Structural basis of G-quadruplex recognition by a camelid antibody fragment

Apart from the iconic Watson-Crick duplex, DNA can fold into different noncanonical structures, of which the most studied are G-quadruplexes (G4s). Despite mounting structural and biophysical evidence, their existence in cells was controversial until their detection using G4-specific antibodies. However, it remains unknown how antibodies recognize G4s at the molecular level and why G4-specific antibodies have low selectivity and are unable to distinguish different G4 sequences. Here, we present the crystal structure of a nanobody bound to the archetypical G4 structure, the thrombin-binding aptamer (TBA). The nanobody exhibits strong selectivity against different G4 sequences and utilizes an unusual scaffold-based paratope, with very limited involvement of complementarity-determining region. The nanobody effectively mimics the binding interface of thrombin, a natural binding partner of TBA, by using isosteric interactions at key positions. The presented structure sheds light on the molecular basis of how antibodies, essential G4-detection tools, recognize noncanonical G4 structures.