L-tryptophan 2',3'-oxidase from Chromobacterium violaceum. Substrate specificity and mechanistic implications.
L-Tryptophan 2',3'-oxidase, an amino acid alpha,beta-dehydrogenase isolated from Chromobacterium violaceum, catalyzes the formation of a double bond between the C alpha and C beta carbons of various tryptophan derivatives provided that they possess: (i) a L-enantiomeric configuration, (ii) an alpha-carbonyl group, and (iii) an unsubstituted and unmodified indole nucleus. Kinetic parameters were evaluated for a series of tryptophan analogues, providing information on the contribution of each chemical group to substrate binding. The stereochemistry of the dehydro product was determined to be a Z-configuration from proton nuclear magnetic resonance assignments. No reaction can be observed in the presence of other aromatic beta-substituted alanyl residues which behave neither as substrates nor as inhibitors and therefore do not compete against this reaction. The enzymatic synthesis of alpha,beta-dehydrotryptophanyl peptides from 5 to 24 residues was successfully achieved without side product formation, irrespective of the position of the tryptophan residue in the amino acid sequence. A reactional mechanism involving a direct alpha,beta-dehydrogenation of the tryptophan side chain is proposed.
Genet
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CEA/Saclay, Département d'Ingénierie et d'Etudes des Protéines, F91191 Gif-sur-Yvette, France.
Bénetti
P H
PH
Hammadi
A
A
Ménez
A
A
D000595:Amino Acid Sequence; D001665:Binding Sites; D002861:Chromobacterium / Q000201:enzymology*; D007211:Indoles / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008969:Molecular Sequence Data; D010088:Oxidoreductases / Q000378:metabolism*; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D013237:Stereoisomerism; D013379:Substrate Specificity; D014364:Tryptophan / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism
D007211:Indoles; D010455:Peptides; C016952:N-acetyltryptophanamide; D014364:Tryptophan; D010088:Oxidoreductases; C088650:L-tryptophan 2',3'-oxidase