Abstract
Tryptophan hydroxylase (TPH) is a tetrahydrobiopterin (BH4)- and iron-dependent enzyme that hydroxylates L-Trp to 5-hydroxy-L-Trp using molecular oxygen. This is the rate limiting step in the synthesis of serotonin. Due to the scarcity of the enzyme in animal tissues and its instability in vitro, TPH is the least characterized of the three aromatic amino acid hydroxylases and its 3D structure is still not known. Based on the high sequence identity between the three mammalian hydroxylases we have prepared a structural model for TPH (1) (Fig. 1A). We have also determined the structure of the bound conformation of L-Trp and the inactive cofactor analogue 7,8-dihydrobiopterin (BH2) complexed with a stable form of the catalytic domain of human TPH (Δ90TPH) by NMR and by molecular docking (McKinney et al., submitted; Fig. 1B). From the structure of the complex it was inferred that residue F313 (W326 in phenylalanine hydroxylase (PAH)) may have a role in substrate specificity. In this work we report the kinetic characterization of Δ90TPH and of the mutants F313/W326.
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References
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McKinney, J., Teigen, K., Frøystein, N.Å., Knappskog, P.M., Haavik, J., Martínez, A. (2002). Role of PHE313/TRP326 in Determining Substrate Specificity in Tryptophan and Phenylalanine Hydroxylases. In: Milstien, S., Kapatos, G., Levine, R.A., Shane, B. (eds) Chemistry and Biology of Pteridines and Folates. Springer, Boston, MA. https://doi.org/10.1007/978-1-4615-0945-5_16
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DOI: https://doi.org/10.1007/978-1-4615-0945-5_16
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