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Lipase chemoselectivity towards alcohol and thiol acyl acceptors in a transacylation reaction
KTH, School of Biotechnology (BIO), Biochemistry.
KTH, School of Biotechnology (BIO), Biochemistry.
KTH, School of Biotechnology (BIO), Biochemistry.ORCID iD: 0000-0002-2993-9375
2010 (English)In: Journal of Molecular Catalysis B: Enzymatic, ISSN 1381-1177, E-ISSN 1873-3158, Vol. 66, no 1-2, 120-123 p.Article in journal (Refereed) Published
Abstract [en]

The lipase chemoselectivity towards an alcohol and a thiol was investigated for the two lipases Candida antarctica lipase B (CalB) and Rhizomucor miehei lipase (Rml). Hexanol and hexanethiol were used as acyl acceptors in a transacylation reaction with ethyl octanoate in cyclohexane. CalB showed the highest chemoselectivity ratio (k(cat)/K-M)(OH)/(k(cat)/K-M)(SH), of 88,000 while the ratio for Rml was 1200. That could be compared with the ratio, k(OH)/k(SH), of 120 for the non-catalyzed reaction. Thus, the enzyme contribution to the chemoselectivity between hexanol and hexanethiol was 730 for CalB and 10 for Rml. High K-M values displayed towards hexanethiol (above 1.8 M) were the largest contribution to the selectivity. No saturation was achieved. The K-M values were more than two orders of magnitude higher than those of hexanol.

Place, publisher, year, edition, pages
2010. Vol. 66, no 1-2, 120-123 p.
Keyword [en]
Chemoselectiviry, Acylation, Lipase, Kinetics, Active site titration
National Category
Biochemistry and Molecular Biology
Identifiers
URN: urn:nbn:se:kth:diva-25232DOI: 10.1016/j.molcatb.2010.04.005ISI: 000280928200015Scopus ID: 2-s2.0-77957827548OAI: oai:DiVA.org:kth-25232DiVA: diva2:356637
Note
QC 20101013Available from: 2010-10-13 Created: 2010-10-13 Last updated: 2017-12-12Bibliographically approved
In thesis
1. Lipase chemoselectivity - kinetics and applications
Open this publication in new window or tab >>Lipase chemoselectivity - kinetics and applications
2009 (English)Licentiate thesis, comprehensive summary (Other academic)
Abstract [en]

 

A chemoselective catalyst is preferred in a chemical reaction where protecting groups otherwise are needed. The two lipases Candida antarctica lipase B and Rhizomucor miehei lipase showed large chemoselectivity ratios, defined as (kcat/KM)OH / (kcat/KM)SH, in a transacylation reaction with ethyl octanoate as acyl donor and hexanol or hexanethiol as acyl acceptor (paper I). The chemoselectivity ratio of the uncatalyzed reaction was 120 in favour of the alcohol. Compared to the uncatalyzed reaction, the chemoselectivity was 730 times higher for Candida antarctica lipase B and ten times higher for Rhizomucor miehei lipase. The KM towards the thiol was more than two orders of magnitude higher than the KM towards the corresponding alcohol. This was the dominating contribution to the high chemoselectivity displayed by the two lipases. In a novel approach, Candida antarctica lipase B was used as catalyst for enzymatic synthesis of thiol-functionalized polyesters in a one-pot reaction without using protecting groups (paper II). Poly(e-caprolactone) with a free thiol at one of the ends was synthesized in an enzymatic ring-opening polymerization initiated with mercaptoethanol or terminated with either 3-mercaptopropionic acid or g-thiobutyrolactone.

 

Place, publisher, year, edition, pages
Stockholm: KTH, 2009. 36 p.
Series
Trita-BIO-Report, ISSN 1654-2312 ; 2009:7
Keyword
Candida antarctica lipase B, Rhizomucor miehei lipase, active site titration, kinetics, chemoselectivity, thiol, alcohol, thiol-functionalized polyesters
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:kth:diva-10232 (URN)978-91-7415-275-3 (ISBN)
Presentation
2009-05-14, 10:00 (English)
Supervisors
Available from: 2009-05-06 Created: 2009-04-20 Last updated: 2010-10-13Bibliographically approved
2. Lipase selectivity in functional polyester synthesis
Open this publication in new window or tab >>Lipase selectivity in functional polyester synthesis
2011 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

Enzyme selectivity means that the enzyme´s preferences towards competing substrates will be different. In this thesis, the enzyme selectivity has been studied for utilization in synthesis of functionalized macromonomers. The aim was to study how the inherent –or introduced – selectivity of lipases can be used to introduce thiol‐ or enefunctionalities into short polyesters. Thiol‐ and ene‐functionalized renewable organic precursor molecules in combination with thiol‐ene click chemistry opens up for a sustainable material production. Lipases do not normally affect ene‐moieties and the preference towards thiols is low, enabling introduction of these functional groups for further modifications. In addition, lipases have been shown to be good catalysts in the formation of polyesters, both via ring‐opening and polycondensation polymerization.

In paper I Candida antarctica lipase B was used to end‐functionalize poly(ε‐caprolactone) with free thiols in a one‐pot reaction. The advantage of using achemoselective lipase as catalyst was that no protection of the thiol was needed. The chemoselectivity displayed by Candida antarctica lipase B turned out to be 88 000 in favour of the alcohol (paper II). Rhizomucor miehei lipase showed less pronounced chemoselectivity. The largest contribution to the selectivities was derived from the more than two orders of magnitude higer KM towards the thiol compared to the alcohol. Thiols can be cross‐linked with enes in radical reactions to form networks, enabling formation of materials.

One promising renewable molecule containing an acrylate moiety is itaconic acid. In paper III the selectivity towards the two esters in dimethyl itaconate was investigated and the active site of Candida antarctica lipase B was redesigned to generate variants with increased and decreased selectivity. One variant showed 14‐fold higher selectivity and could regioselectively add dimethyl itaconate onto a diol. This variant could be used in end‐functionalizations of polymers, introducing acrylate‐ester end‐groups.

The enzyme selectivity towards lactones and their corresponding polyesters is of importance when designing a ring‐opening polymerization reaction. In paper IV Candida antarctica lipase B was found to prefer ω‐pentadecalactone and polyesters over ε‐caprolactone ten‐fold, while Humicola insolens cutinase preferred ε‐caprolactone and its corresponding polyester four‐fold over ω‐pentadecalactone and its polyester. From a selectivity point of view, Candida antarctica lipase B and Humicola insolens cutinase would be equally good in ring‐opening polymerization of ω‐pentadecalactone, while in the case of ε‐caprolactone Humicola insolens cutinase would be the preferred choice.

Place, publisher, year, edition, pages
Stockholm: KTH Royal Institute of Technology, 2011. 49 p.
Series
Trita-BIO-Report, ISSN 1654-2312 ; 2011:20
National Category
Biochemistry and Molecular Biology
Identifiers
urn:nbn:se:kth:diva-34023 (URN)978-91-7501-007-6 (ISBN)
Public defence
2011-06-17, FD5, AlbaNova University Center, Roslagstullsbacken 21, Stockholm, 10:00 (English)
Opponent
Supervisors
Note
QC 20110608Available from: 2011-06-08 Created: 2011-05-23 Last updated: 2011-06-08Bibliographically approved

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