More than just recruitment: the X-domain influences catalysis of the first phenolic coupling reaction in A47934 biosynthesis by Cytochrome P450 StaH
| Autor: | Madeleine Peschke, Max J. Cryle, Clara Brieke, Veronika Ulrich |
|---|---|
| Jazyk: | angličtina |
| Rok vydání: | 2016 |
| Předmět: |
0301 basic medicine
Models Molecular Stereochemistry medicine.drug_class Molecular Conformation Peptide Biology Glycopeptide antibiotic Substrate Specificity 03 medical and health sciences chemistry.chemical_compound Enzyme activator Biosynthesis Cytochrome P-450 Enzyme System Phenols Oxidoreductase Catalytic Domain Aromatic amino acids medicine Protein Interaction Domains and Motifs Molecular Biology chemistry.chemical_classification Molecular Structure Spectrum Analysis Glycopeptides Glycopeptide Recombinant Proteins Enzyme Activation 030104 developmental biology Enzyme chemistry Ristocetin Cyclization Biotechnology |
| Zdroj: | Molecular BioSystems |
| Popis: | Glycopeptide antibiotic biosynthesis involves a complex cascade of reactions centred on a non-ribosomal peptide synthetase and modifiying proteins acting in trans, such as Cytochrome P450 enzymes. These P450s are responsible for cyclisation of the peptide via cross-linking aromatic amino acid side chains, which are a hallmark of the glycopeptide antibiotics. Here, we analysed the first cyclisation reaction in the biosynthesis of the glycopeptide antibiotic A47934. Our results demonstrate that the P450 StaH is recruited to the NRPS machinery through interaction with the X-domain present in the last A47934 NRPS module. We determined the crystal structure of StaH and showed that it is responsible for the first cyclisation in A47934 biosynthesis and additionally exhibits flexible substrate specificity. Our results further point out that the X-domain has an impact on the efficiency of the in vitro cyclisation reaction: hybrid PCP-X constructs obtained by domain exchange between A47934 and teicoplanin biosynthesis NRPS modules reveal that the X-domain from A47934 leads to decreased P450 activity and alternate stereochemical preference for the substrate peptide. We determined that a tight interaction between StaH and the A47934 X-domain correlates with decreased in vitro P450 activity: this highlights the need for glycopeptide antibiotic cyclisation to be a dynamic system, with an overly tight interaction interfering with substrate turnover in vitro. |
| Databáze: | OpenAIRE |
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