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Use of a Synthetic Biosensor for Neutralizing Activity-Biased Selection of Monoclonal Antibodies against Atroxlysin-I, an Hemorrhagic Metalloproteinase from Bothrops atrox Snake Venom
- Source :
- PLoS Neglected Tropical Diseases, PLoS Neglected Tropical Diseases, Public Library of Science, 2014, 8 (4), pp.e2826. ⟨10.1371/journal.pntd.0002826⟩, PLoS Neglected Tropical Diseases, Vol 8, Iss 4, p e2826 (2014)
- Publication Year :
- 2014
- Publisher :
- HAL CCSD, 2014.
-
Abstract
- Background The snake Bothrops atrox is responsible for the majority of envenomings in the northern region of South America. Severe local effects, including hemorrhage, which are mainly caused by snake venom metalloproteinases (SVMPs), are not fully neutralized by conventional serum therapy. Little is known about the immunochemistry of the P-I SVMPs since few monoclonal antibodies (mAbs) against these molecules have been obtained. In addition, producing toxin-neutralizing mAbs remains very challenging. Methodology/Principal Findings Here, we report on the set-up of a functional screening based on a synthetic peptide used as a biosensor to select neutralizing mAbs against SVMPs and the successful production of neutralizing mAbs against Atroxlysin-I (Atr-I), a P-I SVMP from B. atrox. Hybridomas producing supernatants with inhibitory effect against the proteolytic activity of Atr-I towards the FRET peptide Abz-LVEALYQ-EDDnp were selected. Six IgG1 Mabs were obtained (named mAbatr1 to mAbatr6) and also two IgM. mAbatrs1, 2, 3 and 6 were purified. All showed a high specific reactivity, recognizing only Atr-I and B. atrox venom in ELISA and a high affinity, showing equilibrium constants in the nM range for Atr-I. These mAbatrs were not able to bind to Atr-I overlapping peptides, suggesting that they recognize conformational epitopes. Conclusions/Significance For the first time a functional screening based on a synthetic biosensor was successfully used for the selection of neutralizing mAbs against SVMPs.<br />Author Summary In this work, we propose a new screening strategy to produce monoclonal antibodies against Atr-I, a P-I class SVMP from Bothrops atrox, which is the snake responsible for the majority of the accidents in South America. SVMPs are the main toxic factors in Bothrops venom causing systemic and local hemorrhage, which may evolve to inflammation and/or necrosis. Since the toxic effects of SVMPs are related to their proteolytic activity, we have produced a peptide which was used as a biosensor for Atr-I hydrolysis. Hydrolysis of this substrate was monitored and the clones possessing inhibitory activity against the proteolytic activity of Atr-I upon the peptide were selected. Using our new approach, we have obtained four monoclonal antibodies highly specific and with neutralizing capacity against the hemorrhagic activity of either Atr-I alone or Bothrops atrox whole venom. To the best of the authors' knowledge, this is the first time where a functional screening is used for the selection of neutralizing mAbs against SVMPs. It is also the first description of mAbs anti-Atr-I, with inhibitory potential against its toxic activities which may be useful for diagnosis and treatment in the future.
- Subjects :
- lcsh:Arctic medicine. Tropical medicine
lcsh:RC955-962
medicine.drug_class
[SDV]Life Sciences [q-bio]
Immunology
Peptide
Venom
Biosensing Techniques
Monoclonal antibody
Biochemistry
Epitope
Immunochemistry
Fluorescence Resonance Energy Transfer
medicine
Animals
Humans
Mass Screening
Bothrops
Mass screening
ComputingMilieux_MISCELLANEOUS
chemistry.chemical_classification
biology
lcsh:Public aspects of medicine
Immunity
Public Health, Environmental and Occupational Health
Antibodies, Monoclonal
Metalloendopeptidases
Biology and Life Sciences
lcsh:RA1-1270
South America
biology.organism_classification
Antibodies, Neutralizing
Vaccination and Immunization
Molecular biology
3. Good health
Infectious Diseases
chemistry
Snake venom
Humoral Immunity
Clinical Immunology
Antitoxins
Immunotherapy
Peptides
Snake Venoms
Research Article
Subjects
Details
- Language :
- English
- ISSN :
- 19352727 and 19352735
- Database :
- OpenAIRE
- Journal :
- PLoS Neglected Tropical Diseases, PLoS Neglected Tropical Diseases, Public Library of Science, 2014, 8 (4), pp.e2826. ⟨10.1371/journal.pntd.0002826⟩, PLoS Neglected Tropical Diseases, Vol 8, Iss 4, p e2826 (2014)
- Accession number :
- edsair.doi.dedup.....7bedc1ad2053e1fa8dd47487bb2001f5