Parasitic nematodes are responsible for numerous chronically incapacitating and deforming diseases in Africa, Asia, and the Americas. Among these diseases is lymphatic filariasis, which is a mosquito-transmitted disease, endemic to 81 countries. It is estimated that 120 million people are infected with this disease. Enzymes that are essential for the parasitic nematodes but that do not have a human homologue are potential drug targets for therapeutic intervention. The availability of the genome from B. malayi, the representative organism for filarial nematodes, has enabled the ranking of potential drug targets from this parasitic organism. One such enzyme is trehalose-6-phosphate phosphatase (T6PP), which is required for the biosynthesis of trehalose. The oblation of T6PP activity in the C. elegans model commonly used for parasitic nematodes ultimately leads to organism death, probably due to the accumulation of trehalose 6-phosphate (T6P). Because T6PP is a member of the haloalkanoate dehalogenase superfamily of phosphatases, knowledge about the structure/function relationships in this family can be used to define T6PP for drug development. The objective of the proposed study is to identify the steric and electrostatic features of the T6PP active-site region that can be exploited in the design of lead inhibitors. Because the two domains of the enzyme can rotate to open the active site for ligand exchange, the surface area that can be potentially targeted by an inhibitor will be obtained by determining structures of bot open and closed conformers. The research plan is focused on a single Aim: Define the Target Site for the Development of Drug-like Inhibitors of B. malayi Trehalose-6-phosphate Phosphatase. The ?lead? T6PP X-ray crystal structure will be determined using the recombinant enzyme from B. malayi and C. elegans to define the overall structure of the protein. Co-crystallization and crystal soaking experiments will capture the closed state (using an inactive T6PP mutant and T6P or using T6PP plus an inert substrate analog) and transition-state conformations (T6PP and trehalose plus vanadate, beryllium fluoride, or aluminum fluoride). Analysis of structure-activity relationships will be used to define the binding interactions which dominate the contributions to the ligand binding energy. The structure of the enzyme in the cap-open conformation will be determined (using apo T6PP and T6PP domain-domain binding mutants) in order to design and evaluate bidentate inhibitors that can fill and complement the expanded binding crevice of the cap-open conformer. This work will deliver the foundation for the development of a drug for the treatment of disease(s) inflicting the large segment of the world's population suffering from infection by parasitic nematodes.
It is estimated that 120 million people are infected with lymphatic filariasis, one of the debilitating diseases caused by parasitic nematodes. The proposed work will develop trehalose 6-phosphate phosphatase (T6PP), an enzyme essential to parasitic nematodes, as a drug target for therapeutic intervention. The findings will identify the steric and electrostatic features of T6PP that can be exploited in the design of lead inhibitors and deliver the foundation for drug discovery.
| Farelli, Jeremiah D; Galvin, Brendan D; Li, Zhiru et al. (2014) Structure of the trehalose-6-phosphate phosphatase from Brugia malayi reveals key design principles for anthelmintic drugs. PLoS Pathog 10:e1004245 |
