Final Report Summary - EXIVC (Expanding the Potential of in vitro Compartmentalised Screening and Selection Approaches)
Western blotting showed that antibody binding is not compromised under these conditions.
Building in this procedure I wanted to conduct selections with carbohydrate binding domains (CBDs) and started to clone a selection of these proteins. A number of these proteins did not show good expression in in vitro expression systems, neither in PURE, nor in commercial Roche or Invitrogen kits. Towards the end of the project I have established that visible enrichment can be observed in model selections with one CBD. Proper selections could not be performed in the timeframe of the project, because of the many practical obstacles encountered. However, other members of the host group will take up this project in the future. The – by now robust – protocol to perform selections from protein libraries with disulfide bonds will help to achieve the goal of actual selection of CBDs. I have set up assays for catalytic selections that will allow screening of a catalytically competent protein after evolving the binding capabilities of the CBD.