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Marylène Vandevenne, Mathieu Dondelinger, Sami Yunus, Astrid Freischels, Régine Freischels, Oscar Crasson, Noureddine Rhazi, Pierre Bogaerts, Moreno Galleni, Patrice Filée
Biosensors are becoming increasingly important and implemented in various fields such as pathogen detection, molecular diagnosis, environmental monitoring, and food safety control. In this context, we used β-lactamases as efficient reporter enzymes in several protein-protein interaction studies. Furthermore, their ability to accept insertions of peptides or structured proteins/domains strongly encourages the use of these enzymes to generate chimeric proteins. In a recent study, we inserted a single-domain antibody fragment into the Bacillus licheniformis BlaP β-lactamase...
February 1, 2018: Journal of Visualized Experiments: JoVE
Masahito Yamagata, Joshua R Sanes
Sensitive and specific antibodies are essential for detecting molecules in cells and tissues. However, currently used polyclonal and monoclonal antibodies are often less specific than desired, difficult to produce, and available in limited quantities. A promising recent approach to circumvent these limitations is to employ chemically defined antigen-combining domains called "nanobodies," derived from single-chain camelid antibodies. Here, we used nanobodies to prepare sensitive unimolecular detection reagents by genetically fusing cDNAs encoding nanobodies to enzymatic or antigenic reporters...
February 13, 2018: Proceedings of the National Academy of Sciences of the United States of America
Sunanda Singh, Genoveva Murillo, Dong Chen, Ashutosh S Parihar, Rajendra G Mehta
Background: The serendipitous discovery of heavy-chain antibodies devoid of light chains in camelids and the subsequent development of VHHs (variable region of camelid heavy chain) have provided a very important tool for research and possibly for therapeutics. In this study, we synthesized single-domain 15-kDa antibody SBT-100 (anti-STAT3 B VHH13) against human STAT3 (signal transducer and activator of transcription) that binds selectively to STAT3 and suppresses the function of phosphorylated STAT3 (p-STAT3)...
2018: Breast Cancer: Basic and Clinical Research
Conor McMahon, Alexander S Baier, Roberta Pascolutti, Marcin Wegrecki, Sanduo Zheng, Janice X Ong, Sarah C Erlandson, Daniel Hilger, Søren G F Rasmussen, Aaron M Ring, Aashish Manglik, Andrew C Kruse
Camelid single-domain antibody fragments ('nanobodies') provide the remarkable specificity of antibodies within a single 15-kDa immunoglobulin V HH domain. This unique feature has enabled applications ranging from use as biochemical tools to therapeutic agents. Nanobodies have emerged as especially useful tools in protein structural biology, facilitating studies of conformationally dynamic proteins such as G-protein-coupled receptors (GPCRs). Nearly all nanobodies available to date have been obtained by animal immunization, a bottleneck restricting many applications of this technology...
February 12, 2018: Nature Structural & Molecular Biology
B Schauer, R Krametter-Frötscher, F Knauer, R Ehricht, S Monecke, A T Feßler, S Schwarz, T Grunert, J Spergser, I Loncaric
The aim of this study was to determine the prevalence, the antimicrobial resistance patterns and the genetic diversity of methicillin-resistant Staphylococcus aureus (MRSA) from Austrian ruminants and New World camelids that were treated at the University of Veterinary Medicine, Vienna. Between April 2014 and January 2017, 723 nasal swabs originating from ruminants and New World camelids were examined. MRSA isolates were characterized by mecA/mecA1/mecC PCRs and by DNA microarray analysis. They were genotyped by spa typing, dru typing, MLST and MLVA...
February 2018: Veterinary Microbiology
Amanda Sparkes, Patrick De Baetselier, Lea Brys, Inês Cabrito, Yann G-J Sterckx, Steve Schoonooghe, Serge Muyldermans, Geert Raes, Richard Bucala, Peter Vanlandschoot, Jo A Van Ginderachter, Benoît Stijlemans
Sepsis-leading to septic shock-is the leading cause of death in intensive care units. The systemic inflammatory response to infection, which is initiated by activated myeloid cells, plays a key role in the lethal outcome. Macrophage migration inhibitory factor (MIF) is an upstream immunoregulatory mediator, released by myeloid cells, that underlies a common genetic susceptibility to different infections and septic shock. Accordingly, strategies that are aimed at inhibiting the action of MIF have therapeutic potential...
January 25, 2018: FASEB Journal: Official Publication of the Federation of American Societies for Experimental Biology
Naoya Shinozaki, Ryuji Hashimoto, Masanori Noda, Susumu Uchiyama
Recently, we showed that immunized rabbit heavy chain variable regions (rVHs) can have strong antigen binding activity comparable to that of the camelid variable domain of the heavy chain of heavy chain antibody (VHH). These rVHs lack the light chain variable regions (rVLs), which exist in the authentic Fab format; thus, molecular surfaces at the interface region of rVHs are exposed to solvent. This physical feature may change physicochemical properties, such as causing reduced stability. By overcoming potential physicochemical issues through engineering the interface region, rVHs could become more useful as single-domain antibodies...
February 1, 2018: Journal of Bioscience and Bioengineering
Traian Sulea, Greg Hussack, Shannon Ryan, Jamshid Tanha, Enrico O Purisima
Assisted Design of Antibody and Protein Therapeutics (ADAPT) is an affinity maturation platform interleaving predictions and testing that was previously validated on monoclonal antibodies (mAbs). This study expands the applicability of ADAPT to single-domain antibodies (sdAbs), a promising class of recombinant antibody-based biologics. As a test case, we used the camelid sdAb A26.8, a VHH that binds Clostridium difficile toxin A (TcdA) relatively weakly but displays a reasonable level of TcdA neutralization...
February 2, 2018: Scientific Reports
Polina L Perelman, Rudolf Pichler, Anna Gaggl, Denis M Larkin, Terje Raudsepp, Fahad Alshanbari, Heather M Holl, Samantha A Brooks, Pamela A Burger, Kathiravan Periasamy
The availability of genomic resources including linkage information for camelids has been very limited. Here, we describe the construction of a set of two radiation hybrid (RH) panels (5000RAD and 15000RAD) for the dromedary (Camelus dromedarius) as a permanent genetic resource for camel genome researchers worldwide. For the 5000RAD panel, a total of 245 female camel-hamster radiation hybrid clones were collected, of which 186 were screened with 44 custom designed marker loci distributed throughout camel genome...
January 31, 2018: Scientific Reports
Abdullah Hoter, Mahdi Amiri, Abdelbary Prince, Hassan Amer, Mohamad Warda, Hassan Y Naim
Increased expression of heat shock proteins (HSPs) following heat stress or other stress conditions is a common physiological response in almost all living organisms. Modification of cytosolic proteins including HSPs by O-GlcNAc has been shown to enhance their capabilities for counteracting lethal levels of cellular stress. Since HSPs are key players in stress resistance and protein homeostasis, we aimed to analyze their forms at the cellular and molecular level using camel and human HSPs as models for efficient and moderate thermotolerant mammals, respectively...
January 30, 2018: International Journal of Molecular Sciences
Ryota Saito, Yutaro Saito, Hikaru Nakazawa, Takamitsu Hattori, Izumi Kumagai, Mitsuo Umetsu, Koki Makabe
Biomolecules which recognize inorganic materials and metal surfaces gain much attention for creating new type of nanomaterials and sensors. 4F2, a camelid VHH antibody, recognizes ZnO surface and has been applied for sensor applications. 4F2 was constructed sequential CDR replacement on the parental VHH antibody, termed the Construction of Antibody by Integrating Grafting and Evolution Technology; CAnIGET procedure. Here, we evaluate the influence of CDR replacements during 4F2 generation using calorimetric technique...
January 22, 2018: Journal of Biochemistry
Antony S K Yerabham, Andreas Müller-Schiffmann, Tamar Ziehm, Andreas Stadler, Sabrina Köber, Xela Indurkhya, Rita Marreiros, Svenja V Trossbach, Nicholas J Bradshaw, Ingrid Prikulis, Dieter Willbold, Oliver H Weiergräber, Carsten Korth
Accumulating evidence suggests an important role for the Disrupted-in-Schizophrenia 1 (DISC1) protein in neurodevelopment and chronic mental illness. In particular, the C-terminal 300 amino acids of DISC1 have been found to mediate important protein-protein interactions and to harbor functionally important phosphorylation sites and disease-associated polymorphisms. However, long disordered regions and oligomer-forming subdomains have so far impeded structural analysis. VHH domains derived from camelid heavy chain only antibodies are minimal antigen binding modules with appreciable solubility and stability, which makes them well suited for the stabilizing proteins prior to structural investigation...
2018: PloS One
Ellen R Goldman, Aeris Broussard, George P Anderson, Jinny L Liu
Single domain antibodies, recombinantly expressed variable domains derived from camelid heavy chain antibodies, are often expressed as multimers for detection and therapeutic applications. Constructs in which several single domain antibodies are genetically fused serially, as well as those in which single domain antibodies are genetically linked with domains that naturally form multimers, yield improvement in apparent binding affinity due to avidity. Here, using a single domain antibody that binds envelope protein from the Dengue virus, we demonstrated the construction of single domain antibody dimers using the Bglbrick cloning strategy...
December 2017: Heliyon
Somayeh Bahrami, Mohammad Reza Tabandeh, Ali Reza Ganjali Tafreshi
Camels ( Camelus dromedarius) are important, multipurpose local animals in Iran. Despite their importance, camelid parasitic diseases have not received adequate attention in the veterinary literature. The present study investigated the prevalence of, and molecularly identified, camel piroplasms in Iran. Blood samples from 248 camels from five different regions of Iran were screened for the presence of piroplasmid infection using an 18SrRNA polymerase chain reaction (PCR) sequencing method. Of the 248 samples, 16 were positive for piroplasms via PCR (6...
December 2017: Journal of Zoo and Wildlife Medicine: Official Publication of the American Association of Zoo Veterinarians
Alexandra Hund, Thomas Wittek
Abomasal ulcers are frequent incidental findings in necropsies of domestic ruminants and South American Camelids (SAC) or in slaughter animals and are a frequent cause of death in the most affected group of cattle, veal calves. Their true prevalence and significance is unknown owing to limitations in diagnosing the condition in live animals. This article discusses types of ulcers, possible causes of ulceration, and the clinical consequences, symptoms, and differential diagnoses, as well as further diagnostics in cattle, small ruminants and SAC...
December 21, 2017: Veterinary Clinics of North America. Food Animal Practice
Nesreen Allam Tantawy Allam, Doaa Sedky, Enshrah Khalil Mira
Background and Aim: Recently, cases of mastitis refractory to treatment have been reported frequently. There are limited routine laboratory investigations on Camelidae infections. Mastitis has been estimated to affect more than 25% of lactating she-camel with up to 70% milk loss. The details of Bacillus spp. pathogenesis in mastitis are not yet fully described. The present study is the first detailed phenotypic and genotypic characterization of Bacillus licheniformis isolates from recurrent mastitic she-camels with sepsis in Egypt...
November 2017: Veterinary World
Christopher Bachran, Matthias Schröder, Lena Conrad, Juan J Cragnolini, Fikadu G Tafesse, Laura Helming, Hidde L Ploegh, Lee Kim Swee
The central role of myeloid cells in driving autoimmune diseases and cancer has raised interest in manipulating their function or depleting them for therapeutic benefits. To achieve this, antibodies are used to antagonize differentiation, survival and polarization signals or to kill target cells, for example in the form of antibody-drug conjugates (ADC). The action of ADC in vivo can be hard to predict based on target expression pattern alone. The biology of the targeted receptor as well as its interplay with the ADC can have drastic effects on cell apoptosis versus survival...
December 20, 2017: Scientific Reports
Ahmed Tibary
This paper reviews that state of our knowledge concerning follicular wave dynamics, monitoring and manipulation. All camelids have overlapping follicular waves in absence of ovulation which is induced by a seminal plasma factor (βNGF). The interval between follicular waves varies. The size of the ovulatory follicle varies between 11 and 25 mm in camels and between in 6 and 13 mm in South American Camelids. The interval between induction of ovulation and next ovulatory follicle is 15 ± 1 day for all camelids...
December 8, 2017: Theriogenology
Rasoul Baharlou, Nader Tajik, Mahdi Behdani, Mohammad Ali Shokrgozar, Amir Hassan Zarnani, Fatemeh Shahhosseini, Mahdi Habibi-Anbouhi
Antibody-based targeting of angiogenesis is a key approach for cancer treatment. Delta-like ligand 4 (DLL4) plays a pivotal role in tumor neovascular development and angiogenesis during tumor progression. It forecasts the prognosis of human malignancies and blocking its signaling can help to inhibit neovascularization and tumor metastasis. Nanobodies are the smallest antigen-binding domains of heavy chain antibodies in camelidae. The aim of this study was to develop a Nanobody against DLL4 and apply binding and functional approaches to target it...
December 12, 2017: Analytical Biochemistry
Jinny L Liu, Lisa C Shriver-Lake, George P Anderson, Dan Zabetakis, Ellen R Goldman
BACKGROUND: A key advantage of recombinant antibody technology is the ability to optimize and tailor reagents. Single domain antibodies (sdAbs), the recombinantly produced variable domains derived from camelid and shark heavy chain antibodies, provide advantages of stability and solubility and can be further engineered to enhance their properties. In this study, we generated sdAbs specific for Ebola virus envelope glycoprotein (GP) and increased their stability to expand their utility for use in austere locals...
December 12, 2017: Microbial Cell Factories
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