| The examples of CsA, FKpYEEI, and SLFpYEEI have shown that the activity of a small-molecule ligand can be altered by a non-target protein. For CsA, recruitment of the protein cyclophilin makes it an effective inhibitor of calcineurin's phosphatase activity. In the case of FKpYEEI, recruitment of the protein FKBP52 makes it a more effective binder to FynSH2. By contrast, the involvement of FKBP12 makes SLFpYEEI a less effective binder to FynSH2. Several questions are raised by these examples. First, what can we know about the trimeric complexes that are formed? The dissociation constant for either bifunctional molecule·protein complex can be measured by an array of standard methods (e.g., competition binding, fluorescence quenching, etc.), but the binding of a bifunctional molecule·protein dimeric complex to the remaining protein is more difficult to determine. The second chapter of this thesis addresses solutions to that problem and analyzes the activity of the bifunctional molecule SLFpYEEI. Can the presence of a presenter protein control the biological activity of a bifunctional molecule for its target protein by decreasing affinity? How do protein concentration and ligand dissociation constants affect the ability of the presenter protein to control bifunctional molecule activity? These questions will be addressed by the bifunctional molecules PMPSLF and MTXSLF. The third chapter explores the extreme condition of protein-protein interactions so energetically unfavorable that no trimeric complex can form. Finally, what is the maximum amount of ligand enhancement that can be achieved? Are discrete, monomeric proteins the only targets and presenters that are available? In the fourth chapter the ability of daunorubicin-taxol and daunorubicin-colchine dimers will be tested for the ability to simultaneously bind to the biopolymers DNA and microtubules in an attempt to create polyvalent arrays. |
