Hints

The Histidine Triad Nucleotide binding proteins (Hints) are have been the subject of considerable structural and biophysical study in our laboratory. While the cellular role of human Hints is still mysterious, cytosolic human Hint1 is important because its unique phosphoramidase catalytic activity is required for the metabolic activation of nucleoside phosphoramidate prodrugs. All our work on Hints is completed in collaboration with the Wagner lab, which was the first to study the substrate selectivity of Hint enzymes.

Our lab determined the first structure of the mitochondrial human Hint2[1], and has continued to try to understand the molecular basis for Hint enzyme substrate selectivity and reactivity by examining the structures of complexes with substrate phosphoramidates and nucleotide products. Complex structures are used to rationalize the observed thermodynamics of binding and catalytic efficiency[2]. With a substrate chemically engineered to react slowly, we were able to trap the adenylylated histidine reaction intermediate (E* in Figure at left) using time-resolved crystallography as catalysis occurs in crystals of hHint1[3].

The Wagner group continues to investigate the role played by Hints in the development of insensitivity to opioids and neuropathic pain, and it is likely that future work with Hint enzymes will be directed toward the discovery and optimization of potent and selective Hint inhibitors[4].