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Heterogeneous catalysis in racemization and kinetic resolution along a journey in protein engineering
Stockholm University, Faculty of Science, Department of Organic Chemistry.
2018 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

The first part of my thesis concerns the use of heterogeneous acidic resins for racemization of tert-alcohols without any side-product formation. The focus was to develop a system which can be further extended to a DKR protocol consisting of an enzymatic KR reaction. Based on our knowledge of the resins, an unexpected migratory DKR protocol turned out to be an efficient method for the synthesis of carbocyclic allylic carbinols.

The development of enzyme and metal catalyst hybrids was already an ongoing theme in our group. A supporter-free biohybrid catalyst was developed which can be used in several different types of reactions. The Pd(0)-CalB CLEA catalyst was applied in a two-step-cascade transformation and in the DKR of benzylic primary amines. The catalyst was characterized by different analytical techniques, to understand its composition and structure.

The enzymes have always been the main focus of the studies and therefore wild type enzymes were initially utilized. However, these natural biocatalysts are associated with certain limitations. In contrast, protein engineering allows for enzymes to be modified and optimized. We have used the technique to create a subtilisin Carlsberg mutant, which was studied both by modeling and in vitro. The mutant was found to catalyze the (S)-selective transesterification of sec-alcohols containing long aliphatic carbon chains, and it also exhibited higher performance in organic solvent.

The last project concerned the protein engineering of CalA enzyme towards tert-alcohols. The kinetic resolution of tert-alcohols with this enzyme is very slow but it occurs with good enantioselectivity. The aim was therefore to improve the activity of CalA via protein engineering. Seven amino acids were mutated close to the active site and a library was created based on our prediction. Throughout the screening, a few variants showed higher activity, which were sequenced and further analyzed in the transesterification of tert-alcohols.

Place, publisher, year, edition, pages
Stockholm: Department of Organic Chemistry, Stockholm University , 2018. , p. 81
Keyword [en]
racemization, protein engineering, kinetic resolution, heterogeneous catalysis, biohybrid catalyst
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
URN: urn:nbn:se:su:diva-153921ISBN: 978-91-7797-185-6 (print)ISBN: 978-91-7797-186-3 (electronic)OAI: oai:DiVA.org:su-153921DiVA, id: diva2:1188639
Public defence
2018-04-26, Magnélisalen, Kemiska övningslaboratoriet, Svante Arrhenius väg 16 B, Stockholm, 10:00 (English)
Opponent
Supervisors
Note

At the time of the doctoral defense, the following papers were unpublished and had a status as follows: Paper 4: Manuscript. Paper 6: Manuscript.

Available from: 2018-04-03 Created: 2018-03-08 Last updated: 2018-03-23Bibliographically approved
List of papers
1. Heterogeneous Acid-Catalyzed Racemization of Tertiary Alcohols
Open this publication in new window or tab >>Heterogeneous Acid-Catalyzed Racemization of Tertiary Alcohols
2018 (English)In: Chemistry - A European Journal, ISSN 0947-6539, E-ISSN 1521-3765, Vol. 24, no 1, p. 77-80Article in journal (Refereed) Published
Abstract [en]

Tertiary alcohols are important structural motifs in natural products and building blocks in organic synthesis but only few methods are known for their enantioselective preparation. Chiral resolution is one of these approaches that leaves one enantiomer (50% of the material) unaffected. An attractive method to increase the efficiency of those resolutions is to racemize the unaffected enantiomer. In the present work, we have developed a practical racemization protocol for tertiary alcohols. Five different acidic resin materials were tested. The Dowex 50WX8 was the resin of choice since it was capable of racemizing tertiary alcohols without any byproduct formation. Suitable solvents and a biphasic system were investigated, and the optimized system was capable of racemizing differently substituted tertiary alcohols.

Keyword
acidic resins, heterogeneous catalysis, racemization, tertiary alcohols
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-152563 (URN)10.1002/chem.201704691 (DOI)000419046600010 ()29131426 (PubMedID)
Available from: 2018-02-19 Created: 2018-02-19 Last updated: 2018-03-08Bibliographically approved
2. Migratory Dynamic Kinetic Resolution of Carbocyclic Allylic Alcohols
Open this publication in new window or tab >>Migratory Dynamic Kinetic Resolution of Carbocyclic Allylic Alcohols
Show others...
2014 (English)In: Organic Letters, ISSN 1523-7060, E-ISSN 1523-7052, Vol. 16, no 22, p. 5952-5955Article in journal (Refereed) Published
Abstract [en]

A novel migratory dynamic kinetic resolution based on the interplay between an enzyme acylation catalyst and a heterogeneous Bronsted acid as an isomerization/racemization catalyst gives rise to carbocyclic allylic esters with excellent stereoselectivity from readily available tertiary carbinols. An easy-to-use teabag setup combining resin-bound catalysts, a biphasic isooctanewater solvent system, and a highly lipophilic acyl donor efficiently suppresses side reactions and allows for the preparation of functionalized carbocyclic building blocks in high yields and optical purity.

National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-121784 (URN)10.1021/ol502979g (DOI)000345470000033 ()
Funder
German Research Foundation (DFG), DE1599/4-1Swedish Research CouncilKnut and Alice Wallenberg Foundation
Available from: 2015-10-16 Created: 2015-10-16 Last updated: 2018-03-08Bibliographically approved
3. Design of a Pd(0)-CalB CLEA Biohybrid Catalyst and Its Application in a One-Pot Cascade Reaction
Open this publication in new window or tab >>Design of a Pd(0)-CalB CLEA Biohybrid Catalyst and Its Application in a One-Pot Cascade Reaction
Show others...
2017 (English)In: ACS Catalysis, ISSN 2155-5435, E-ISSN 2155-5435, Vol. 7, no 3, p. 1601-1605Article in journal (Refereed) Published
Abstract [en]

Herein, a design of a biohybrid catalyst is described, consisting of Pd nanoparticles and a cross-linked network of aggregated lipase B enzyme of Candida antarctica (CalB CLEA) functioning as an active support for the Pd nanoparticles. Both entities of the hybrid catalyst showed good catalytic activity. The applicability was demonstrated in a one-pot reaction, where the Pd-catalyzed cycloisomerization of 4-pentynoic acid afforded a lactone that serves as an acyl donor in a subsequent selective enzymatic kinetic resolution of a set of sec-alcohols. The catalyst proved to be robust and could be recycled five times without a significant loss of activity.

Keyword
kinetic resolution, biohybrid catalyst, Pd nanoparticles, CalB, cycloisomerization, one-pot cascade reaction
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-142484 (URN)10.1021/acscatal.6b03481 (DOI)000395726500011 ()
Available from: 2017-05-16 Created: 2017-05-16 Last updated: 2018-03-09Bibliographically approved
4. Application and further structure elucidation of Pd(0)-CalB CLEA biohybrid catalyst- Chemoenzymatic dynamic kinetic resolution of primary benzylic amines
Open this publication in new window or tab >>Application and further structure elucidation of Pd(0)-CalB CLEA biohybrid catalyst- Chemoenzymatic dynamic kinetic resolution of primary benzylic amines
Show others...
(English)Manuscript (preprint) (Other academic)
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-153897 (URN)
Available from: 2018-03-07 Created: 2018-03-07 Last updated: 2018-03-20Bibliographically approved
5. Improved Enantioselectivity of Subtilisin Carlsberg Towards Secondary Alcohols by Protein Engineering
Open this publication in new window or tab >>Improved Enantioselectivity of Subtilisin Carlsberg Towards Secondary Alcohols by Protein Engineering
2018 (English)In: ChemBioChem (Print), ISSN 1439-4227, E-ISSN 1439-7633, Vol. 19, no 4, p. 338-346Article in journal (Refereed) Published
Abstract [en]

Generally, the catalytic activity of subtilisin Carlsberg (SC) for transacylation reactions with secondary alcohols in organic solvent is low. Enzyme immobilization and protein engineering was performed to improve the enantioselectivity of SC towards secondary alcohols. Possible amino-acid residues for mutagenesis were found by combining available literature data with molecular modeling. SC variants were created by site-directed mutagenesis and were evaluated for a model transacylation reaction containing 1-phenylethanol in THF. Variants showing high E values (>100) were found. However, the conversions were still low. A second mutation was made, and both the E values and conversions were increased. Relative to that shown by the wild type, the most successful variant, G165L/M221F, showed increased conversion (up to 36 %), enantioselectivity (E values up to 400), substrate scope, and stability in THF.

Keyword
biocatalysis, enzymes, immobilization, kinetic resolution, molecular modeling, transacylation
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-153920 (URN)10.1002/cbic.201700408 (DOI)000425511500006 ()
Available from: 2018-03-07 Created: 2018-03-07 Last updated: 2018-03-20Bibliographically approved
6. Transesterification of tert-Alcohols by Engineered Candida antarctica Lipase A
Open this publication in new window or tab >>Transesterification of tert-Alcohols by Engineered Candida antarctica Lipase A
Show others...
(English)Manuscript (preprint) (Other academic)
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:su:diva-153918 (URN)
Available from: 2018-03-07 Created: 2018-03-07 Last updated: 2018-03-20Bibliographically approved

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