dc.identifier.uri |
http://dx.doi.org/10.15488/1250 |
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dc.identifier.uri |
http://www.repo.uni-hannover.de/handle/123456789/1275 |
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dc.contributor.author |
Roberts, Douglas M.
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dc.contributor.author |
Bartel, Christoph
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dc.contributor.author |
Scott, Alan
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dc.contributor.author |
Ivison, David
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dc.contributor.author |
Simpson, Thomas J.
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dc.contributor.author |
Cox, Russell J.
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dc.date.accessioned |
2017-03-31T08:16:06Z |
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dc.date.available |
2017-03-31T08:16:06Z |
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dc.date.issued |
2017 |
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dc.identifier.citation |
Roberts, Douglas M.; Bartel, Christoph; Scott, A.; Ivison, D.; Simpson, T.J. et al.: Substrate selectivity of an isolated enoyl reductase catalytic domain from an iterative highly reducing fungal polyketide synthase reveals key components of programming. In: Chemical Science 8 (2017), Nr. 2, S. 1116-1126. DOI: https://doi.org/10.1039/c6sc03496a |
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dc.description.abstract |
A cis-acting enoyl reductase (ER) catalytic domain was isolated from a fungal highly reducing iterative polyketide synthase (HR-iPKS) for the first time and studied in vitro. The ER from the squalestatin tetraketide synthase forms a discrete dimeric protein in solution. The ER shows broad substrate selectivity, reducing enoyl species including both natural and unnatural substrates. Pantetheine-bound substrate thiolesters reacted much faster than the corresponding SNAC thiolesters. The unnatural substrates included Z-olefins, 2-ethyl olefins and pentaketides. Methylation of the substrate modifies the activity of the ER such that the 2,4-dimethyl oct-2-enoyl substrate fits into the active site but cannot be reduced. A new NMR-based assay was developed for the direct observation of the stereochemical preferences at the 4′ position of the NADPH cofactor and the C-2 and C-3 positions of the substrates. The assay reveals that the fungal iPKS ER-catalysed reaction is stereochemically identical to that of the vertebrate FAS (vFAS) at the cofactor 4′ position and the substrate 3-position, but the high stereoselectivity displayed by intact SQTKS is lost such that reprotonation at the 2-position is unselective by the isolated ER. A 3D model of ER was consistent with these observations and showed that the ER may sequester its final substrate to prevent further chain extension. The results support a developing model for programming by HR-iPKS in which competition for substrates between restrictive and permissive catalytic domains chaperones the growing polyketide to completion, while allowing for errors and evolution. |
eng |
dc.description.sponsorship |
EPSRC/EP/F066104/1 |
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dc.description.sponsorship |
DFG/INST 187/621 |
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dc.description.sponsorship |
BBSRC/BB/I003355/1 |
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dc.description.sponsorship |
School of Chemistry, University of Bristol |
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dc.description.sponsorship |
MINAS |
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dc.language.iso |
eng |
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dc.publisher |
Cambridge : Royal Society of Chemistry |
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dc.relation.ispartofseries |
Chemical Science 8 (2017), Nr. 2 |
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dc.rights |
CC BY 3.0 Unported |
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dc.rights.uri |
https://creativecommons.org/licenses/by/3.0/ |
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dc.subject |
Alkylation |
eng |
dc.subject |
Fungi |
eng |
dc.subject |
Ketones |
eng |
dc.subject |
Olefins |
eng |
dc.subject |
Catalytic domains |
eng |
dc.subject |
Chain extension |
eng |
dc.subject |
Dimeric proteins |
eng |
dc.subject |
Direct observations |
eng |
dc.subject |
High stereoselectivities |
eng |
dc.subject |
Polyketide synthases |
eng |
dc.subject |
Reprotonation |
eng |
dc.subject |
Substrate selectivity |
eng |
dc.subject |
Substrates |
eng |
dc.subject.ddc |
540 | Chemie
|
ger |
dc.title |
Substrate selectivity of an isolated enoyl reductase catalytic domain from an iterative highly reducing fungal polyketide synthase reveals key components of programming |
eng |
dc.type |
Article |
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dc.type |
Text |
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dc.relation.issn |
2041-6520 |
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dc.relation.doi |
https://doi.org/10.1039/c6sc03496a |
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dc.bibliographicCitation.issue |
2 |
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dc.bibliographicCitation.volume |
8 |
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dc.bibliographicCitation.firstPage |
1116 |
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dc.bibliographicCitation.lastPage |
1126 |
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dc.description.version |
publishedVersion |
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tib.accessRights |
frei zug�nglich |
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