Virtually all organisms utilize adenosine triphosphate (ATP) for substrate level phosphorylations during glycolysis; however, it has been recognized that the parasitic protists lacking mitochondria have glycolytic enzymes which utilize inorganic pyrophosphate (PPi) as the high energy phosphate donor instead of ATP. PPi-phosphofructokinase (PPi-PFK) is the rate limiting glycolytic enzyme found in Entamoeba histolytica, Giardia lamblia, Toxoplasma gondii, Trichomonas vaginalis, and Naegleria fowleri, and as such represents an important drug target. Current therapy for these infections is not ideal, especially for pregnant women and children, and newer, selective agents would be welcome. The primary amino acid sequences of PPi-PFK from E. histolytica, G. lamblia and N. fowleri have been compared to gain insight into how these sequences differ from those of known ATP-PFKs, including human. Although the primary amino acid sequences of the known PPi-PFKs and ATP-PFKs share little overall identity, there are areas of strong homology (60-80%) in several regions representing the known active sites of these enzymes. Because of this, structure based molecular modeling has the potential to make useful predictions about the structure of the PPi-PFK pyrophosphate binding site, and potential drug-target interactions at the site. From the crystallographic structure of B. stearothermophilus ATP-PFK, a stereochemically reasonable model for E. histolytica PPi-PFK has been constructed using Look. This model has been validated by several computational strategies including comparison with a backbone=dependent rotamer library for analysis of side chain orientation, The model has been used with single mode DOCK to predict the binding site of pyrophosphate and to evaluate the interaction of known pyrophosphate analogs at this site. Two Ciba-Geigy compounds were found to have good drug-target interaction based on the DOCK strategy. These compounds were then tested in vivo against E. histolytica and they significantly inhibited the growth of the amoebae. The model will also be used to screen the Available Chemicals Directory for potential inhibitors of pyrophosphate binding, using Dock 3.5. The most promising candidates will be tested in-vivo. These modeling studies have the potential to identify clinically useful broad spectrum antiparasitic agents, and would not be possible without the use of the Computer Graphics Lab.