Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2
Enzyme-catalyzed ring-opening polymerization of lactones is a method of increasing interest for the synthesis of polyesters. In the present work, we investigated which changes in the structure of Candida antarctica lipase B (CaLB) shift the catalytic equilibrium between esterification and hydrolysis...
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ftmdpi:oai:mdpi.com:/2073-4360/10/5/524/ 2023-08-20T04:02:01+02:00 Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 Heidi Höck Stefan Engel Simone Weingarten Helmut Keul Ulrich Schwaneberg Martin Möller Marco Bocola 2018-05-14 application/pdf https://doi.org/10.3390/polym10050524 EN eng Multidisciplinary Digital Publishing Institute https://dx.doi.org/10.3390/polym10050524 https://creativecommons.org/licenses/by/4.0/ Polymers; Volume 10; Issue 5; Pages: 524 Candida antarctica lipase B enzymatic ring-opening polymerization enzyme engineering microgel immobilization Text 2018 ftmdpi https://doi.org/10.3390/polym10050524 2023-07-31T21:31:13Z Enzyme-catalyzed ring-opening polymerization of lactones is a method of increasing interest for the synthesis of polyesters. In the present work, we investigated which changes in the structure of Candida antarctica lipase B (CaLB) shift the catalytic equilibrium between esterification and hydrolysis towards polymerization. Therefore, we present two concepts: (i) removing the glycosylation of CaLB to increase the surface hydrophobicity; and (ii) introducing a hydrophobic lid adapted from Pseudomonas cepacia lipase (PsCL) to enhance the interaction of a growing polymer chain to the elongated lid helix. The deglycosylated CaLB (CaLB-degl) was successfully generated by site-saturation mutagenesis of asparagine 74. Furthermore, computational modeling showed that the introduction of a lid helix at position Ala148 was structurally feasible and the geometry of the active site remained intact. Via overlap extension PCR the lid was successfully inserted, and the variant was produced in large scale in Pichia pastoris with glycosylation (CaLB-lid) and without (CaLB-degl-lid). While the lid variants show a minor positive effect on the polymerization activity, CaLB-degl showed a clearly reduced hydrolytic and enhanced polymerization activity. Immobilization in a hydrophobic polyglycidol-based microgel intensified this effect such that a higher polymerization activity was achieved, compared to the “gold standard” Novozym® 435. Text Antarc* Antarctica MDPI Open Access Publishing Polymers 10 5 524 |
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Open Polar |
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MDPI Open Access Publishing |
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English |
topic |
Candida antarctica lipase B enzymatic ring-opening polymerization enzyme engineering microgel immobilization |
spellingShingle |
Candida antarctica lipase B enzymatic ring-opening polymerization enzyme engineering microgel immobilization Heidi Höck Stefan Engel Simone Weingarten Helmut Keul Ulrich Schwaneberg Martin Möller Marco Bocola Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 |
topic_facet |
Candida antarctica lipase B enzymatic ring-opening polymerization enzyme engineering microgel immobilization |
description |
Enzyme-catalyzed ring-opening polymerization of lactones is a method of increasing interest for the synthesis of polyesters. In the present work, we investigated which changes in the structure of Candida antarctica lipase B (CaLB) shift the catalytic equilibrium between esterification and hydrolysis towards polymerization. Therefore, we present two concepts: (i) removing the glycosylation of CaLB to increase the surface hydrophobicity; and (ii) introducing a hydrophobic lid adapted from Pseudomonas cepacia lipase (PsCL) to enhance the interaction of a growing polymer chain to the elongated lid helix. The deglycosylated CaLB (CaLB-degl) was successfully generated by site-saturation mutagenesis of asparagine 74. Furthermore, computational modeling showed that the introduction of a lid helix at position Ala148 was structurally feasible and the geometry of the active site remained intact. Via overlap extension PCR the lid was successfully inserted, and the variant was produced in large scale in Pichia pastoris with glycosylation (CaLB-lid) and without (CaLB-degl-lid). While the lid variants show a minor positive effect on the polymerization activity, CaLB-degl showed a clearly reduced hydrolytic and enhanced polymerization activity. Immobilization in a hydrophobic polyglycidol-based microgel intensified this effect such that a higher polymerization activity was achieved, compared to the “gold standard” Novozym® 435. |
format |
Text |
author |
Heidi Höck Stefan Engel Simone Weingarten Helmut Keul Ulrich Schwaneberg Martin Möller Marco Bocola |
author_facet |
Heidi Höck Stefan Engel Simone Weingarten Helmut Keul Ulrich Schwaneberg Martin Möller Marco Bocola |
author_sort |
Heidi Höck |
title |
Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 |
title_short |
Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 |
title_full |
Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 |
title_fullStr |
Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 |
title_full_unstemmed |
Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium—Part 2 |
title_sort |
comparison of candida antarctica lipase b variants for conversion of ε-caprolactone in aqueous medium—part 2 |
publisher |
Multidisciplinary Digital Publishing Institute |
publishDate |
2018 |
url |
https://doi.org/10.3390/polym10050524 |
genre |
Antarc* Antarctica |
genre_facet |
Antarc* Antarctica |
op_source |
Polymers; Volume 10; Issue 5; Pages: 524 |
op_relation |
https://dx.doi.org/10.3390/polym10050524 |
op_rights |
https://creativecommons.org/licenses/by/4.0/ |
op_doi |
https://doi.org/10.3390/polym10050524 |
container_title |
Polymers |
container_volume |
10 |
container_issue |
5 |
container_start_page |
524 |
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1774712409766756352 |