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Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.

Iman Omar Orcid Logo, Michele Crotti Orcid Logo, Chuhan Li Orcid Logo, Krisztina Pisak, Blazej Czemerys, Salvatore Ferla Orcid Logo, Aster van Noord, Caroline E. Paul Orcid Logo, Kersti Karu Orcid Logo, Cagakan Ozbalci Orcid Logo, Ulrike Eggert Orcid Logo, Richard Lloyd, Sarah M. Barry Orcid Logo, Daniele Castagnolo Orcid Logo

Angewandte Chemie International Edition

Swansea University Author: Salvatore Ferla Orcid Logo

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DOI (Published version): 10.1002/anie.202403493

Abstract

Cyclopropane fatty acid synthases (CFAS) are a class of S-adenosylmethionine (SAM) dependent methyltransferase enzymes able to catalyse the cyclopropanation of unsaturated phospholipids. Since CFAS enzymes employ SAM as a methylene source to cyclopropanate alkene substrates, they have the potential...

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Published in: Angewandte Chemie International Edition
ISSN: 1433-7851 1521-3773
Published: Wiley 2024
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URI: https://cronfa.swan.ac.uk/Record/cronfa66386
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spelling v2 66386 2024-05-09 Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation. d4c62248f510e3b221916989a7bbe6a6 0000-0002-5918-9237 Salvatore Ferla Salvatore Ferla true false 2024-05-09 MEDS Cyclopropane fatty acid synthases (CFAS) are a class of S-adenosylmethionine (SAM) dependent methyltransferase enzymes able to catalyse the cyclopropanation of unsaturated phospholipids. Since CFAS enzymes employ SAM as a methylene source to cyclopropanate alkene substrates, they have the potential to be mild and more sustainable biocatalysts for cyclopropanation transformations than current carbene based approaches. This work describes the characterisation of E. coli CFAS enzyme (ecCFAS) and its exploitation in the stereoselective biocatalytic synthesis of cyclopropyl lipids. ecCFAS was found to convert phosphatidylglycerol (PG) to methyl dihydrosterculate 1 from in up to 58% conversion and 73% ee and the absolute configuration (9S,10R) was established. Substrate tolerance of ecCFAS was found to be correlated with the electronic properties of phospholipid headgroups and for the first time ecCFAS was found to catalyse cyclopropanation of both phospholipid chains to form dicyclopropanated products. In addition, mutagenesis and in-silico experiments were carried out to identify the enzyme residues with key roles in catalysis and to provide structural insights into the lipid substrate preference of ecCFAS. Finally, the biocatalytic synthesis of methyl dihydrosterculate 1 and its deuterated analogue was also accomplished combining pure ecCFAS with the SAM regenerating AtHMT enzyme in presence of CH3I and CD3I. Journal Article Angewandte Chemie International Edition 0 Wiley 1433-7851 1521-3773 14 6 2024 2024-06-14 10.1002/anie.202403493 COLLEGE NANME Medical School COLLEGE CODE MEDS Swansea University Another institution paid the OA fee Biotechnology and Biological Sciences Research Council. Grant Numbers: BB/M009513/1, BIDS3000037082, POC-045, R116824, BB/P019811/1 Chinese Government Scholarship. Grant Number: 202108310078 H2020 Marie Skłodowska-Curie Actions. Grant Number: 101027045 H2020 European Research Council. Grant Number: 949910 2024-06-27T15:38:27.0000222 2024-05-09T15:23:09.4818876 Faculty of Medicine, Health and Life Sciences Swansea University Medical School - Pharmacy Iman Omar 0000-0002-4216-5394 1 Michele Crotti 0000-0003-3274-9393 2 Chuhan Li 0000-0001-5989-0093 3 Krisztina Pisak 4 Blazej Czemerys 5 Salvatore Ferla 0000-0002-5918-9237 6 Aster van Noord 7 Caroline E. Paul 0000-0002-7889-9920 8 Kersti Karu 0000-0002-3974-0271 9 Cagakan Ozbalci 0000-0002-7572-5511 10 Ulrike Eggert 0000-0003-0932-5525 11 Richard Lloyd 12 Sarah M. Barry 0000-0001-8188-2153 13 Daniele Castagnolo 0000-0002-7517-5732 14 66386__30772__01c5effe77b145cb901e73dec438b44e.pdf 66386.VoR.pdf 2024-06-27T15:36:24.3514976 Output 8912595 application/pdf Version of Record true © 2024 The Authors. This is an open access article under the terms of the Creative Commons Attribution License. true eng http://creativecommons.org/licenses/by/4.0/
title Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
spellingShingle Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
Salvatore Ferla
title_short Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
title_full Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
title_fullStr Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
title_full_unstemmed Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
title_sort Insights into E. coli Cyclopropane Fatty Acid Synthase (CFAS) Towards Enantioselective Carbene Free Biocatalytic Cyclopropanation.
author_id_str_mv d4c62248f510e3b221916989a7bbe6a6
author_id_fullname_str_mv d4c62248f510e3b221916989a7bbe6a6_***_Salvatore Ferla
author Salvatore Ferla
author2 Iman Omar
Michele Crotti
Chuhan Li
Krisztina Pisak
Blazej Czemerys
Salvatore Ferla
Aster van Noord
Caroline E. Paul
Kersti Karu
Cagakan Ozbalci
Ulrike Eggert
Richard Lloyd
Sarah M. Barry
Daniele Castagnolo
format Journal article
container_title Angewandte Chemie International Edition
container_volume 0
publishDate 2024
institution Swansea University
issn 1433-7851
1521-3773
doi_str_mv 10.1002/anie.202403493
publisher Wiley
college_str Faculty of Medicine, Health and Life Sciences
hierarchytype
hierarchy_top_id facultyofmedicinehealthandlifesciences
hierarchy_top_title Faculty of Medicine, Health and Life Sciences
hierarchy_parent_id facultyofmedicinehealthandlifesciences
hierarchy_parent_title Faculty of Medicine, Health and Life Sciences
department_str Swansea University Medical School - Pharmacy{{{_:::_}}}Faculty of Medicine, Health and Life Sciences{{{_:::_}}}Swansea University Medical School - Pharmacy
document_store_str 1
active_str 0
description Cyclopropane fatty acid synthases (CFAS) are a class of S-adenosylmethionine (SAM) dependent methyltransferase enzymes able to catalyse the cyclopropanation of unsaturated phospholipids. Since CFAS enzymes employ SAM as a methylene source to cyclopropanate alkene substrates, they have the potential to be mild and more sustainable biocatalysts for cyclopropanation transformations than current carbene based approaches. This work describes the characterisation of E. coli CFAS enzyme (ecCFAS) and its exploitation in the stereoselective biocatalytic synthesis of cyclopropyl lipids. ecCFAS was found to convert phosphatidylglycerol (PG) to methyl dihydrosterculate 1 from in up to 58% conversion and 73% ee and the absolute configuration (9S,10R) was established. Substrate tolerance of ecCFAS was found to be correlated with the electronic properties of phospholipid headgroups and for the first time ecCFAS was found to catalyse cyclopropanation of both phospholipid chains to form dicyclopropanated products. In addition, mutagenesis and in-silico experiments were carried out to identify the enzyme residues with key roles in catalysis and to provide structural insights into the lipid substrate preference of ecCFAS. Finally, the biocatalytic synthesis of methyl dihydrosterculate 1 and its deuterated analogue was also accomplished combining pure ecCFAS with the SAM regenerating AtHMT enzyme in presence of CH3I and CD3I.
published_date 2024-06-14T15:38:26Z
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