Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules
ABSTRACT The bacterial envelope plays a critical role in maintaining essential cellular functions by selectively regulating import and export. The selectivity of this envelope can restrict the utilisation of externally provided compounds, thereby restricting the functional space of cellular engineer...
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| Format: | Article |
| Language: | English |
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Wiley
2025-03-01
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| Series: | Microbial Biotechnology |
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| Online Access: | https://doi.org/10.1111/1751-7915.70122 |
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| author | Ivan Casas‐Rodrigo Tobias Vornholt Kathrin Castiglione Tania Michelle Roberts Markus Jeschek Thomas R. Ward Sven Panke |
| author_facet | Ivan Casas‐Rodrigo Tobias Vornholt Kathrin Castiglione Tania Michelle Roberts Markus Jeschek Thomas R. Ward Sven Panke |
| author_sort | Ivan Casas‐Rodrigo |
| collection | DOAJ |
| description | ABSTRACT The bacterial envelope plays a critical role in maintaining essential cellular functions by selectively regulating import and export. The selectivity of this envelope can restrict the utilisation of externally provided compounds, thereby restricting the functional space of cellular engineering. This study systematically investigates the potential of large pore outer membrane proteins (OMPs) to enhance outer membrane permeability for diverse challenging compounds. We focus on the general porin OmpF, which facilitates the diffusion of water and small molecules, and specific OMP transporters FhuA and FepA, which mediate the translocation of small hydrophilic compounds. Through comprehensive characterisation, we evaluate the effects of recombinant expression of OMPs and engineered variants for small and hydrophilic compounds, aromatic molecules and bulky molecules and apply our findings to address two critical contemporary transport challenges: the uptake of large metal‐containing cofactors for artificial metalloenzymes and non‐permeant fluorescent Halo‐ligands for in vivo protein labelling. Notably, we demonstrate significant improvements in ArM‐catalysis and labelling. This study provides a practical guide for designing experiments that include outer‐membrane‐transport‐limiting steps. This study highlights the potential of engineered OMPs to overcome the limitations imposed by the cell envelope, enabling the incorporation of complex molecules and expanding the frontiers of cellular engineering. |
| format | Article |
| id | doaj-art-ee19e20f7aac46d8bb6b2f1021373eae |
| institution | Kabale University |
| issn | 1751-7915 |
| language | English |
| publishDate | 2025-03-01 |
| publisher | Wiley |
| record_format | Article |
| series | Microbial Biotechnology |
| spelling | doaj-art-ee19e20f7aac46d8bb6b2f1021373eae2025-08-20T03:43:31ZengWileyMicrobial Biotechnology1751-79152025-03-01183n/an/a10.1111/1751-7915.70122Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex MoleculesIvan Casas‐Rodrigo0Tobias Vornholt1Kathrin Castiglione2Tania Michelle Roberts3Markus Jeschek4Thomas R. Ward5Sven Panke6Department of Biosystems Science and Engineering ETH Zurich Basel SwitzerlandDepartment of Biosystems Science and Engineering ETH Zurich Basel SwitzerlandInstitute of Bioprocess Engineering Friedrich‐Alexander‐Universität Erlangen‐Nürnberg Erlangen GermanyDepartment of Biosystems Science and Engineering ETH Zurich Basel SwitzerlandDepartment of Biosystems Science and Engineering ETH Zurich Basel SwitzerlandDepartment of Chemistry University of Basel Basel SwitzerlandDepartment of Biosystems Science and Engineering ETH Zurich Basel SwitzerlandABSTRACT The bacterial envelope plays a critical role in maintaining essential cellular functions by selectively regulating import and export. The selectivity of this envelope can restrict the utilisation of externally provided compounds, thereby restricting the functional space of cellular engineering. This study systematically investigates the potential of large pore outer membrane proteins (OMPs) to enhance outer membrane permeability for diverse challenging compounds. We focus on the general porin OmpF, which facilitates the diffusion of water and small molecules, and specific OMP transporters FhuA and FepA, which mediate the translocation of small hydrophilic compounds. Through comprehensive characterisation, we evaluate the effects of recombinant expression of OMPs and engineered variants for small and hydrophilic compounds, aromatic molecules and bulky molecules and apply our findings to address two critical contemporary transport challenges: the uptake of large metal‐containing cofactors for artificial metalloenzymes and non‐permeant fluorescent Halo‐ligands for in vivo protein labelling. Notably, we demonstrate significant improvements in ArM‐catalysis and labelling. This study provides a practical guide for designing experiments that include outer‐membrane‐transport‐limiting steps. This study highlights the potential of engineered OMPs to overcome the limitations imposed by the cell envelope, enabling the incorporation of complex molecules and expanding the frontiers of cellular engineering.https://doi.org/10.1111/1751-7915.70122cell envelopeOMPouter membranepermeabilitypore |
| spellingShingle | Ivan Casas‐Rodrigo Tobias Vornholt Kathrin Castiglione Tania Michelle Roberts Markus Jeschek Thomas R. Ward Sven Panke Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules Microbial Biotechnology cell envelope OMP outer membrane permeability pore |
| title | Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules |
| title_full | Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules |
| title_fullStr | Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules |
| title_full_unstemmed | Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules |
| title_short | Permeabilisation of the Outer Membrane of Escherichia coli for Enhanced Transport of Complex Molecules |
| title_sort | permeabilisation of the outer membrane of escherichia coli for enhanced transport of complex molecules |
| topic | cell envelope OMP outer membrane permeability pore |
| url | https://doi.org/10.1111/1751-7915.70122 |
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