Baculovirus Surface Display

The importance and variety of applications of surface display libraries as a tool in basic research as well as for the development of diagnostic and clinical drugs has become evident during the past years. Bacterial "phage-display- libraries" can be used to isolate specific ligands out of pools of different binding, proteins, which are being expressed on the viral surface, as fusion proteins to a phage coat protein. Specific binders can be screened and identified very easily by affinity selection ("panning"). However, there are limitations to this approach when it comes to complex, eukaryotic proteins, that are not processed properly in the bacterial expression system (glycosylation, folding, assembly). An attractive, eukaryotic expression system, is the insect-cell-baculovirus expression system. In the course of our work we compared different viral constructs in terms of expression of foreign proteins on the surface of baculovirus particles as well as on infected insect cells. In the course of this project we are planing to further optimize the cloning strategy and increase the relative portion of the foreign protein on the baculovirus surface. Combining optimal -viral constructs with a refined sorting protocol should result in the establishment of an easy to handle expression and selection system. The ability of creating large libraries of complex eukaryotic proteins on the surface of reproducing particles will further be tested. For this purpose we will generate first an epitope library (representing the entire envelope protein gpl60 of HIV-1) and second a subtractive cDNA library. After determining size- and diversity of constructed libraries, cell sorting by a fluorescent activated cell sorter. (FACS) will be used for selection of individual viral clones. Successful screening of these libraries would confirm the usefulness and the flexibility of our system. By optimizing, the baculovirus surface expression system we are attempting a major contribution to ligand technology in general.