Structure–Function Relationships in the Aromatic Amino Acid Hydroxylases Enzyme Family: Evolutionary Insights

Abstract

The nonheme iron and tetrahydrobiopterin dependent aromatic amino acid hydroxylases (AAAHs) comprise a family of enzymes that catalyse the hydroxylation of l‐Phe, l‐Tyr and l‐Trp. These reactions are of central physiological importance. Consequently, dysfunction of the AAAHs is associated with serious disorders, that is, the genetic metabolic disease phenylketonuria for phenylalanine hydroxylase (PAH), and neurological and neuropsychiatric disorders for tyrosine hydroxylase (TH) and tryptophan hydroxylase 1 and 2 (TPH1, TPH2). Mammalian AAAHs are tetrameric proteins that present a three‐domain structure with a remarkable high similarity, in particularly for the catalytic domains. Structural analyses have provided valuable insights on the effect of disease‐associated mutations, mechanism of catalysis, the determinants for substrate specificity and regulation of enzymatic activity. The best characterised AAAH is PAH, which also seems to be the precursor of the family. The AAAHs have developed sophisticated regulatory mechanisms during evolution to control substrate l‐Phe (PAH), catecholamine (TH), serotonin and melatonin (TPHs) levels.

Key Concepts:

  • The aromatic amino acid hydroxylases (AAAHs) are tetrahydrobiopterin (BH4) dependent enzymes that use dioxygen as additional substrate.

  • The AAAHs are involved in serious inborn errors of metabolism with neuronal impact.

  • The AAAHs are highly homologous, contain a catalytic mononuclear nonheme iron coordinated by a 2‐His‐1‐carboxylate facial triad and show similar reaction mechanism.

  • Metazoans have at least three AAAH genes: phenylalanine hydroxylase (PAH), tyrosine hydroxylase (TH) and tryptophan hydroxylase(s) (TPH(s)), where PAH appears as the precursor of the family.

  • Mammalian PAH, TH, TPH1 and TPH2 are tetrameric, with a three‐domain subunit structure; substrate specificity is provided by the catalytic domain.

  • The AAAHs are highly regulated by mechanisms at the transcriptional, translational, posttranslational and allosteric levels.

  • Evolutionary adaptation of the enzymes is most evident at the regulatory mechanisms, along the complexity of the organisms, especially of the central nervous system (CNS).

Keywords: tetrahydrobiopterin; mononuclear nonheme iron; 2‐His‐1‐carboxylate facial triad; enzyme regulation; structure–function relationships; inborn errors of metabolism; neurometabolic disorders

Figure 1.

The reactions catalysed by mammalian AAAHs and the synthesis and regeneration of BH4. The BH4‐dependent reaction of the AAAHs leads to the hydroxylation of the respective aromatic amino acid (grey panel) and the conversion of BH4 into BH4–4α‐carbinolamine. The two enzymes pterin 4α‐carbinolamine dehydratase and dihydropteridine reductase (DHPR) are responsible for the regeneration of the cofactor, necessary for a new cycle of hydroxylation (light blue panel). In the absence of DHPR, quinoid dihydrobiopterin can be converted non‐enzymatically to BH2 and reduced to BH4 by dihydrofolate reductase (DHFR) through a salvage pathway (dashed arrows in light blue panel). BH4 is synthesised de novo from guanosine tripshophate (GTP) by the action of three enzymes; GTP cyclohydrolase I, pyruvoyltetrahydropterin synthase and SR (yellow panel). There are a number of alternate enzymes and salvage pathways that also lead to the synthesis of BH4 (dashed arrows in yellow panel).

Figure 2.

Sequence and structure of the AAAHs. (a) Schematic representation of the sequence alignment of human PAH, TH (isoform 1), TPH1 and TPH2. Residues conserved in all four proteins are depicted with a red bar above the alignment scheme. Phosphorylation sites are marked with a yellow star and correspond to positions Ser16 for PAH; Thr8, Ser19, Ser31, Ser40 for TH1; Ser58 for TPH1 and Ser19 for TPH2. (b–d) Structure of the AAAHs subunits coloured yellow (regulatory domain), blue (catalytic domain) and red (oligomerization domain). The cofactor BH4 and the amino acid substrates are shown in orange and yellow spheres, respectively. The solution structure of the regulatory ACT domain of TH is shown in white cartoon (PDB id 2MDA). (e–f) Tetrameric assemblies of PAH and TH generated by combining the quaternary structures of the oligomerization and catalytic domains (PDB id 2PAH for PAH; 1TOH for TH) with the structure of (e) the regulatory and catalytic domains (PDB id 1PHZ for PAH) or (f) the solution structure of dimeric regulatory domain (PDB id 2MDA for TH) in accordance with (Zhang et al., ). Only subunits I and IV are coloured for visualisation purposes. (g) Hypothetical tetrameric model of TPH1. For this enzyme only structural data of the truncated catalytic domain is so far available (shown here: PDB id 3E2T), and the tetramer is, therefore, blurred.

Figure 3.

Binding interactions of the AAAHs. (a) Cofactor binding site of PAH (PDB id 1J8U) with the cofactor BH4 in ball and stick representation (orange) showing stacking interactions with Phe254 and electrostatic interactions with Glu286, which are conserved in all four AAAHs. (b) Substrate binding site of TPH1 (PDB id 3E2T) with bound amino acid substrate (l‐Trp; in yellow) showing the interaction with conserved Arg257. (c) l‐DOPA bound to PAH (PDB id 6PAH); the hydroxyl groups of the catechol replace two iron‐bound water molecules. (d) Ser19‐phosphorylated N‐terminal peptide of TH bound to 14‐3‐3γ (PDB id 4J6S).

Figure 4.

Substrate‐dependent conformational changes in the catalytic domain. (a) Structural comparison of TPH1 with cofactor (PDB id 1MLW) and with substrate (PDB id 3E2T). Cofactor‐ and substrate‐bound structures are shown in white and grey cartoon, respectively. Loops consisting of residues Val122‐Phe136, Gln362 and Asp371 (in TPH1) closing around the active site upon substrate binding are highlighted in red (without substrate) and yellow (with substrate). (b–c) Principal component analysis of the available X‐ray structures of the AAAHs. (b) Visualisation of principal component 1 which consists of a subdomain (coloured red) moving in over the substrate binding site relative to a more rigid core. (c) Available X‐ray structures projected onto the two first principal components with each dot representing one structure of the catalytic domain. The subunits can be divided into two major groups based on their pairwise structural deviations: (i) substrate bound, and (ii) apo and pterin bound, with a group of inhibitor bound structures of TPH1 obtaining an intermediate state.

Figure 5.

AAAHs phylogenetic analysis of representative organisms. Nodes coloured red and blue depict PAH from bacteria and protozoa, respectively. Sequences marked with an asterisk represent unreviewed sequences in the Uniprot database http://www.uniprot.org/.

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References

Andersen OA, Flatmark T and Hough E (2001) High resolution crystal structures of the catalytic domain of human phenylalanine hydroxylase in its catalytically active Fe(II) form and binary complex with tetrahydrobiopterin. Journal of Molecular Biology 314: 279–291.

Aravind L and Koonin EV (1999) Gleaning non‐trivial structural, functional and evolutionary information about proteins by iterative database searches. Journal of Molecular Biology 287: 1023–1040.

Blau N, van Spronsen FJ and Levy HL (2010) Phenylketonuria. Lancet 376: 1417–1427.

Calvo AC, Pey AL, Miranda‐Vizuete A, Doskeland AP and Martinez A (2011) Divergence in enzyme regulation between Caenorhabditis elegans and human tyrosine hydroxylase, the key enzyme in the synthesis of dopamine. Biochemical Journal 434: 133–141.

Daubner SC, Le T and Wang S (2011) Tyrosine hydroxylase and regulation of dopamine synthesis. Archives of Biochemistry and Biophysics 508: 1–12.

Daubner SC, Melendez J and Fitzpatrick PF (2000) Reversing the substrate specificities of phenylalanine and tyrosine hydroxylase: aspartate 425 of tyrosine hydroxylase is essential for l‐DOPA formation. Biochemistry 39: 9652–9661.

Dunkley PR, Bobrovskaya L, Graham ME et al. (2004) Tyrosine hydroxylase phosphorylation: regulation and consequences. Journal of Neurochemistry 91: 1025–1043.

Faust DM, Catherin AM, Barbaux S et al. (1996) The activity of the highly inducible mouse phenylalanine hydroxylase gene promoter is dependent upon a tissue‐specific, hormone‐inducible enhancer. Molecular Cell Biology 16: 3125–3137.

Fitzpatrick PF (1999) Tetrahydropterin‐dependent amino acid hydroxylases. Annual Review of Biochemistry 68: 355–381.

Fitzpatrick PF (2012) Allosteric regulation of phenylalanine hydroxylase. Archives of Biochemistry and Biophysics 519: 194–201.

Flydal MI and Martinez A (2013) Phenylalanine hydroxylase: function, structure, and regulation. IUBMB Life 65: 341–349.

Fossbakk A, Kleppe R, Knappskog PM, Martinez A and Haavik J (2014) Functional studies of tyrosine hydroxylase missense variants reveal distinct patterns of molecular defects in DOPA responsive dystonia. Human Mutation 35: 880–890.

Fusetti F, Erlandsen H, Flatmark T and Stevens RC (1998) Structure of tetrameric human phenylalanine hydroxylase and its implications for phenylketonuria. Journal of Biological Chemistry 273: 16962–16967.

Fölling A (1934) Über ausscheidung von phenylbrenztraubensäure in den harn als stoffwechselanomalie in verbindung mit imbezillitat. Hoppe‐Seyler's Zeitschrift für Physiologische Chemie 227: 169–176.

Gardino AK and Yaffe MB (2011) 14‐3‐3 Proteins as signaling integration points for cell cycle control and apoptosis. Seminars in Cell and Developmental Biology 22: 688–695.

Goodwill KE, Sabatier C, Marks C et al. (1997) Crystal structure of tyrosine hydroxylase at 2.3 A and its implications for inherited neurodegenerative diseases. Nature Structural & Molecular Biology 4: 578–585.

Gordon SL, Bobrovskaya L, Dunkley PR and Dickson PW (2009) Differential regulation of human tyrosine hydroxylase isoforms 1 and 2 in situ: isoform 2 is not phosphorylated at Ser35. Biochimica et Biophysica Acta 1793: 1860–1867.

Grenett HE, Ledley FD, Reed LL and Woo SL (1987) Full‐length cDNA for rabbit tryptophan hydroxylase: functional domains and evolution of aromatic amino acid hydroxylases. Proceedings of the National Academy of Sciences of the USA 84: 5530–5534.

Grohmann M, Hammer P, Walther M et al. (2010) Alternative splicing and extensive RNA editing of human TPH2 transcripts. PLoS One 5: e8956.

Haavik J, Blau N and Thony B (2008) Mutations in human monoamine‐related neurotransmitter pathway genes. Human Mutation 29: 891–902.

Iida Y, Sawabe K, Kojima M et al. (2002) Proteasome‐driven turnover of tryptophan hydroxylase is triggered by phosphorylation in RBL2H3 cells, a serotonin producing mast cell line. European Journal of Biochemistry 269: 4780–4788.

Jaffe EK, Stith L, Lawrence SH, Andrake M and Dunbrack RL Jr (2013) A new model for allosteric regulation of phenylalanine hydroxylase: implications for disease and therapeutics. Archives of Biochemistry and Biophysics 530: 73–82.

Kappock TJ and Caradonna JP (1996) Pterin‐dependent amino acid hydroxylases. Chemical Reviews 96: 2659–2756.

Kaufman S (1993) The phenylalanine hydroxylating system. Advances in Enzymology and Related Areas of Molecular Biology 67: 77–264.

Kawahata I, Tokuoka H, Parvez H and Ichinose H (2009) Accumulation of phosphorylated tyrosine hydroxylase into insoluble protein aggregates by inhibition of an ubiquitin‐proteasome system in PC12D cells. Journal of Neural Transmission 116: 1571–1578.

Kobe B, Jennings IG, House CM et al. (1999) Structural basis of autoregulation of phenylalanine hydroxylase. Nature Structural & Molecular Biology 6: 442–448.

Kumer SC, Mockus SM, Rucker PJ and Vrana KE (1997) Amino‐terminal analysis of tryptophan hydroxylase: protein kinase phosphorylation occurs at serine‐58. Journal of Neurochemistry 69: 1738–1745.

Kumer SC and Vrana KE (1996) Intricate regulation of tyrosine hydroxylase activity and gene expression. Journal of Neurochemistry 67: 443–462.

Lundin LG (1999) Gene duplications in early metazoan evolution. Seminars in Cell and Developmental Biology 10: 523–530.

Lye LF, Kang SO, Nosanchuk JD, Casadevall A and Beverley SM (2011) Phenylalanine hydroxylase (PAH) from the lower eukaryote Leishmania major. Molecular and Biochemical Parasitology 175: 58–67.

Martinez A, Calvo AC, Teigen K and Pey AL (2008) Rescuing proteins of low kinetic stability by chaperones and natural ligands phenylketonuria, a case study. Progress in Molecular Biology and Translational Science 83: 89–134.

McKinney J, Teigen K, Froystein NA et al. (2001) Conformation of the substrate and pterin cofactor bound to human tryptophan hydroxylase. Important role of Phe313 in substrate specificity. Biochemistry 40: 15591–15601.

Murphy KL, Zhang X, Gainetdinov RR, Beaulieu JM and Caron MG (2008) A regulatory domain in the N terminus of tryptophan hydroxylase 2 controls enzyme expression. Journal of Biological Chemistry 283: 13216–13224.

Nagatsu T (1995) Tyrosine hydroxylase: human isoforms, structure and regulation in physiology and pathology. Essays in Biochemistry 30: 15–35.

Nagatsu T and Ichinose H (1999) Regulation of pteridine‐requiring enzymes by the cofactor tetrahydrobiopterin. Molecular Neurobiology 19: 79–96.

Nakamura K and Hasegawa H (2007) Developmental role of tryptophan hydroxylase in the nervous system. Molecular Neurobiology 35: 45–54.

Olsson E, Teigen K, Martinez A and Jensen VR (2010) The aromatic amino acid hydroxylase mechanism: a perspective from computational chemistry. Advances in Inorganic Chemistry 62: 437–500.

Pey AL, Stricher F, Serrano L and Martinez A (2007) Predicted effects of missense mutations on native‐state stability account for phenotypic outcome in phenylketonuria, a paradigm of misfolding diseases. American Journal of Human Genetics 81: 1006–1024.

Pey AL, Ying M, Cremades N et al. (2008) Identification of pharmacological chaperones as potential therapeutic agents to treat phenylketonuria. Journal of Clinical Investigation 118: 2858–2867.

Roberts KM and Fitzpatrick PF (2013) Mechanisms of tryptophan and tyrosine hydroxylase. IUBMB Life 65: 350–357.

Sarkissian CN, Gamez A, Wang L et al. (2008) Preclinical evaluation of multiple species of PEGylated recombinant phenylalanine ammonia lyase for the treatment of phenylketonuria. Proceedings of the National Academy of Sciences of the USA 105: 20894–20899.

Scriver CR and Waters PJ (1999) Monogenic traits are not simple: lessons from phenylketonuria. Trends in Genetics 15: 267–272.

Siltberg‐Liberles J, Steen IH, Svebak RM and Martinez A (2008) The phylogeny of the aromatic amino acid hydroxylases revisited by characterizing phenylalanine hydroxylase from Dictyostelium discoideum. Gene 427: 86–92.

Teigen K, McKinney JA, Haavik J and Martinez A (2007) Selectivity and affinity determinants for ligand binding to the aromatic amino acid hydroxylases. Current Medicinal Chemistry 14: 455–467.

Walther DJ and Bader M (2003) A unique central tryptophan hydroxylase isoform. Biochemical Pharmacology 66: 1673–1680.

Werner ER, Blau N and Thony B (2011) Tetrahydrobiopterin: biochemistry and pathophysiology. Biochemical Journal 438: 397–414.

Willemsen MA, Verbeek MM, Kamsteeg EJ et al. (2010) Tyrosine hydroxylase deficiency: a treatable disorder of brain catecholamine biosynthesis. Brain 133: 1810–1822.

Windahl MS, Petersen CR, Christensen HE and Harris P (2008) Crystal structure of tryptophan hydroxylase with bound amino acid substrate. Biochemistry 47: 12087–12094.

Winge I, McKinney JA, Ying M et al. (2008) Activation and stabilization of human tryptophan hydroxylase 2 by phosphorylation and 14‐3‐3 binding. Biochemical Journal 410: 195–204.

Ying M, Pey AL, Aarsaether N and Martinez A (2010) Phenylalanine hydroxylase expression in primary rat hepatocytes is modulated by oxygen concentration. Molecular Genetics and Metabolism 101: 279–281.

Zhang S, Huang T, Ilangovan U, Hinck AP and Fitzpatrick PF (2014) The solution structure of the regulatory domain of tyrosine hydroxylase. Journal of Molecular Biology 426: 1483–1497.

Further Reading

Bruijnincx PC, van Koten G and Klein Gebbink RJ (2008) Mononuclear non‐heme iron enzymes with the 2‐His‐1‐carboxylate facial triad: recent developments in enzymology and modeling studies. Chemical Society Reviews 37: 2716–2744.

Di Giovanni G, Pessia M and Di Maio R (2012) Redox sensitivity of tyrosine hydroxylase activity and expression in dopaminergic dysfunction. CNS & Neurological Disorders – Drug Targets 11: 419–429.

Kern D and Zuiderweg ER (2003) The role of dynamics in allosteric regulation. Current Opinion in Structural Biology 13: 748–757.

Kobe B and Kemp BE (1999) Active site‐directed protein regulation. Nature 402: 373–376.

Skjevik AA, Mileni M, Baumann A et al. (2014) The N‐terminal sequence of tyrosine hydroxylase is a conformationally versatile motif that binds 14‐3‐3 proteins and membranes. Journal of Molecular Biology 426: 150–168.

van Spronsen FJ (2010) Phenylketonuria: a 21st century perspective. Nature Reviews Endocrinology 6: 509–514.

Torrente MP, Gelenberg AJ and Vrana KE (2012) Boosting serotonin in the brain: is it time to revamp the treatment of depression? Journal of Psychopharmacology 26: 629–635.

Underhaug J, Aubi O and Martinez A (2012) Phenylalanine hydroxylase misfolding and pharmacological chaperones. Current Topics in Medicinal Chemistry 12: 2534–2545.

Yamamoto K and Vernier P (2011) The evolution of dopamine systems in chordates. Frontiers in Neuroanatomy 5: 21. doi:10.3389/fnana.2011.00021.

Zill P, Buttner A, Eisenmenger W et al. (2009) Predominant expression of tryptophan hydroxylase 1 mRNA in the pituitary: a postmortem study in human brain. Neuroscience 159: 1274–1282.

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Skjærven, Lars, Teigen, Knut, and Martinez, Aurora(Aug 2014) Structure–Function Relationships in the Aromatic Amino Acid Hydroxylases Enzyme Family: Evolutionary Insights. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0025581]