdt, Catarina Xavier, Sophie Hernot
Advances in optical imaging technologies have stimulated the development of near-infrared (NIR) fluorescently labeled targeted probes for use in image-guided surgery. As nanobodies have already proven to be excellent candidates for molecular imaging, we aimed in this project to design NIR-conjugated nanobodies targeting the tumor biomarker HER2 for future applications in this field and to evaluate the effect of dye and dye conjugation chemistry on their pharmacokinetics during development. IRDye800CW or IRdye680RD were conjugated either randomly (via lysines) or site-specifically (via C-terminal cysteine) to the anti-HER2 nanobody 2Rs15d...
March 9, 2017: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/28239762/-1-h-13-c-and-15-n-backbone-chemical-shift-assignments-of-camelid-single-domain-antibodies-against-active-state-%C3%A2%C2%B5-opioid-receptor
#14
Remy Sounier, Yinshan Yang, Joanna Hagelberger, Sébastien Granier, Hélène Déméné
Nanobodies are single chain antibodies that have become a highly valuable and versatile tool for biomolecular and therapeutic research. One application field is the stabilization of active states of flexible proteins, among which G-protein coupled receptors represent a very important class of membrane proteins. Here we present the backbone and side-chain assignment of the (1)H, (13)C and (15)N resonances of Nb33 and Nb39, two nanobodies that recognize and stabilize the µ-opioid receptor to opioids in its active agonist-bound conformation...
February 26, 2017: Biomolecular NMR Assignments
https://www.readbyqxmd.com/read/28238765/cytoplasmic-versus-periplasmic-expression-of-site-specifically-and-bioorthogonally-functionalized-nanobodies-using-expressed-protein-ligation
#15
Brecht Billen, Cécile Vincke, Rebekka Hansen, Nick Devoogdt, Serge Muyldermans, Peter Adriaensens, Wanda Guedens
Site-specific functionalization of nanobodies after introducing bioorthogonal groups offers the possibility to biofunctionalize surfaces with a uniformly oriented layer of nanobodies. In this paper, expressed protein ligation (EPL) was used for site-specific alkynation of the model nanobody NbBcII10. In contrast to EPL constructs, which are typically expressed in the cytoplasm, nanobodies are expressed in the periplasm where its oxidizing environment ensures a correct folding and disulfide bond formation. Different pathways were explored to express the EPL constructs in the periplasm but simultaneously, the effect of cytoplasmic expression on the functionality of NbBcII10 was also evaluated...
February 24, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28238060/novel-biotechnology-approaches-in-colorectal-cancer-diagnosis-and-therapy
#16
REVIEW
Soudabeh Kavousipour, Fathemeh Khademi, Mozhdeh Zamani, Bahareh Vakili, Pooneh Mokarram
With ever-increasing molecular information about colorectal cancer (CRC), there is an expectation to detect more sensitive and specific molecular markers for new advanced diagnostic methods that can surpass the limitations of current screening tests. Moreover, enhanced molecular pathology knowledge about cancer has led to the development of targeted therapies, designed to interfere with specific aberrant biological pathways in cancer. Furthermore, biotechnology has opened a new window in CRC diagnosis and treatment by introducing different application of antibodies, antibody fragments, non-Ig scaffold proteins, and aptamers in targeted therapy and drug delivery...
February 25, 2017: Biotechnology Letters
https://www.readbyqxmd.com/read/28235780/inhibitory-cortactin-nanobodies-delineate-the-role-of-nta-and-sh3-domain-specific-functions-during-invadopodium-formation-and-cancer-cell-invasion
#17
Laurence Bertier, Ciska Boucherie, Olivier Zwaenepoel, Berlinda Vanloo, Marleen Van Troys, Isabel Van Audenhove, Jan Gettemans
Cancer cells exploit different strategies to escape from the primary tumor, gain access to the circulation, disseminate throughout the body, and form metastases, the leading cause of death by cancer. Invadopodia, proteolytically active plasma membrane extensions, are essential in this escape mechanism. Cortactin is involved in every phase of invadopodia formation, and its overexpression is associated with increased invadopodia formation, extracellular matrix degradation, and cancer cell invasion. To analyze endogenous cortactin domain function in these processes, we characterized the effects of nanobodies that are specific for the N-terminal acidic domain of cortactin and expected to target small epitopes within this domain...
February 24, 2017: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
https://www.readbyqxmd.com/read/28235665/an-in-silico-approach-to-find-a-peptidomimetic-targeting-extracellular-domain-of-her3-from-a-her3-nanobody
#18
Z Pourhashem, M Mehrpouya, N Yardehnavi, A Eslamparast, F Kazemi-Lomedasht
HER3 is an important therapeutic target in cancer treatments. HER3 Nanobodies (Nbs) are a novel class of antibodies with several competitive advantages over conventional antibodies. A peptidomimetic derived from these Nbs can be considered to be a small peptide mimicking some of the molecular recognition interactions of a natural peptide or protein in a three-dimensional (3D) space, with a receptor that has improved properties. In this study, we introduce a new approach to design a peptidomimetic derived from HER3 Nb through an in silico analysis...
February 10, 2017: Computational Biology and Chemistry
https://www.readbyqxmd.com/read/28202688/super-resolution-mapping-of-scaffold-nucleoporins-in-the-nuclear-pore-complex
#19
Jiong Ma, Joseph M Kelich, Samuel L Junod, Weidong Yang
The nuclear pore complex (NPC), composed of ∼30 different nucleoporins (Nups), is one of the largest supramolecular structures in eukaryotic cells. Its octagonal ring-scaffold perforates the nuclear envelope and features a unique molecular machinery that regulates nucleocytoplasmic transport. However, the precise copy number and the spatial location of each Nup in the native NPC remain obscure due to the inherent difficulty of counting and localizing proteins inside the sub-micrometer supramolecular complex...
February 15, 2017: Journal of Cell Science
https://www.readbyqxmd.com/read/28202367/exosomes-therapy-delivery-tools-and-biomarkers-of-diseases
#20
REVIEW
Lucio Barile, Giuseppe Vassalli
Virtually all cells in the organism secrete extracellular vesicles (EVs), a heterogeneous population of lipid bilayer membrane-enclosed vesicles that transport and deliver payloads of proteins and nucleic acids to recipient cells, thus playing central roles in cell-cell communications. Exosomes, nanosized EVs of endosomal origin, regulate many pathophysiological processes including immune responses and inflammation, tumour growth, and infection. Healthy subjects and patients with different diseases release exosomes with different RNA and protein contents into the circulation, which can be measured as biomarkers...
February 12, 2017: Pharmacology & Therapeutics
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