Technological watch

AbstractPhage display and biopanning are powerful tools for generating binding molecules for a specific target. However, the selection process based only on binding affinity provides no assurance for the antibody's affinity to the target epitope. In this study, we propose a molecular?evolution approach guided by native protein–protein interactions to generate epitope?targeting antibodies. The binding?site sequence in a native protein was grafted into a complementarity?determining region (CDR) in the nanobody, and a nonrelated CDR loop (in the grafted nanobody) was randomized to create a phage display library. In this construction of nanobodies by integrating graft and evolution technology (CAnIGET method), suitable grafting of the functional sequence added functionality to the nanobody, and the molecular?evolution approach enhanced the binding function to inhibit the native protein–protein interactions. To apply for biological tool with growth screening, model nanobodies with an affinity for filamenting temperature?sensitive mutant Z (FtsZ) from Staphylococcus aureus were constructed and completely inhibited the polymerization of FtsZ as a function. Consequently, the expression of these nanobodies drastically decreased the cell division rate. We demonstrate the potential of the CAnIGET method with the use of native protein–protein interactions for steady epitope?specific evolutionary engineering.This article is protected by copyright. All rights reserved