RINAD A. ALJOHANI

THERAPEUTIC APPLICATIONS OF PHENYLALANINE AMMONIA LYASE AND

PHENYLALANINE HYDROXYLSE IN PHENYLKETONURIA

WHAT IS PHENYLKETONURIA

• Phenylketonuria is an inherited disorder of amino acid metabolism caused by phenylalanine hydroxylase deficiency• It is characterized with high levels of the amino

acid phenylalanine in the blood• The patients urine has a characteristic

“mousey” odor • If not treated it would cause behavioral

problems, seizures, mental retardation, and failure to grow.

WHAT IS PHENYLKETONURIA

• Early diagnosis of PKU is important because a lot of the central nervous system symptoms can be avoided by controlled level of (Phe) through strict diet with no very low protein intake. Although, simple dietary restriction of (Phe) levels will not reverse the central nervous system effects due to neurotransmitters deficiencies.

PHENYLALANINE HYDROXYLSE• 1. What is Phenylalanine Hydroxylse• Phenylalanine Hydroxylse PAH is an enzyme

responsible for the conversion of the amino acid phenylalanine (phe) to tyrosine

• 2. Structure Phenylalanine Hydroxylse• PAH consist of two or four identical subunits,

present in equilibrium it has two forms and is is regulated by phenylalanine (phe), tetrahydropiopterin (BH4), and phosphorylation

STRUCTURE PHENYLALANINE HYDROXYLSE

FUNCTION OF PHENYLALANINE HYDROXYLSE

• It has two binding sites, a catalytic site, and a regulatory site. The binding site of which is located near the hinge domain might be the factor that causes the drastic change in the enzyme’s shape which will result in (Phe) substrate cleavage.

• In order for the PAH to catalyze the (Phe) in the dephosphorylated state, (Phe) levels must be significantly high enough for this step to occur.

DIETARY TREATMENT OF PHENYLKETONURIA

• 1. Current treatment of PKU• For decades patients with PKU had to maintain their

(Phe) levels through their diet. The primary therapeutic approach has been to lower the elevated (Phe) concentrations to be within the therapeutic range as higher (Phe) concentrations are considered toxic

• There have been newer approaches in dietary management of PKU such as Tetrahydrobiopterin and other pharmacological chaperones, large neutral amino acids (LAAs), and Glycomacropeptide.

MATERNAL PKU:

• When female patients of PKU become pregnant while their dietary levels of (Phe) exceed the required low range the offspring will get affected with “maternal PKU Syndrome”. • Increased levels of (Phe) in the blood will cause

mental retardation, intrauterine growth retardation• dietary control of (Phe) levels in the blood must

begin prior to conception and be maintained through-out the pregnancy to avoid damaging the fetus.

A NEW ERA FOR THE TREATMENT OF PHENYLKETONURIA

• So far treatment for PKU has been Successful in many ways but something better is needed. New treatments for PKU are emerging right now. Such as Sapropterin which is a synthetic form of tetrahydrobiopterin, the cofactor in the PAH reaction. There is also enzyme replacement therapy (ERT) with two enzyme systems that are being developed for PKU treatment (PAH enzyme and the Phe-degrading enzyme phenylalanine ammonia-lyase (PAL))

1. TREATMENT OF PKU USING PHENYLALANINE AMMONIA LYASE

• PAL is an enzyme substitution therapy that break down (Phe) into ammonia and trans-cinnamic acid

• PAL exists in bacteria and yeast; not in mammals.

• In therapy in order to reduce its immunogenicity it is conjugated with polyethylene glycol (PEG).

• Subcutaneous administration of PAL result in lowering plasma and brain (Phe) concentrations. However the metabolic effect was not sustained due to an immune response

. TREATMENT OF PKU USING PHENYLALANINE AMMONIA LYASE

• Scientists have chemically modified PAL by pegylation producing a form of PAL with higher more prolonged half-life, better specific activity, and reduced immunogenicity to not cause serious immune reactions.

• PAL therapy is more favorable than PAH because it requires no cofactors for degrading the (Phe) and its metabolites has low toxicity levels and no embryotoxic effects. in experimental animals, The PAL was very stable under wide range of temperatures.

. TREATMENT OF PKU USING PHENYLALANINE AMMONIA LYASE

2. ENZYME REPLACEMENT THERAPY USING PAH

• Since PAH is the enzyme involved in Phe metabolism, it comes as a natural choice for enzyme based therapeutics in the management of PKU.

• PAH is not an autocatalytic enzyme, it requires several co-factors to function which make the therapy quite complex and challenging because it would mean having to maintain a multi-component enzyme in a stable state which is a herculean task.

CONCLUSIONS

• The future is very bright for PKU patients with the considerable advances in ERT to treat PKU. Multiple treatment options are now available

• These include the use of various forms of PAH, PEGylated PAH as well as the Phe degrading enzyme PAL.

• Although a great deal of work has been conducted to date, there are still problems to overcome, including the stability of the enzymes, consistency of response, and avoiding immune responses.