Structural insights from a P450 Carrier Protein complex reveal how specificity is achieved in the P450_BioIACP complex

Abstract

Cytochrome P450BioI (CYP107H1) from the biotin operon of Bacillus subtilis forms a seven-carbon diacid through a multistep oxidative cleavage of a fatty acid linked to acyl carrier protein (ACP). Crystal structures of P450 BioI in complex with three different length fatty acyl-ACP (Escherichia coli) ligands show that P450 BioI binds the fatty acid such as to force the carbon chain into a U-shape above the active site heme. This positions the C7 and C8 carbons for oxidation, with a large additional cavity extending beyond the heme to accommodate the methyl termini of fatty acids beyond the site of cleavage. The structures explain the experimentally observed lack of stereo-and regiospecificity in the hydroxylation and cleavage of free fatty acids. The P450 BioI-ACP complexes represent the only structurally characterized P450-carrier protein complexes to date, which has allowed the generation of a model of the interaction of the vancomycin biosynthetic P450 OxyB with its proposed carrier protein bound substrate.acyl carrier protein ͉ biotin biosynthesis ͉ crystal structure ͉ cytochrome P450 ͉ protein-protein interaction

Structural Characterization of OxyD, a Cytochrome P450 Involved in β-Hydroxytyrosine Formation in Vancomycin Biosynthesis

“…Exceptions are: (1) Small residue (S, G, A), (2) Positively Charged Residue (K, H, R), (3) Majority Hydrophobic (L, V; also S and G), (4) Majority Small (S, A; also V). b Percentage of residues matching to the overall consensus rule, based upon similarity.…”

“…P450s that utilize carrier protein-bound substrates form a small but significant subgroup of the superfamily. One example is P450 BioI , which is responsible for the carbon-carbon bond cleavage of acyl carrier protein-bound fatty acids during the first step of biotin biosynthesis in Bacillus subtilis (1). Additional examples are found in the biosynthesis of the vancomycin-type antibiotics: balhimycin (10), chloroeremomycin (11), and vancomycin, produced by different strains of Amycolatopsis ( Fig.…”

Using the pimeloyl-CoA synthetase adenylation fold to synthesize fatty acid thioesters

“…Subsequently, crystal structures of BioI captured in complex with acyl-ACPs of varying chain lengths have provided a mechanistic understanding of how heme chemistry is used to act upon the acyl chain and generate pimelic acid 20,21 . BioW from Bacillus sphaericus has been characterised to have pimeloyl-CoA synthetase activity (EC number 6.2.1.14) 22,23 .…”

Structural analysis of HmtT and HmtN involved in the tailoring steps of himastatin biosynthesis

“…P450 BioI is the only protein in the PDB database whose F-G loop adopts a similar conformation as that of HmtT [22]. In the complex structure of P450 BioI and its substrate, the space directly above the heme prosthetic group is partially occupied by the F-G loop while the substrate reaches the heme group from the side.…”

“…HmtT, HmtN and HmtS have an F-G loop of the same length while the loops in EryK from Saccharopolyspora erythraea and P450 BioI (CYP107H1) from Bacillus subtilis are much shorter. The additional residues in the F-G loop could increase the flexibility of this region, which may facilitate the transportation of exceptionally large substrates of these enzymes [21,22].…”