Cargando…

Structure of a low-population intermediate state in the release of an enzyme product

Enzymes can increase the rate of biomolecular reactions by several orders of magnitude. Although the steps of substrate capture and product release are essential in the enzymatic process, complete atomic-level descriptions of these steps are difficult to obtain because of the transient nature of the...

Descripción completa

Detalles Bibliográficos
Autores principales: De Simone, Alfonso, Aprile, Francesco A, Dhulesia, Anne, Dobson, Christopher M, Vendruscolo, Michele
Formato: Online Artículo Texto
Lenguaje:English
Publicado: eLife Sciences Publications, Ltd 2015
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4383205/
https://www.ncbi.nlm.nih.gov/pubmed/25575179
http://dx.doi.org/10.7554/eLife.02777
_version_ 1782364694686728192
author De Simone, Alfonso
Aprile, Francesco A
Dhulesia, Anne
Dobson, Christopher M
Vendruscolo, Michele
author_facet De Simone, Alfonso
Aprile, Francesco A
Dhulesia, Anne
Dobson, Christopher M
Vendruscolo, Michele
author_sort De Simone, Alfonso
collection PubMed
description Enzymes can increase the rate of biomolecular reactions by several orders of magnitude. Although the steps of substrate capture and product release are essential in the enzymatic process, complete atomic-level descriptions of these steps are difficult to obtain because of the transient nature of the intermediate conformations, which makes them largely inaccessible to standard structure determination methods. We describe here the determination of the structure of a low-population intermediate in the product release process by human lysozyme through a combination of NMR spectroscopy and molecular dynamics simulations. We validate this structure by rationally designing two mutations, the first engineered to destabilise the intermediate and the second to stabilise it, thus slowing down or speeding up, respectively, product release. These results illustrate how product release by an enzyme can be facilitated by the presence of a metastable intermediate with transient weak interactions between the enzyme and product. DOI: http://dx.doi.org/10.7554/eLife.02777.001
format Online
Article
Text
id pubmed-4383205
institution National Center for Biotechnology Information
language English
publishDate 2015
publisher eLife Sciences Publications, Ltd
record_format MEDLINE/PubMed
spelling pubmed-43832052015-04-03 Structure of a low-population intermediate state in the release of an enzyme product De Simone, Alfonso Aprile, Francesco A Dhulesia, Anne Dobson, Christopher M Vendruscolo, Michele eLife Biophysics and Structural Biology Enzymes can increase the rate of biomolecular reactions by several orders of magnitude. Although the steps of substrate capture and product release are essential in the enzymatic process, complete atomic-level descriptions of these steps are difficult to obtain because of the transient nature of the intermediate conformations, which makes them largely inaccessible to standard structure determination methods. We describe here the determination of the structure of a low-population intermediate in the product release process by human lysozyme through a combination of NMR spectroscopy and molecular dynamics simulations. We validate this structure by rationally designing two mutations, the first engineered to destabilise the intermediate and the second to stabilise it, thus slowing down or speeding up, respectively, product release. These results illustrate how product release by an enzyme can be facilitated by the presence of a metastable intermediate with transient weak interactions between the enzyme and product. DOI: http://dx.doi.org/10.7554/eLife.02777.001 eLife Sciences Publications, Ltd 2015-01-09 /pmc/articles/PMC4383205/ /pubmed/25575179 http://dx.doi.org/10.7554/eLife.02777 Text en © 2014, De Simone et al http://creativecommons.org/licenses/by/4.0/ This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited.
spellingShingle Biophysics and Structural Biology
De Simone, Alfonso
Aprile, Francesco A
Dhulesia, Anne
Dobson, Christopher M
Vendruscolo, Michele
Structure of a low-population intermediate state in the release of an enzyme product
title Structure of a low-population intermediate state in the release of an enzyme product
title_full Structure of a low-population intermediate state in the release of an enzyme product
title_fullStr Structure of a low-population intermediate state in the release of an enzyme product
title_full_unstemmed Structure of a low-population intermediate state in the release of an enzyme product
title_short Structure of a low-population intermediate state in the release of an enzyme product
title_sort structure of a low-population intermediate state in the release of an enzyme product
topic Biophysics and Structural Biology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4383205/
https://www.ncbi.nlm.nih.gov/pubmed/25575179
http://dx.doi.org/10.7554/eLife.02777
work_keys_str_mv AT desimonealfonso structureofalowpopulationintermediatestateinthereleaseofanenzymeproduct
AT aprilefrancescoa structureofalowpopulationintermediatestateinthereleaseofanenzymeproduct
AT dhulesiaanne structureofalowpopulationintermediatestateinthereleaseofanenzymeproduct
AT dobsonchristopherm structureofalowpopulationintermediatestateinthereleaseofanenzymeproduct
AT vendruscolomichele structureofalowpopulationintermediatestateinthereleaseofanenzymeproduct