Functional analysis and crystallographic structure of clotrimazole bound OleP, a cytochrome P450 epoxidase from Streptomyces antibioticus involved in oleandomycin biosynthesis

Montemiglio, Linda Celeste; Parisi, Giacomo; Scaglione, Antonella; Sciara, Giuliano; Savino, C.; Vallone, B.

doi:10.1016/j.bbagen.2015.10.009

Cited by 19 publications

(41 citation statements)

References 43 publications

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“…This unique feature clearly sets this group of enzymes apart from the structurally related non‐Oxy P450 enzymes identified by Dali search for both molecules present in the asymmetric unit (e.g. CYP105‐P1 /‐AS1 /‐N1 , MycG , P450 nor and OleP ). Compared to the OxyB and OxyC structures, in OxyA tei these helices are pulled down toward the heme, whereas in the OxyE structure these helices adopt an even more closed conformation (Fig.…”

Section: Resultsmentioning

confidence: 88%

“…This structure not only completes the collection of peptide tailoring P450s involved in GPA biosynthesis but also reveals striking differences compared to the previously determined structures of OxyB, C and E. In particular the central cavity of OxyA is narrowed by the presence of an additional N terminal helix, the A'‐helix and the orientation of the F‐ and G‐helices toward the I‐helix. Since this closed conformation of the F‐ and G‐helices is more prominent in protomer A with the active site occupied by the N terminus of the protomer B, it is highly likely that binding of the N terminus induced a closing of the capping helices in a similar manner as described for ligand‐bound P450s, yet with less dramatic conformational changes . In the literature, there is one example of a P450 forming a homodimer in the crystal in a similar manner as OxyA tei , the structure of P450 sky (PBD: 4LOE) .…”

Section: Discussionmentioning

confidence: 92%

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Structure of OxyA_tei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Haslinger

Cryle

2016

FEBS Letters

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Edited by Richard CogdellCyclization of glycopeptide antibiotic precursors occurs in either three or four steps catalyzed by Cytochrome P450 enzymes. Three of these enzymes have been structurally characterized to date with the second enzyme along the pathway, OxyA, escaping structural analysis. We are now able to present the structure of OxyA tei involved in teicoplanin biosynthesis -the same enzyme recently shown to be the first active OxyA homolog. In spite of the hydrophobic character of the teicoplanin precursor, the polar active site of OxyA tei and its affinity for certain azole inhibitors hint at its preference for substrates with polar decorations.

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Section: Resultsmentioning

confidence: 88%

Section: Discussionmentioning

confidence: 92%

Structure of OxyA_tei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Haslinger

Cryle

2016

FEBS Letters

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“…Other P450 enzymes with high structural similarity to StaF are those from secondary metabolism and involve oxidative functionalisation of large substrates, such as pravastatin (CYP105AS, PDB ID: 4OQS) [42], oleandomycin (OleP, PDB ID: 4XE3) [43], mycinamicin (MycG, PDB ID: 2YCA) [44] and filipin (CYP105P1, PDB ID: 3E5L) [45] (Table 3). StaF also shows moderate levels of structural similarity to other P450s that oxidise carrier protein-bound substrates, including the fattyacyl-ACP oxidase P450 BioI (PDB ID: 3EJD) [46–48] and the aminoacyl-PCP hydroxylases OxyD (PDB ID: 3MGX) and P450 sky (PDB ID: 4PXH) [49–50] (Table 3).…”

Section: Resultsmentioning

confidence: 99%

Biochemical and structural characterisation of the second oxidative crosslinking step during the biosynthesis of the glycopeptide antibiotic A47934

Ulrich¹,

Brieke²,

Cryle³

2016

Beilstein J. Org. Chem.

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The chemical complexity and biological activity of the glycopeptide antibiotics (GPAs) stems from their unique crosslinked structure, which is generated by the actions of cytochrome P450 (Oxy) enzymes that affect the crosslinking of aromatic side chains of amino acid residues contained within the GPA heptapeptide precursor. Given the crucial role peptide cyclisation plays in GPA activity, the characterisation of this process is of great importance in understanding the biosynthesis of these important antibiotics. Here, we report the cyclisation activity and crystal structure of StaF, the D-O-E ring forming Oxy enzyme from A47934 biosynthesis. Our results show that the specificity of StaF is reduced when compared to Oxy enzymes catalysing C-O-D ring formation and that this activity relies on interactions with the non-ribosomal peptide synthetase via the X-domain. Despite the interaction of StaF with the A47934 X-domain being weaker than for the preceding Oxy enzyme StaH, StaF retains higher levels of in vitro activity: we postulate that this is due to the ability of the StaF/X-domain complex to allow substrate reorganisation after initial complex formation has occurred. These results highlight the importance of testing different peptide/protein carrier constructs for in vitro GPA cyclisation assays and show that different Oxy homologues can display significantly different catalytic propensities despite their overall similarities.

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“…3), is unique in that it catalyzes epoxidation of two aliphatic carbons, although there is evidence that this transformation proceeds through an OleP-generated olefinic intermediate. 72,73 …”

Section: Functionmentioning

confidence: 99%

Cytochromes P450 for natural product biosynthesis in Streptomyces: sequence, structure, and function

et al. 2017

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Cytochrome P450 enzymes (P450s) are some of the most exquisite and versatile biocatalysts found in nature. In addition to their well-known roles in steroid biosynthesis and drug metabolism in humans, P450s are key players in natural product biosynthetic pathways. Natural products, the most chemically and structurally diverse small molecules known, require an extensive collection of P450s to accept and functionalize their unique scaffolds. In this review, we survey the current catalytic landscape of P450s within the Streptomyces genus, one of the most prolific producers of natural products, and comprehensively summarize the functionally characterized P450s from Streptomyces. A sequence similarity network of >8500 P450s revealed insights into the sequence–function relationships of these oxygen-dependent metalloenzymes. Although only ~2.4% and <0.4% of streptomycete P450s have been functionally and structurally characterized, respectively, the study of streptomycete P450s involved in the biosynthesis of natural products has revealed their diverse roles in nature, expanded their catalytic repertoire, created structural and mechanistic paradigms, and exposed their potential for biomedical and biotechnological applications. Continued study of these remarkable enzymes will undoubtedly expose their true complement of chemical and biological capabilities.

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Functional analysis and crystallographic structure of clotrimazole bound OleP, a cytochrome P450 epoxidase from Streptomyces antibioticus involved in oleandomycin biosynthesis

Cited by 19 publications

References 43 publications

Structure of OxyA_tei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Structure of OxyA_tei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Biochemical and structural characterisation of the second oxidative crosslinking step during the biosynthesis of the glycopeptide antibiotic A47934

Cytochromes P450 for natural product biosynthesis in Streptomyces: sequence, structure, and function

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Functional analysis and crystallographic structure of clotrimazole bound OleP, a cytochrome P450 epoxidase from Streptomyces antibioticus involved in oleandomycin biosynthesis

Cited by 19 publications

References 43 publications

Structure of OxyAtei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Structure of OxyAtei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Biochemical and structural characterisation of the second oxidative crosslinking step during the biosynthesis of the glycopeptide antibiotic A47934

Cytochromes P450 for natural product biosynthesis in Streptomyces: sequence, structure, and function

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Structure of OxyA_tei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade

Structure of OxyA_tei: completing our picture of the glycopeptide antibiotic producing Cytochrome P450 cascade