Annotated Bibliography for Phenylketonuria

 Blainey, J. D., and R. Gulliford. “Phenylalanine-Restricted Diets in the Treatment of Phenylketonuria.” Archives of Disease in Childhood 31, no. 160 (December 1956): 452–66.

This article was an extension off of the original work by Bickel et al. who identified the most successful treatment for PKU to date. This treatment defined in this article is the implementation of a phenylalanine-restricted diet which successful prevents the neurological pathology due to elevated phenylalanine levels in the blood.


Carluccio, Carla, Franca Fraternali, Francesco Salvatore, Arianna Fornili, and Adriana Zagari. “Structural Features of the Regulatory ACT Domain of Phenylalanine Hydroxylase.” PLoS ONE 8, no. 11 (November 14, 2013). doi:10.1371/journal.pone.0079482.

This article demonstrates the importance of the regulatory domain of PAH. The authors demonstrate a direct link between missense phenylalanine-causing mutations of PAH that results in changes to the hydrophobic interactions at the N-terminal regulatory domain, which affects its catalytic function.


Centerwall, Siegried A., and Willard R. Centerwall. “The Discovery of Phenylketonuria: The Story of a Young Couple, Two Retarded Children, and a Scientist.” Pediatrics 105, no. 1 (January 1, 2000): 89–103.
This article was essential for defining recalling the history of this first inborn error in metabolism, PKU. The discovery of PKU was an interesting story of a Swedish doctor who identified the presence of phenylpyruvate in the urine of patients experiencing an unusual occurence of mental retardation with a musty smelling urine.

Cunningham, Amy, Heather Bausell, Mary Brown, Maggie Chapman, Kari DeFouw, Sharon Ernst, Julie McClure, et al. “Recommendations for the Use of Sapropterin in Phenylketonuria.” Molecular Genetics and Metabolism 106, no. 3 (July 2012): 269–76. doi:10.1016/j.ymgme.2012.04.004.

This article outlines current recommendations for the use of sapropterin in PKU patients. This article attempts to develop a uniform approach for treating PKU along with a low phenylalanine diet, which is the classic approach for preventing PKU’s negative effects. This article will aid in my discussion regarding treatment and therapy using sapropterin, which is a synthetic version of BH4.


Embury, Jennifer E., Catherine E. Charron, Anatoly Martynyuk, Andreas G. Zori, Bin Liu, Syed F. Ali, Neil E. Rowland, and Philip J. Laipis. “PKU Is a Reversible Neurodegenerative Process within the Nigrostriatum That Begins as Early as 4 Weeks of Age in Pahenu2 Mice.” Brain Research 1127, no. 1 (January 5, 2007): 136–50. doi:10.1016/j.brainres.2006.09.101.

I utilized this article for background information on PKU neurodegeneration.


Fitzpatrick, Paul F. “Mechanism of Aromatic Amino Acid Hydroxylation.” Biochemistry 42, no. 48 (December 9, 2003): 14083–91. doi:10.1021/bi035656u.
This source helps elucidate the kinetic mechanism that is in the literature for phenylalanine hydroxylase. The enzymatic reaction is split into the the formation of the Fe(IV)-oxo species and the hydroxylation of phenylalanine to the tyrosine.

Fusetti, Fabrizia, Heidi Erlandsen, Torgeir Flatmark, and Raymond C. Stevens. “Structure of Tetrameric Human Phenylalanine Hydroxylase and Its Implications for Phenylketonuria.” Journal of Biological Chemistry 273, no. 27 (July 3, 1998): 16962–67. doi:10.1074/jbc.273.27.16962.

The authors of this article were able to crystallize human phenylalanine hydroxylase. They identify that the resultant enzyme is in fact a catalytically active tetramer. The authors identify how common mutations implicated with PKU affect protein structural stability at the quaternary structural level.



Gersting, Soren W., Kristina F. Kemter, Michael Staudigl, Dunja D. Messing, Marta K. Danecka, Florian B. Lagler, Christian P. Sommerhoff, Adelbert A. Roscher, and Ania C. Muntau. “Loss of Function in Phenylketonuria Is Caused by Impaired Molecular Motions and Conformational Instability.” American Journal of Human Genetics 83, no. 1 (July 11, 2008): 5–17. doi:10.1016/j.ajhg.2008.05.013.

The authors look at 10 different PAH mutations that have been shown to be BH4-responsive and look at their kinetics, conformation, and stability in order to identify the cause of the PAH deficiency. Most of the mutations pointed to protein misfolding as the cause, due to conformational changes that obstruct its catalytic capabilities. The authors emphasize the importance of the regulatory domain for catalytic functionality and protein stabilization. This article will help explain mechanism and causes of the disease.



Gersting, Søren W, Michael Staudigl, Marietta S Truger, Dunja D Messing, Marta K Danecka, Christian P Sommerhoff, Kristina F Kemter, and Ania C Muntau. “Activation of Phenylalanine Hydroxylase Induces Positive Cooperativity toward the Natural Cofactor.” The Journal of Biological Chemistry 285, no. 40 (October 1, 2010): 30686–97. doi:10.1074/jbc.M110.124016.

The main assertion made in this article that was rather interesting was that there is a relevance to the patient’s metabolic state that plays a more crucial role on drug efficacy and conformation and function of PAH than previously thought. They determined that BH4 cooperativity of the activated enzyme acts independent of phenylalanine, but the cooperativity is due to the activated conformational changes of the enzyme. This was a novel approach to cooperativity of PAH in this field of research up to 2010.


Guthrie, Robert, and Ada Susi. “A Simple Phenylalanine Method for Detecting Phenylketonuria in Large Populations of Newborn Infants.” Pediatrics 32, no. 3 (September 1, 1963): 338–43.
This article demonstrates a significant breakthrough for PKU screening and PKU diet management. This article was written by the creater of the Guthrie screening test that has identified infant PKU patients in order to start immediate implementation of the phenylalanine restricted diet. The Guthrie test is able to monitor phenylalanine concentrations in the blood and urine.

Jaffe, Eileen K, Linda Stith, Sarah H Lawrence, Mark Andrake, and Roland L Dunbrack Jr. “A New Model for Allosteric Regulation of Phenylalanine Hydroxylase: Implications for Disease and Therapeutics.” Archives of Biochemistry and Biophysics 530, no. 2 (February 15, 2013): 73–82. doi:10.1016/

This article discusses a new model of allosteric regulation in rat phenylalanine hydroxylase. The authors identify a morpheem model of PAH allostery and also suggest how many PKU mutations can force conformational changes that result in PAH being less active. In light of the allosteric mechanism identified by the authors, new pharmocological chaperones might offer protein stability by shifting towards the active tetramer PAH in PKU patients with mutations, which have been forced PAH quaternary structure to a less active conformation.



This article demonstrates many theories that could potentially link elevated phenylalanine concentrations with hypomyelination, which could be the cause of the neurological pathology. The direct correlation between the two was not defined, however other hypotheses were explored.


Leandro, João, Nina Simonsen, Jaakko Saraste, Paula Leandro, and Torgeir Flatmark. “Phenylketonuria as a Protein Misfolding Disease: The Mutation pG46S in Phenylalanine Hydroxylase Promotes Self-Association and Fibril Formation.” Biochimica et Biophysica Acta 1812, no. 1 (January 2011): 106–20. doi:10.1016/j.bbadis.2010.09.015.

This paper conducts an in depth analysis of how a missense mutation pG46S in the regulatory domain of human PAH affects the enzyme’s catalytic capabilities via oligomerization changes caused by the mutation. The authors identify a direct link for how this mutation leads to structural differences resulting in fibril formation that can be associated with the severe PKU phenotype. This will give some insight into one of the plethora of different mutations and how it affects phenotype in humans.


Li, Jun, Lawrence J. Dangott, and Paul F. Fitzpatrick. “Regulation of Phenylalanine Hydroxylase: Conformational Changes Upon Phenylalanine Binding Detected by Hydrogen/Deuterium Exchange and Mass Spectrometry.” Biochemistry 49, no. 15 (April 20, 2010): 3327–35. doi:10.1021/bi1001294.

The authors use hydrogen/deuterium exchange monitored by mass spectrometry to demonstrate that the N-terminis of PAH acts as an inhibitory peptide. They demonstrate that there exists a direct link between phenylalanine binding which leads to a conformational change at the regulatory domain, which can affect regulatory and catalytic domain activity. They also show how a mutation that could lead to loss of functionality at the regulatory domain could lead to the disruption of such conformational change in PAH.


Li, Jun, and Paul F. Fitzpatrick. “Regulation of Phenylalanine Hydroxylase: Conformational Changes upon Phosphorylation Detected by H/D Exchange and Mass Spectrometry.” Archives of Biochemistry and Biophysics 535, no. 2 (July 15, 2013): 115–19. doi:10.1016/

The authors explore via hydrogen/deuterium exchange monitored by mass spectrometry how phosphorlyation of Ser16 affects the conformation changes associated with the phosphorylation event. They demonstrated that phosphorylation has less of an impact on PAH conformation changes than that of phenylalanine activation.


Moraes, Tarsila Barros, Giovana Reche Dalazen, Carlos Eduardo Jacques, Raylane Silva de Freitas, Andrea Pereira Rosa, and Carlos Severo Dutra-Filho. “Glutathione Metabolism Enzymes in Brain and Liver of Hyperphenylalaninemic Rats and the Effect of Lipoic Acid Treatment.” Metabolic Brain Disease, February 2, 2014. doi:10.1007/s11011-014-9491-x.

Moraes, Tarsila Barros, Carlos Eduardo Diaz Jacques, Andrea Pereira Rosa, Giovana Reche Dalazen, Melaine Terra, Juliana Gonzalez Coelho, and Carlos Severo Dutra-Filho. “Role of Catalase and Superoxide Dismutase Activities on Oxidative Stress in the Brain of a Phenylketonuria Animal Model and the Effect of Lipoic Acid.” Cellular and Molecular Neurobiology 33, no. 2 (March 2013): 253–60. doi:10.1007/s10571-012-9892-5.

Moraes et al. demonstrate an interesting concern associated with PKU patients experiencing relatively high levels of phenylalanine in the blood. PKU and high levels of phenylalanine have been associated with oxidative stress in the brain because phenylalanine inhibits catalase and phenyllactic acids stimulate superoxide dismutase activity. The authors identify that daily administration of lipoic acid, an antioxidant, can decrease the effect reactive oxygen species, that are associated with high phenylalanine concentrations, in the brain. The authors expand on this analysis in the second paper which discusses the importance of glutathione function in PKU, which also supports treatment with lipoic acid. Both of these papers give good insight into preventative treatment and therapies.

Nowacki, P M, S Byck, L Prevost, and C R Scriver. “PAH Mutation Analysis Consortium Database: 1997. Prototype for Relational Locus-Specific Mutation Databases.” Nucleic Acids Research 26, no. 1 (January 1, 1998): 220–25.

The authors do a global analysis of all known mutations of PAH to date in 1998. There are over 300 mutations of PAH that have been associated with PKU.


Pey, Angel L., Lourdes R. Desviat, Alejandra Gámez, Magdalena Ugarte, and Belén Pérez. “Phenylketonuria: Genotype–phenotype Correlations Based on Expression Analysis of Structural and Functional Mutations in PAH.” Human Mutation 21, no. 4 (April 1, 2003): 370–78. doi:10.1002/humu.10198.

The authors attempted to determine the effects of 18 known PKU mutations. They demonstrated that the amount of mutant PAH expression and its activity can explain why individuals with the same genotype might have varying phenotypes in severity. The authors focused on the genotype and phenotype relationship, which could offer a different understanding of PKU at a DNA level all the way to the physical phenotype level.


Pietz, J, R Kreis, A Rupp, E Mayatepek, D Rating, C Boesch, and H J Bremer. “Large Neutral Amino Acids Block Phenylalanine Transport into Brain Tissue in Patients with Phenylketonuria.” The Journal of Clinical Investigation 103, no. 8 (April 1999): 1169–78. doi:10.1172/JCI5017.
This article demonstrated a potential driving force behind neurological pathology between increased phenylalanine blood levels in PKU patients and the phenylalanine competition across the blood brain barrier. The authors also demonstrated that supplementation with other large neutral amino acids could contradict the increased transport of phenylalanine across the blood brain barrier.

Roberts, Kenneth M., Jorge Alex Pavon, and Paul F. Fitzpatrick. “Kinetic Mechanism of Phenylalanine Hydroxylase: Intrinsic Binding and Rate Constants from Single-Turnover Experiments.” Biochemistry 52, no. 6 (February 12, 2013): 1062–73. doi:10.1021/bi301675e.

This article specifically focuses on the kinetic mechanism of the catalytic portion of rat phenylalanine hydroxylase enzyme, excluding the effects of allostery by omitting the regulatory subunit. Complete mechanism was demonstrated with experimental proof, and this article will contain most of the information in regards to mechanism.


Sarkissian, Christineh N, and Alejandra Gámez. “Phenylalanine Ammonia Lyase, Enzyme Substitution Therapy for Phenylketonuria, Where Are We Now?” Molecular Genetics and Metabolism 86 Suppl 1 (December 2005): S22–26. doi:10.1016/j.ymgme.2005.06.016.
This source was helpful for describing the progression of a new treatment of utilizing phenylalanine ammonia lyase which will be PEGylated in order to surpass our immune system and decrease phenylalanine concentration in the blood. Phenylalanine ammonia lyase converts phenylalanine to trans-cinnamic acid, which is a non-toxic metabolite.

Vockley, Jerry, Hans C. Andersson, Kevin M. Antshel, Nancy E. Braverman, Barbara K. Burton, Dianne M. Frazier, John Mitchell, et al. “Phenylalanine Hydroxylase Deficiency: Diagnosis and Management Guideline.” Genetics in Medicine 16, no. 2 (February 2014): 188–200. doi:10.1038/gim.2013.157.

This article was published in February of 2014 as a means to collect all the known treatment and screening of PKU in order to give a PKU diagnosis and management guideline. This article’s purpose is to be an educational resource for clinicians to treat PKU, and I will use this article to demonstrate the field’s approach to treating PKU currently.

One Reply to “Annotated Bibliography for Phenylketonuria”

  1. The Carluccio et al paper is a Molecular Modeling/Dynamics Simulation paper – no wet/bench science. That should be mentioned in your 1-3 sentence summary.

    Also, even if we do not have access to the full text of the paper – the link should be to through the DOI to the journal itself (rather than PubMed) unless the paper is too old to be online (e.g. the Moraes paper)

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