Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions
Abstract Michael‐type additions of various thiols and α,β‐unsaturated carbonyl compounds were performed in organic solvent catalyzed by wild‐type and a rationally redesigned mutant of Candida antarctica lipase B. The mutant lacks the nucleophilic serine 105 in the active‐site; this results in a chan...
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crwiley:10.1002/cbic.200400213 2024-06-23T07:47:40+00:00 Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions Carlqvist, Peter Svedendahl, Maria Branneby, Cecilia Hult, Karl Brinck, Tore Berglund, Per 2005 http://dx.doi.org/10.1002/cbic.200400213 https://api.wiley.com/onlinelibrary/tdm/v1/articles/10.1002%2Fcbic.200400213 https://onlinelibrary.wiley.com/doi/full/10.1002/cbic.200400213 en eng Wiley http://onlinelibrary.wiley.com/termsAndConditions#vor ChemBioChem volume 6, issue 2, page 331-336 ISSN 1439-4227 1439-7633 journal-article 2005 crwiley https://doi.org/10.1002/cbic.200400213 2024-06-11T04:45:08Z Abstract Michael‐type additions of various thiols and α,β‐unsaturated carbonyl compounds were performed in organic solvent catalyzed by wild‐type and a rationally redesigned mutant of Candida antarctica lipase B. The mutant lacks the nucleophilic serine 105 in the active‐site; this results in a changed catalytic mechanism of the enzyme. The possibility of utilizing this mutant for Michael‐type additions was initially explored by quantum‐chemical calculations on the reaction between acrolein and methanethiol in a model system. The model system was constructed on the basis of docking and molecular‐dynamics simulations and was designed to simulate the catalytic properties of the active site. The catalytic system was explored experimentally with a range of different substrates. The k cat values were found to be in the range of 10 −3 to 4 min −1 , similar to the values obtained with aldolase antibodies. The enzyme proficiency was 10 7 . Furthermore, the Michael‐type reactions followed saturation kinetics and were confirmed to take place in the enzyme active site. Article in Journal/Newspaper Antarc* Antarctica Wiley Online Library ChemBioChem 6 2 331 336 |
institution |
Open Polar |
collection |
Wiley Online Library |
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crwiley |
language |
English |
description |
Abstract Michael‐type additions of various thiols and α,β‐unsaturated carbonyl compounds were performed in organic solvent catalyzed by wild‐type and a rationally redesigned mutant of Candida antarctica lipase B. The mutant lacks the nucleophilic serine 105 in the active‐site; this results in a changed catalytic mechanism of the enzyme. The possibility of utilizing this mutant for Michael‐type additions was initially explored by quantum‐chemical calculations on the reaction between acrolein and methanethiol in a model system. The model system was constructed on the basis of docking and molecular‐dynamics simulations and was designed to simulate the catalytic properties of the active site. The catalytic system was explored experimentally with a range of different substrates. The k cat values were found to be in the range of 10 −3 to 4 min −1 , similar to the values obtained with aldolase antibodies. The enzyme proficiency was 10 7 . Furthermore, the Michael‐type reactions followed saturation kinetics and were confirmed to take place in the enzyme active site. |
format |
Article in Journal/Newspaper |
author |
Carlqvist, Peter Svedendahl, Maria Branneby, Cecilia Hult, Karl Brinck, Tore Berglund, Per |
spellingShingle |
Carlqvist, Peter Svedendahl, Maria Branneby, Cecilia Hult, Karl Brinck, Tore Berglund, Per Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions |
author_facet |
Carlqvist, Peter Svedendahl, Maria Branneby, Cecilia Hult, Karl Brinck, Tore Berglund, Per |
author_sort |
Carlqvist, Peter |
title |
Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions |
title_short |
Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions |
title_full |
Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions |
title_fullStr |
Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions |
title_full_unstemmed |
Exploring the Active‐Site of a Rationally Redesigned Lipase for Catalysis of Michael‐Type Additions |
title_sort |
exploring the active‐site of a rationally redesigned lipase for catalysis of michael‐type additions |
publisher |
Wiley |
publishDate |
2005 |
url |
http://dx.doi.org/10.1002/cbic.200400213 https://api.wiley.com/onlinelibrary/tdm/v1/articles/10.1002%2Fcbic.200400213 https://onlinelibrary.wiley.com/doi/full/10.1002/cbic.200400213 |
genre |
Antarc* Antarctica |
genre_facet |
Antarc* Antarctica |
op_source |
ChemBioChem volume 6, issue 2, page 331-336 ISSN 1439-4227 1439-7633 |
op_rights |
http://onlinelibrary.wiley.com/termsAndConditions#vor |
op_doi |
https://doi.org/10.1002/cbic.200400213 |
container_title |
ChemBioChem |
container_volume |
6 |
container_issue |
2 |
container_start_page |
331 |
op_container_end_page |
336 |
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1802651809790033920 |