Affinity is an important determinant of the anti-trypanosome activity of nanobodies.

TitleAffinity is an important determinant of the anti-trypanosome activity of nanobodies.
Publication TypeJournal Article
Year of Publication2012
AuthorsCaljon, G., B. Stijlemans, D. Saerens, J. Van Den Abbeele, S. Muyldermans, S. Magez, and P. De Baetselier
JournalPLoS Negl Trop Dis
Volume6
Issue11
Paginatione1902
Date Published2012
Type of Articleparasites
ISSN1935-2735
KeywordsAnimals, Antibodies, Protozoan, Antibody Affinity, Mice, Mice, Inbred C57BL, Microbial Viability, Single-Domain Antibodies, Trypanosoma brucei brucei
Abstract

BACKGROUND: The discovery of Nanobodies (Nbs) with a direct toxic activity against African trypanosomes is a recent advancement towards a new strategy against these extracellular parasites. The anti-trypanosomal activity relies on perturbing the highly active recycling of the Variant-specific Surface Glycoprotein (VSG) that occurs in the parasite's flagellar pocket.METHODOLOGY/PRINCIPAL FINDINGS: Here we expand the existing panel of Nbs with anti-Trypanosoma brucei potential and identify four categories based on their epitope specificity. We modified the binding properties of previously identified Nanobodies Nb_An05 and Nb_An33 by site-directed mutagenesis in the paratope and found this to strongly affect trypanotoxicity despite retention of antigen-targeting properties. Affinity measurements for all identified anti-trypanosomal Nbs reveal a strong correlation between trypanotoxicity and affinity (K(D)), suggesting that it is a crucial determinant for this activity. Half maximal effective (50%) affinity of 57 nM was calculated from the non-linear dose-response curves. In line with these observations, Nb humanizing mutations only preserved the trypanotoxic activity if the K(D) remained unaffected.CONCLUSIONS/SIGNIFICANCE: This study reveals that the binding properties of Nanobodies need to be compatible with achieving an occupancy of >95% saturation of the parasite surface VSG in order to exert an anti-trypanosomal activity. As such, Nb-based approaches directed against the VSG target would require binding to an accessible, conserved epitope with high affinity.

DOI10.1371/journal.pntd.0001902
Alternate JournalPLoS Negl Trop Dis
PubMed ID23166849
PubMed Central IDPMC3499403
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