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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fttibhannoverren:oai:oa.tib.eu:123456789/8646 2024-09-15T17:42:17+00:00 Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium-Part 2 Höck, Heidi Engel, Stefan Weingarten, Simone Keul, Helmut Schwaneberg, Ulrich Möller, Martin Bocola, Marco 2018 application/pdf https://oa.tib.eu/renate/handle/123456789/8646 https://doi.org/10.34657/7684 eng eng Basel : MDPI ESSN:2073-4360 DOI:https://doi.org/10.3390/polym10050524 https://oa.tib.eu/renate/handle/123456789/8646 https://doi.org/10.34657/7684 CC BY 4.0 Unported https://creativecommons.org/licenses/by/4.0/ frei zugänglich ddc:540 Candida antarctica lipase B Enzymatic ring-opening polymerization Enzyme engineering Immobilization Microgel status-type:publishedVersion doc-type:Article doc-type:Text 2018 fttibhannoverren https://doi.org/10.34657/768410.3390/polym10050524 2024-07-03T23:33:53Z 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. Article in Journal/Newspaper Antarc* Antarctica Renate - Repositorium für Naturwissenschaften und Technik (TIB Hannover) |
institution |
Open Polar |
collection |
Renate - Repositorium für Naturwissenschaften und Technik (TIB Hannover) |
op_collection_id |
fttibhannoverren |
language |
English |
topic |
ddc:540 Candida antarctica lipase B Enzymatic ring-opening polymerization Enzyme engineering Immobilization Microgel |
spellingShingle |
ddc:540 Candida antarctica lipase B Enzymatic ring-opening polymerization Enzyme engineering Immobilization Microgel Höck, Heidi Engel, Stefan Weingarten, Simone Keul, Helmut Schwaneberg, Ulrich Möller, Martin Bocola, Marco Comparison of Candida antarctica Lipase B Variants for Conversion of ε-Caprolactone in Aqueous Medium-Part 2 |
topic_facet |
ddc:540 Candida antarctica lipase B Enzymatic ring-opening polymerization Enzyme engineering Immobilization Microgel |
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 |
Article in Journal/Newspaper |
author |
Höck, Heidi Engel, Stefan Weingarten, Simone Keul, Helmut Schwaneberg, Ulrich Möller, Martin Bocola, Marco |
author_facet |
Höck, Heidi Engel, Stefan Weingarten, Simone Keul, Helmut Schwaneberg, Ulrich Möller, Martin Bocola, Marco |
author_sort |
Höck, Heidi |
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 |
Basel : MDPI |
publishDate |
2018 |
url |
https://oa.tib.eu/renate/handle/123456789/8646 https://doi.org/10.34657/7684 |
genre |
Antarc* Antarctica |
genre_facet |
Antarc* Antarctica |
op_relation |
ESSN:2073-4360 DOI:https://doi.org/10.3390/polym10050524 https://oa.tib.eu/renate/handle/123456789/8646 https://doi.org/10.34657/7684 |
op_rights |
CC BY 4.0 Unported https://creativecommons.org/licenses/by/4.0/ frei zugänglich |
op_doi |
https://doi.org/10.34657/768410.3390/polym10050524 |
_version_ |
1810488799156764672 |