Souders, C. | Search bioRxiv

bioRxiv2025

The Therapeutic Nanobody Profiler: characterising and predicting nanobody developability to improve therapeutic design

Developability optimisation is an important step for successful biotherapeutic design. For monoclonal antibodies, developability is relatively well characterised. However, progress for novel biotherapeutics such as nanobodies is more limited. Differences in structural features between antibodies and nanobodies render current antibody computational methods unsuitable for direct application to nanobodies. Following the principles of the Therapeutic Antibody Profiler (TAP), we have built the Therapeutic Nanobody Profiler (TNP), an open-source computational tool for predicting nanobody developability. Tailored specifically for nanobodies, it accounts for their unique properties compared to conventional antibodies for more efficient development of this novel therapeutic format. We calibrate TNP metrics using the 36 currently available clinical-stage nanobody sequences. We also collected experimental developability data for 108 nanobodies and examine how these results are related to the TNP guidelines. TNP is available as a web application at opig.stats.ox.ac.uk/webapps/tnp.

bioinformatics↗

bioRxiv2020

Conformational Tuning of Amylin by Charged SMA Copolymers

Human amylin forms structurally heterogeneous amyloids that have been linked to type-2 diabetes. Thus, understanding the molecular interactions governing amylin aggregation can provide mechanistic insights in its pathogenic formation. Here, we demonstrate that fibril formation of amylin is altered by synthetic amphipathic copolymer derivatives of the styrene- maleic-acid (SMAQA and SMAEA). High-speed AFM is used to follow the real-time aggregation of amylin by observing the rapid formation of de novo globular oligomers and arrestment of fibrillation by the positively-charged SMAQA. We also observed an accelerated fibril formation in the presence of the negatively-charged SMAEA. These findings were further validated by fluorescence, SOFAST-HMQC, DOSY and STD NMR experiments. Conformational analysis by CD and FT-IR revealed that the SMA copolymers modulate the conformation of amylin aggregates. While the species formed with SMAQA are -helical, the ones formed with SMAEA are rich in {beta}-sheet structure. The interacting interfaces between SMAEA or SMAQA and amylin are mapped by NMR and microseconds all-atom MD simulation. SMAEA displayed {pi}-{pi} interaction with Phe23, electrostatic {pi}-cation interaction with His18 and hydrophobic packing with Ala13 and Val17; whereas SMAQA showed a selective interaction with amylins C-terminus (residues 31-37) that belongs to one of the two {beta}-sheet regions (residues 14-19 and 31-36) involved in amylin fibrillation. Toxicity analysis showed both SMA copolymers to be non-toxic in vitro and the amylin species formed with the copolymers showed minimal deformity to zebrafish embryos. Together, this study demonstrates that chemical tools, such as copolymers, can be used to modulate amylin aggregation, alter the conformation of species.

biophysics↗