UC San Diego

Carbonic anhydrase was used as a model system to examine the influence of metalloenzyme active site environments on coordination by metal-binding pharmacophores (MBPs). This dissertation will first discuss metalloenzymes as therapeutic targets, along with inhibitors of these enzymes that have gained FDA approval in Chapter 1. The characteristics of MBPs that have generally been implicated in the low clinical success rate of therapeutics targeting these enzymes are then discussed, followed by an explanation of how the use of a more diverse collection of MBPs can potentially solve these problems. The coordination of a series of novel MBPs inspired by Zn²⁺ sensors to Co²⁺, Ni²⁺, Cu²⁺, and Zn²⁺ will then be discussed in Chapter 2. These studies into the fundamental coordination chemistry of MBPs provide valuable information into the preferred binding modes of these molecules to active site metal ions. Inhibition of human carbonic anhydrase II (hCAII) by MBPs was investigated and those molecules showing activity were cocrystallized in the hCAII active site. The binding modes were then compared to those observed in small-molecule model complexes. Unexpected monodentate coordination by the O,S-donor ligand 3-hydroxy-2-methylpyran-4-thione is described in Chapter 3. Further studies using substituted derivatives of the 1-hydroxy-1H-pyridin-2-thione MBP are then discussed that show the monodentate coordination mode to be a result of interactions with the surrounding hCAII active site environment formed upon metal coordination. In Chapter 4, mutants of hCAII were then used to examine the influence of the metal coordination sphere on MBP binding. The affinity and binding mode of benzenesulfonamide, a prototypical hCAII inhibitor, to mutants containing altered coordination spheres will be discussed that show decreased binding affinity despite a common binding mode. Finally, in Chapter 5, a class of inhibitors that bind in the hCAII active site by hydrogen bonding to the metal- bound water molecule rather than direct coordination of the active site metal ion, are discussed