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Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191

The arylacetonitrilase from the bacterium Pseudomonas fluorescens EBC191 has been intensively studied as a model to understand the molecular basis for the substrate-, reaction-, and enantioselectivity of nitrilases. The nitrilase converts various aromatic and aliphatic nitriles to the corresponding...

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Autores principales: Stolz, Andreas, Eppinger, Erik, Sosedov, Olga, Kiziak, Christoph
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2019
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6930498/
https://www.ncbi.nlm.nih.gov/pubmed/31766372
http://dx.doi.org/10.3390/molecules24234232
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author Stolz, Andreas
Eppinger, Erik
Sosedov, Olga
Kiziak, Christoph
author_facet Stolz, Andreas
Eppinger, Erik
Sosedov, Olga
Kiziak, Christoph
author_sort Stolz, Andreas
collection PubMed
description The arylacetonitrilase from the bacterium Pseudomonas fluorescens EBC191 has been intensively studied as a model to understand the molecular basis for the substrate-, reaction-, and enantioselectivity of nitrilases. The nitrilase converts various aromatic and aliphatic nitriles to the corresponding acids and varying amounts of the corresponding amides. The enzyme has been analysed by site-specific mutagenesis and more than 50 different variants have been generated and analysed for the conversion of (R,S)-mandelonitrile and (R,S)-2-phenylpropionitrile. These comparative analyses demonstrated that single point mutations are sufficient to generate enzyme variants which hydrolyse (R,S)-mandelonitrile to (R)-mandelic acid with an enantiomeric excess (ee) of 91% or to (S)-mandelic acid with an ee-value of 47%. The conversion of (R,S)-2-phenylpropionitrile by different nitrilase variants resulted in the formation of either (S)- or (R)-2-phenylpropionic acid with ee-values up to about 80%. Furthermore, the amounts of amides that are produced from (R,S)-mandelonitrile and (R,S)-2-phenylpropionitrile could be changed by single point mutations between 2%–94% and <0.2%–73%, respectively. The present study attempted to collect and compare the results obtained during our previous work, and to obtain additional general information about the relationship of the amide forming capacity of nitrilases and the enantiomeric composition of the products.
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spelling pubmed-69304982019-12-26 Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191 Stolz, Andreas Eppinger, Erik Sosedov, Olga Kiziak, Christoph Molecules Review The arylacetonitrilase from the bacterium Pseudomonas fluorescens EBC191 has been intensively studied as a model to understand the molecular basis for the substrate-, reaction-, and enantioselectivity of nitrilases. The nitrilase converts various aromatic and aliphatic nitriles to the corresponding acids and varying amounts of the corresponding amides. The enzyme has been analysed by site-specific mutagenesis and more than 50 different variants have been generated and analysed for the conversion of (R,S)-mandelonitrile and (R,S)-2-phenylpropionitrile. These comparative analyses demonstrated that single point mutations are sufficient to generate enzyme variants which hydrolyse (R,S)-mandelonitrile to (R)-mandelic acid with an enantiomeric excess (ee) of 91% or to (S)-mandelic acid with an ee-value of 47%. The conversion of (R,S)-2-phenylpropionitrile by different nitrilase variants resulted in the formation of either (S)- or (R)-2-phenylpropionic acid with ee-values up to about 80%. Furthermore, the amounts of amides that are produced from (R,S)-mandelonitrile and (R,S)-2-phenylpropionitrile could be changed by single point mutations between 2%–94% and <0.2%–73%, respectively. The present study attempted to collect and compare the results obtained during our previous work, and to obtain additional general information about the relationship of the amide forming capacity of nitrilases and the enantiomeric composition of the products. MDPI 2019-11-21 /pmc/articles/PMC6930498/ /pubmed/31766372 http://dx.doi.org/10.3390/molecules24234232 Text en © 2019 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 Review
Stolz, Andreas
Eppinger, Erik
Sosedov, Olga
Kiziak, Christoph
Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191
title Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191
title_full Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191
title_fullStr Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191
title_full_unstemmed Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191
title_short Comparative Analysis of the Conversion of Mandelonitrile and 2-Phenylpropionitrile by a Large Set of Variants Generated from a Nitrilase Originating from Pseudomonas fluorescens EBC191
title_sort comparative analysis of the conversion of mandelonitrile and 2-phenylpropionitrile by a large set of variants generated from a nitrilase originating from pseudomonas fluorescens ebc191
topic Review
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6930498/
https://www.ncbi.nlm.nih.gov/pubmed/31766372
http://dx.doi.org/10.3390/molecules24234232
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