Biochemical evaluation of the decarboxylation and decarboxylation-deamination activities of plant aromatic amino acid decarboxylases

J Biol Chem. 2013 Jan 25;288(4):2376-87. doi: 10.1074/jbc.M112.401752. Epub 2012 Nov 30.

Abstract

Plant aromatic amino acid decarboxylase (AAAD) enzymes are capable of catalyzing either decarboxylation or decarboxylation-deamination on various combinations of aromatic amino acid substrates. These two different activities result in the production of arylalkylamines and the formation of aromatic acetaldehydes, respectively. Variations in product formation enable individual enzymes to play different physiological functions. Despite these catalytic variations, arylalkylamine and aldehyde synthesizing AAADs are indistinguishable without protein expression and characterization. In this study, extensive biochemical characterization of plant AAADs was performed to identify residues responsible for differentiating decarboxylation AAADs from aldehyde synthase AAADs. Results demonstrated that a tyrosine residue located on a catalytic loop proximal to the active site of plant AAADs is primarily responsible for dictating typical decarboxylase activity, whereas a phenylalanine at the same position is primarily liable for aldehyde synthase activity. Mutagenesis of the active site phenylalanine to tyrosine in Arabidopsis thaliana and Petroselinum crispum aromatic acetaldehyde synthases primarily converts the enzymes activity from decarboxylation-deamination to decarboxylation. The mutation of the active site tyrosine to phenylalanine in the Catharanthus roseus and Papaver somniferum aromatic amino acid decarboxylases changes the enzymes decarboxylation activity to a primarily decarboxylation-deamination activity. Generation of these mutant enzymes enables the production of unusual AAAD enzyme products including indole-3-acetaldehyde, 4-hydroxyphenylacetaldehyde, and phenylethylamine. Our data indicates that the tyrosine and phenylalanine in the catalytic loop region could serve as a signature residue to reliably distinguish plant arylalkylamine and aldehyde synthesizing AAADs. Additionally, the resulting data enables further insights into the mechanistic roles of active site residues.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Aldehydes / metabolism
  • Amino Acid Sequence
  • Arabidopsis / enzymology
  • Aromatic-L-Amino-Acid Decarboxylases / chemistry*
  • Aromatic-L-Amino-Acid Decarboxylases / physiology*
  • Catalytic Domain
  • Chromatography, Liquid / methods
  • DNA, Complementary / metabolism
  • Deamination
  • Decarboxylation
  • Gene Expression Regulation, Enzymologic*
  • Gene Expression Regulation, Plant*
  • Indoleacetic Acids / chemistry
  • Indoles / chemistry
  • Kinetics
  • Mass Spectrometry / methods
  • Models, Chemical
  • Molecular Sequence Data
  • Mutagenesis
  • Mutation
  • Tryptophan / chemistry

Substances

  • Aldehydes
  • DNA, Complementary
  • Indoleacetic Acids
  • Indoles
  • indole-3-acetaldehyde
  • Tryptophan
  • Aromatic-L-Amino-Acid Decarboxylases