scholarly journals Research on the Mechanism of Action of a Citrinin and Anti-Citrinin Antibody Based on Mimotope X27

Toxins ◽  
2020 ◽  
Vol 12 (10) ◽  
pp. 655
Author(s):  
Yanping Li ◽  
Yucheng Hu ◽  
Zhui Tu ◽  
Zhenqiang Ning ◽  
Qinghua He ◽  
...  
Keyword(s):  
Amino Acids ◽  
Molecular Docking ◽  
Mechanism Of Action ◽  
3D Structure ◽  
Antibody Fragment ◽  
Binding Activity ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Binding Model ◽  
Modeling Strategy

Immunoassays are developed based on antigen–antibody interactions. A mimotope is an effective recognition receptor used to study the mechanism of action of antigens and antibodies, and is used for improving the sensitivity of the antibody. In this study, we built a 3D structure of the citrinin (CIT) mimotope X27 and anti-CIT single-chain antibody fragment (ScFv) through a “homologous modeling” strategy. Then, CIT and X27 were respectively docked to anti-CIT ScFv by using the “molecular docking” program. Finally, T28, F29, N30, R31, and Y32 were confirmed as the key binding sites in X27. Furthermore, the result of the phage-ELISA showed that the mutational phage lost the binding activity to the anti-CIT ScFv when the five amino acids were mutated to “alanine”, thereby proving the correctness of the molecular docking model. Lastly, a site-directed saturation strategy was adopted for the sites (T28, F29, N30, R31, and Y32). Eighteen different amino acids were introduced to each site on average. The activities of all mutants were identified by indirect competitive ELISA. The sensitivities of mutants T28F, T28I, F29I, F29V, N30T, and N30V were 1.83-, 1.37-, 1.70-, 2.96-, 1.31-, and 2.01-fold higher than that of the wild-type, respectively. In conclusion, the binding model between the CIT and antibody was elaborated for the first time based on the mimotope method, thereby presenting another strategy for improving the sensitivity of citrinin detection in immunoassays.

scholarly journals Loxoscelism: Advances and Challenges in the Design of Antibody Fragments with Therapeutic Potential

Toxins ◽  
2020 ◽  
Vol 12 (4) ◽  
pp. 256
Author(s):  
Sabrina Karim-Silva ◽  
Alessandra Becker-Finco ◽  
Isabella Gizzi Jiacomini ◽  
Fanny Boursin ◽  
Arnaud Leroy ◽  
...  
Keyword(s):  
Monoclonal Antibody ◽  
Therapeutic Potential ◽  
Antibody Fragment ◽  
Binding Activity ◽  
Antibody Fragments ◽  
Antigen Binding ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Humanized Antibody ◽  
Neutralizing Capacity

Envenoming due to Loxosceles spider bites still remains a neglected disease of particular medical concern in the Americas. To date, there is no consensus for the treatment of envenomed patients, yet horse polyclonal antivenoms are usually infused to patients with identified severe medical conditions. It is widely known that venom proteins in the 30–35 kDa range with sphingomyelinase D (SMasesD) activity, reproduce most of the toxic effects observed in loxoscelism. Hence, we believe that monoclonal antibody fragments targeting such toxins might pose an alternative safe and effective treatment. In the present study, starting from the monoclonal antibody LimAb7, previously shown to target SMasesD from the venom of L. intermedia and neutralize its dermonecrotic activity, we designed humanized antibody V-domains, then produced and purified as recombinant single-chain antibody fragments (scFvs). These molecules were characterized in terms of humanness, structural stability, antigen-binding activity, and venom-neutralizing potential. Throughout this process, we identified some blocking points that can impact the Abs antigen-binding activity and neutralizing capacity. In silico analysis of the antigen/antibody amino acid interactions also contributed to a better understanding of the antibody’s neutralization mechanism and led to reformatting the humanized antibody fragment which, ultimately, recovered the functional characteristics for efficient in vitro venom neutralization.

scholarly journals Stimulation of Innate and Adaptive Immunity by Using Filamentous Bacteriophage fd Targeted to DEC-205

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Luciana D’Apice ◽  
Valerio Costa ◽  
Rossella Sartorius ◽  
Maria Trovato ◽  
Marianna Aprile ◽  
...  
Keyword(s):  
Immune Response ◽  
Dendritic Cells ◽  
Adaptive Immune Response ◽  
Antibody Fragment ◽  
Antigenic Determinants ◽  
Rna Seq ◽  
Single Chain ◽  
Single Chain Antibody ◽  
Filamentous Bacteriophage ◽  
Adaptive Immune

The filamentous bacteriophage fd, codisplaying antigenic determinants and a single chain antibody fragment directed against the dendritic cell receptor DEC-205, is a promising vaccine candidate for its safety and its ability to elicit innate and adaptive immune response in absence of adjuvants. By using a system vaccinology approach based on RNA-Sequencing (RNA-Seq) analysis, we describe a relevant gene modulation in dendritic cells pulsed with anti-DEC-205 bacteriophages fd. RNA-Seq data analysis indicates that the bacteriophage fd virions are sensed as a pathogen by dendritic cells; they activate the danger receptors that trigger an innate immune response and thus confer a strong adjuvanticity that is needed to obtain a long-lasting adaptive immune response.

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