Modification of the zonal elution method for detection of transient protein-protein interactions involving ligand exchange.

ABSTRACT: A new affinity chromatography method was developed by modifying a zonal elution method. The new method targets transient protein-protein interactions, such as those encountered during direct ligand transfer between the ligand transporter and its cognate receptor. A ligand-loaded transport protein is immobilized on the solid support, and a plug containing a putative receptor is flowed through the column. Elution profiles of proteins not interacting with the immobilized transporter can be approximated with a simple Gaussian curve, while the elution profiles of cognate receptors show significant delay and exhibit complex shape. Ligand transfer from the immobilized transporter molecules to the receptors is verified by both UV absorbance measurements and mass spectrometry. The sensitivity of th

SUBMITTER: Sjoelund V 

PROVIDER: S-EPMC3352988 | biostudies-literature | 2012 May

REPOSITORIES: biostudies-literature