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Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate
The cytochrome P450 OleP catalyzes the epoxidation of aliphatic carbons on both the aglycone 8.8a-deoxyoleandolide (DEO) and the monoglycosylated L-olivosyl-8.8a-deoxyoleandolide (L-O-DEO) intermediates of oleandomycin biosynthesis. We investigated the substrate versatility of the enzyme. X-ray and...
Autores principales: | , , , , , , , , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
MDPI
2020
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7600006/ https://www.ncbi.nlm.nih.gov/pubmed/33036250 http://dx.doi.org/10.3390/biom10101411 |
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author | Parisi, Giacomo Freda, Ida Exertier, Cécile Cecchetti, Cristina Gugole, Elena Cerutti, Gabriele D’Auria, Lucia Macone, Alberto Vallone, Beatrice Savino, Carmelinda Montemiglio, Linda Celeste |
author_facet | Parisi, Giacomo Freda, Ida Exertier, Cécile Cecchetti, Cristina Gugole, Elena Cerutti, Gabriele D’Auria, Lucia Macone, Alberto Vallone, Beatrice Savino, Carmelinda Montemiglio, Linda Celeste |
author_sort | Parisi, Giacomo |
collection | PubMed |
description | The cytochrome P450 OleP catalyzes the epoxidation of aliphatic carbons on both the aglycone 8.8a-deoxyoleandolide (DEO) and the monoglycosylated L-olivosyl-8.8a-deoxyoleandolide (L-O-DEO) intermediates of oleandomycin biosynthesis. We investigated the substrate versatility of the enzyme. X-ray and equilibrium binding data show that the aglycone DEO loosely fits the OleP active site, triggering the closure that prepares it for catalysis only on a minor population of enzyme. The open-to-closed state transition allows solvent molecules to accumulate in a cavity that forms upon closure, mediating protein–substrate interactions. In silico docking of the monoglycosylated L-O-DEO in the closed OleP–DEO structure shows that the L-olivosyl moiety can be hosted in the same cavity, replacing solvent molecules and directly contacting structural elements involved in the transition. X-ray structures of aglycone-bound OleP in the presence of L-rhamnose confirm the cavity as a potential site for sugar binding. All considered, we propose L-O-DEO as the optimal substrate of OleP, the L-olivosyl moiety possibly representing the molecular wedge that triggers a more efficient structural response upon substrate binding, favoring and stabilizing the enzyme closure before catalysis. OleP substrate versatility is supported by structural solvent molecules that compensate for the absence of a glycosyl unit when the aglycone is bound. |
format | Online Article Text |
id | pubmed-7600006 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2020 |
publisher | MDPI |
record_format | MEDLINE/PubMed |
spelling | pubmed-76000062020-11-01 Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate Parisi, Giacomo Freda, Ida Exertier, Cécile Cecchetti, Cristina Gugole, Elena Cerutti, Gabriele D’Auria, Lucia Macone, Alberto Vallone, Beatrice Savino, Carmelinda Montemiglio, Linda Celeste Biomolecules Article The cytochrome P450 OleP catalyzes the epoxidation of aliphatic carbons on both the aglycone 8.8a-deoxyoleandolide (DEO) and the monoglycosylated L-olivosyl-8.8a-deoxyoleandolide (L-O-DEO) intermediates of oleandomycin biosynthesis. We investigated the substrate versatility of the enzyme. X-ray and equilibrium binding data show that the aglycone DEO loosely fits the OleP active site, triggering the closure that prepares it for catalysis only on a minor population of enzyme. The open-to-closed state transition allows solvent molecules to accumulate in a cavity that forms upon closure, mediating protein–substrate interactions. In silico docking of the monoglycosylated L-O-DEO in the closed OleP–DEO structure shows that the L-olivosyl moiety can be hosted in the same cavity, replacing solvent molecules and directly contacting structural elements involved in the transition. X-ray structures of aglycone-bound OleP in the presence of L-rhamnose confirm the cavity as a potential site for sugar binding. All considered, we propose L-O-DEO as the optimal substrate of OleP, the L-olivosyl moiety possibly representing the molecular wedge that triggers a more efficient structural response upon substrate binding, favoring and stabilizing the enzyme closure before catalysis. OleP substrate versatility is supported by structural solvent molecules that compensate for the absence of a glycosyl unit when the aglycone is bound. MDPI 2020-10-06 /pmc/articles/PMC7600006/ /pubmed/33036250 http://dx.doi.org/10.3390/biom10101411 Text en © 2020 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/). |
spellingShingle | Article Parisi, Giacomo Freda, Ida Exertier, Cécile Cecchetti, Cristina Gugole, Elena Cerutti, Gabriele D’Auria, Lucia Macone, Alberto Vallone, Beatrice Savino, Carmelinda Montemiglio, Linda Celeste Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate |
title | Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate |
title_full | Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate |
title_fullStr | Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate |
title_full_unstemmed | Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate |
title_short | Dissecting the Cytochrome P450 OleP Substrate Specificity: Evidence for a Preferential Substrate |
title_sort | dissecting the cytochrome p450 olep substrate specificity: evidence for a preferential substrate |
topic | Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7600006/ https://www.ncbi.nlm.nih.gov/pubmed/33036250 http://dx.doi.org/10.3390/biom10101411 |
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