Evaluation of the anti-GFP tandem nano body LaG-16- G4S-LaG2

Nanobodies are an unconventional type of antibodies. Also known as heavy-chain antibodies, they are found in the sera of Camelidae and are devoid of the L-chain polypeptide, while also lacking the first constant domain - CH1. These properties, as well as their relatively smaller size allow them to be well adapted for accessing binding sites of difficult targets. A prime example of a versatile nanobody is the tandem nanobody LaG-16-G4S-LaG-2, developed by the Rout lab, as this group has shown its high specificity and affinity for GFP. The strong affinity of the tandem nanobody for this marker molecule would allow its utilization in the purification of low-abundance proteins or protein complexes. Here I present experiments aimed at rendering the LaG-16-G4S-LaG-2-GFP protein capture system and its subsequent combination with the FimGt-DsF binding complex accessible for future structural studies. Furthermore, I show how the system can be improved upon in the future. I am certain thatthe experiments at hand will help to illuminate the crucial steps of the novel protein purification approach and can lead t