Part:BBa_K200008

Phenylalanine hydroxylase

[http://en.wikipedia.org/wiki/Phenylalanine_hydroxylase Phenylalanine hydroxylase (PAH)] is the enzyme that breaks down [http://en.wikipedia.org/wiki/Phenylalanine phenylalanine] to [http://en.wikipedia.org/wiki/Tyrosine tyrosine]. Deficiency of this enzyme activity has been identified as one of the causes of the autosomal recessive disorder [http://en.wikipedia.org/wiki/Phenylketonuria phenylketonuria (PKU)]#PAH1. Because our project aims to deliver such enzyme to the small intestine, we have also submitted a protease resistant version (BBa_K200028).

Usage and Biology

PAH works well at a PH of around 7.0 (similar to the small intestine), and it is a non-heme iron-dependent enzyme. The enzyme binds three substrates: reduced pterin (PH4), phenylalanine, and oxygen, and releases two products: tyrosine and oxidized pterin #PAH2. Furthermore, the presence of oxygen and an additional cofactor (TPNH) is required for the hydrolysis to take place #PAH4. The enzyme is stable between 7 and 40 degrees celsius#PAH2 and in this range activity increases exponentially with temperature but Km (Michaelis-Menten constant) does not vary significantly between 20 and 40 degrees. The enzyme can be rendered protease resistant via full phosphorylation.#PAH3
Phenylketonuria is a condition whose prevalence varies considerably with populations. In Northern Europe for example the incidence is estimated to 1 in 10000#PAH6. At the other end of the spectrum, Turkey is estimated to have an incidence of 1 in 2600 births#PAH6. This condition is due to the inability of the individuals to catabolise essential amino acid Phenylalanine into Tyrosine. This leads to hyperphenylalaninemia in the blood which, in turn, causes damage to the brain resulting in loss of cognitive abilities and mental retardation [http://www.ncbi.nlm.nih.gov/entrez/dispomim.cgi?id=261600 OMIM]. There are two different types of the disease: classical PKU and BH₄-responsive PKU. The former form of the disease is linked to a deficient PAH enzyme sequence that renders the resulting protein non functional. The latter is due to a deficient co-factor BH₄. Today, only the BH₄ can be tackled, thus encompassing only half of the phenylketonuriac population#PAH5. As part of the Imperial iGEM 2009 [http://2009.igem.org/Team:Imperial_College_London The E.ncapsulator] project, we propose a new, broader therapy strategy where both the functional enzyme and cofactor are delivered to the patient.

Requirements

• Works for PH 7
• Works between 7 to 40 degrees

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 287
  Illegal BamHI site found at 814
  Illegal XhoI site found at 524
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]