Evaluation of three different formats of a neutralizing single chain human antibody against toxin Cn2: neutralization capacity versus thermodynamic stability.
The single-chain antibody fragment (scFv) 6009F, obtained by directed evolution, neutralizes the effects of the Cn2 toxin, which is the major toxic component of Centruroides noxius scorpion venom. In this work we compared the neutralization capacity and the thermodynamic stability of scFv 6009F with those of two other derived formats: Fab 6009F and diabody 6009F. Additionally, the affinity constants to Cn2 toxin of the three recombinant antibody fragments were determined by means of BIAcore. We found a correlation between the thermodynamic stability of these antibody fragments with their neutralization capacity. The order of thermodynamic stability determined was Fab≫scFv>diabody. The Fab and scFv were capable of neutralizing the toxic effects of Cn2 and whole venom but the diabody was unable to fully neutralize intoxication. In silico analysis of the diabody format indicates that the reduction of stability and neutralization capacity could be explained by a less cooperative interface between the heavy and the light variable domains.
Departamento de Medicina Molecular y Bioprocesos, Instituto de Biotecnología, UNAM, Apartado Postal 510-3, Cuernavaca, Morelos 62250, Mexico.
Veronica
V
Quintero-Hernández
Luis
L
Del Pozo-Yauner
Martha
M
Pedraza-Escalona
Victor R
VR
Juárez-González
Israel
I
Alcántara-Recillas
Lourival D
LD
Possani
Baltazar
B
Becerril
D000818:Animals; D057134:Antibodies, Neutralizing / Q000737:chemistry* / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology / Q000378:metabolism; D008958:Models, Molecular; D009500:Neutralization Tests; D011487:Protein Conformation; D017510:Protein Folding; D055550:Protein Stability; D012604:Scorpion Venoms / Q000276:immunology*; D057127:Single-Chain Antibodies / Q000737:chemistry* / Q000235:genetics / Q000276:immunology* / Q000378:metabolism; D013816:Thermodynamics
D057134:Antibodies, Neutralizing; D007140:Immunoglobulin Fab Fragments; D012604:Scorpion Venoms; D057127:Single-Chain Antibodies