Nanobody aided purification and crystallization of G-protein coupled receptors.

Project Details

Description

G-protein coupled receptors (GPCRs) are the largest, most versatile group of membrane proteins with proven therapeutic value. So far it has been very difficult to recombinantly express, purify, characterize and crystallize these receptors for structural analysis. Therefore little is known. about their 3D structure. Within this project I will exploit the unique properties of single domain antibodies or Nanobodies to overcome these bottlenecks in structural analysis of GPCRs. Nanobodies (Nbs) are the smallest antigen binding fragments of naturally occurring heavy chain antibodies found in camelids. I will generate specific Nbs with nanomolar affinity against a small number of chosen target GPCRs. To determine if the Nbs recognize a biological relevant conformation, these GPCR specific Nbs will be characterized as agonist or antagonists. I will also test if the co-expression of the Nbs together with their target GPCRs enhances the yield of functional receptors for structural analysis. Next the characterized Nbs will be used to produce antibody based affinity columns to trap solubilized but correctly folded GPCRs. In the last step the Nbs will be tested for their ability as crystallization chaperones. The final goal of this project is to solve high resolution three-dimensional structures of one of the target GPCRs.
AcronymFWOTM557
StatusFinished
Effective start/end date1/10/1030/09/14

Keywords

  • Applied Biology

Flemish discipline codes

  • Biological sciences