Cyclic dipeptide oxidase from S treptomyces noursei

Abstract

Cyclic dipeptide oxidase is a novel enzyme that specifically catalyzes the formation of α,β‐dehydro‐Phe (ΔPhe) and α,β‐dehydro‐Leu (ΔLeu) residues during the biosynthesis of albonoursin, cyclo(ΔPhe‐ΔLeu), an antibiotic produced by Streptomyces noursei . It was purified 600‐fold with a 30% overall recovery, and consists of the association of a single type of subunit with a relative molecular mass of 21 066 resulting in a large homopolymer of relative molecular mass over 2 000 000. The enzyme exhibits a typical flavoprotein spectrum with maxima at 343.5 and 447.5 nm, the flavin prosthetic group being covalently bound to the protein. The catalytic reaction of the natural substrate cyclo( l ‐Phe‐ l ‐Leu) occurs in a two‐step sequential reaction leading first to cyclo(α,β‐dehydro‐Phe‐ l ‐Leu) and finally to albonoursin. Kinetic parameters for the first step were determined ( K m = 53 µ m ; k = 0.69 s −1 ). The enzyme was shown to catalyze the conversion of a variety of cyclo(dipeptides) and can be reoxidized at the expense of molecular oxygen by producing H 2 O 2 . This reaction mechanism, which differs from those already described for the formation of α,β‐dehydro‐amino acids, might consist of the transient formation of an intermediate imine followed by its rearrangement into an α,β‐dehydro‐residue.