L-TRYPTOPHAN 2',3'-OXIDASE FROM CHROMOBACTERIUM-VIOLACEUM - SUBSTRATE-SPECIFICITY AND MECHANISTIC IMPLICATIONS

L-Tryptophan 2',3'-oxidase, an amino acid alpha,beta-dehydrogenase iso lated from Chromobacterium violaceum, catalyzes the formation of a dou ble bond between the C-alpha and C-beta carbons of various tryptophan derivatives provided that they possess: (i) a L-enantiomeric configura tion, (ii) an alpha-carbonyl group, and (iii) an unsubstituted and unm odified 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 nuc lear magnetic resonance assignments. No reaction can be observed in th e presence of other aromatic beta-substituted alanyl residues which be have neither as substrates nor as inhibitors and therefore do not comp ete against this reaction. The enzymatic synthesis of alpha,beta-dehyd rotryptophanyl peptides from 5 to 24 residues was successfully achieve d 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 c hain is proposed.