Design and synthesis of discontinuous protein binding site mimics of HIV gp120

Essentially all cellular processes are mediated by protein-protein interactions and detailed knowledge about these interactions can aid in determining biological functions or provide opportunities for the treatment of human disease. Extensive research has been performed on the development of modulators of protein-protein interactions, since these molecules could provide better understanding of the mechanisms of molecular recognition, lead to the identification of drug targets or provide access to novel therapeutics. Peptide-based mimics of protein binding sites are promising candidates for this purpose. Despite the progress that has been made in the field of protein mimicry and the increasing understanding of protein-protein interactions, the development of molecules that can modulate these interactions remains a challenging task because of the complex nature of protein binding sites. Protein-protein interaction sites often consist of discontinuous epitopes. An important example of such an interaction is the CD4 binding site of HIV gp120. Since gp120 plays a central role in the ability of HIV to enter cells, it is an interesting target for vaccine research. Therefore, in this thesis the development of a fast and reliable method for the synthesis of collections of discontinuous epitope mimics is described. The method comprises the conjugation of three different cyclic peptides, representing the epitopes, to a scaffold molecule, using the copper(I)-catalyzed alkyne-azide cycloaddition reaction. The main characteristic of our method is the rapid generation in a single experiment of a mixture of multiple discontinuous epitope constructs. Thus, a diversity of combinations of cyclic peptides on a scaffold molecule is accessible in a reproducible manner. These different constructs can be conveniently separated by preparative HPLC and characterized with mass spectrometry. The mixture and the individual constructs were tested for CD4-binding affinity with ELISA for evaluation of their ability to act as gp120 mimics. This synthetic procedure provides rapid access to a diversity sophisticated peptide biomolecular constructs. Moreover, the synthesis and activity screening of multiple products in one approach enhances the probability to rapidly generate a starting point for developing synthetic vaccines derived from gp120 or neutralizing antibodies targeting CD4