page_title: Phenylalanine ammonia-lyase
section_title: Mechanism
hierarchical_section_title: Phenylalanine ammonia-lyase / Mechanism
caption_attribution_description: English: Proposed autocatalytic formation of MIO from the tripeptide Ala-Ser-Gly by two water elimination steps
context_page_description: Phenylalanine ammonia lyase is an enzyme that catalyzes a reaction converting L-phenylalanine to ammonia and trans-cinnamic acid. Phenylalanine ammonia lyase is the first and committed step in the phenyl propanoid pathway and is therefore involved in the biosynthesis of the polyphenol compounds such as flavonoids, phenylpropanoids, and lignin in plants. Phenylalanine ammonia lyase is found widely in plants, as well as some bacteria, yeast, and fungi, with isoenzymes existing within many different species. It has a molecular mass in the range of 270–330 kDa. The activity of PAL is induced dramatically in response to various stimuli such as tissue wounding, pathogenic attack, light, low temperatures, and hormones. PAL has recently been studied for possible therapeutic benefits in humans afflicted with phenylketonuria. It has also been used in the generation of L-phenylalanine as precursor of the sweetener aspartame.
  The enzyme is a member of the ammonia lyase family, which cleaves carbon–nitrogen bonds. Like other lyases, PAL requires only one substrate for the forward reaction, but two for the reverse.
context_section_description: Phenylalanine ammonia lyase is specific for L-phe, and to a lesser extent, L-tyrosine. The reaction catalyzed by PAL is a spontaneous elimination reaction rather than an oxidative deamination.
  L-phenylalanine  trans-cinnamic acid + NH₃
  The cofactor 3,5-dihydro-5-methyldiene-4H-imidazol-4-one (MIO) is involved in the reaction and sits atop the positive pole of three polar helices in the active site, which helps to increase its electrophilicity. MIO is attacked by the aromatic ring of L-phe, which activates the C-H bond on the β carbon for deprotonation by a basic residue. The carbanion intermediate of this E1cB-elimination reaction, which is stabilized by partial positive regions in the active site, then expels ammonia to form the cinnamate alkene. The mechanism of the reaction of PAL is thought to be similar to the mechanism of the related enzyme histidine ammonia lyase.
  A dehydroalanine residue was long thought to be the key electrophilic catalytic residue in PAL and HAL, but the active residue was later found instead to be MIO, which is even more electrophilic. It is formed by cyclization and dehydration of conserved Ala-Ser-Gly tripeptide segment. The first step of MIO formation is a cyclization-elimination by an intramolecular nucleophilic attack of the nitrogen of Gly204 at the carbonyl group of Ala202. A subsequent water elimination from the side chain of Ser203 completes the system of crossconjugated double bonds. Numbers are given for the phenylalanine ammonia lyase from Petroselinum crispum (PDB 1W27). Although MIO is a polypeptide modification, it was proposed to call it a prosthetic group, because it has the quality of an added organic compound.
  PAL is inhibited by trans-cinnamic acid, and, in some species, may be inhibited by trans-cinnamic acid derivatives. The unnatural amino acids D-Phe and D-Tyr, the enantiomeric forms of the normal substrate, are competitive inhibitors.
caption: Proposed autocatalytic formation of MIO cofactor from the tripeptide Ala-Ser-Gly by two water elimination steps.[15]
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