Exploring the substrate specificity of OxyB, a phenol coupling P450 enzyme involved in vancomycin biosynthesis.
OxyB is a cytochrome P450 enzyme that catalyzes the first oxidative phenol coupling reaction during vancomycin biosynthesis. The preferred substrate is a linear peptide linked as a C-terminal thioester to a peptide carrier protein (PCP) domain of the glycopeptide antibiotic non-ribosomal peptide synthetase. Previous studies have shown that OxyB can efficiently oxidize a model hexapeptide-PCP conjugate (R-Leu(1)-R-Tyr(2)-S-Asn(3)-R-Hpg(4)-R-Hpg(5)-S-Tyr(6)-S-PCP) (Hpg = 4-hydroxyphenylglycine) into a macrocyclic product by phenolic coupling of the aromatic rings in residues-4 and -6. In this work, the substrate specificity of OxyB has been explored using a series of N-terminally truncated peptides related in sequence to this model hexapeptide-PCP conjugate. Deletion of one or three residues from the N-terminus afforded a penta- (Ac-Tyr-Asn-Hpg-Hpg-Tyr-S-PCP) and a tri- (Ac-Hpg-Hpg-Tyr-S-PCP) peptide that were also efficiently transformed into the corresponding macrocyclic cross-linked product by OxyB. The tripeptide, representing the core of the macrocycle in vancomycin created by OxyB, is thus sufficient, as a thioester with the PCP domain, for phenol coupling to occur. The related tetrapeptide-PCP thioester was not cyclized by OxyB, neither was a related model hexapeptide containing tryptophan in place of tyrosine-6, nor were tripeptides (related to the natural product K-13) with the sequence Ac-Tyr-Tyr-Tyr-S-PCP cross-linked by OxyB.
Woithe
Katharina
K
Chemistry Department, University of Zurich, Winterthurerstrasse 190, 8057, Zurich, Switzerland.
Geib
Nina
N
Meyer
Odile
O
Wörtz
Tanja
T
Zerbe
Katja
K
Robinson
John A
JA
D001665:Binding Sites; D003577:Cytochrome P-450 Enzyme System / Q000737:chemistry* / Q000378:metabolism; D015394:Molecular Structure; D010636:Phenols / Q000737:chemistry*; D011987:Receptors, Steroid / Q000737:chemistry* / Q000378:metabolism; D013379:Substrate Specificity; D014640:Vancomycin / Q000096:biosynthesis*; D000098825:Oxysterol Binding Proteins
D010636:Phenols; D011987:Receptors, Steroid; D000098825:Oxysterol Binding Proteins; D014640:Vancomycin; D003577:Cytochrome P-450 Enzyme System