Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586

Jack A. Connolly; Amber Wilson; Malgorzata Macioszek; Zhongshu Song; Luoyi Wang; Hadi H. Mohammad; Mukul Yadav; Maura di Martino; Claire E. Miller; Joanne Hothersall; Anthony S. Haines; Elton R. Stephens; Matthew P. Crump; Christine L. Willis; Thomas J. Simpson; Peter J. Winn; Christopher M. Thomas

doi:10.1038/s41598-018-38038-9

Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586

Jack A. Connolly, Amber Wilson, Malgorzata Macioszek, Zhongshu Song, Luoyi Wang, Hadi H. Mohammad, Mukul Yadav, Maura di Martino, Claire E. Miller, Joanne Hothersall, Anthony S. Haines, Elton R. Stephens, Matthew P. Crump, Christine L. Willis, Thomas J. Simpson, Peter J. Winn, Christopher M. Thomas^*

^*Corresponding author for this work

School of Chemistry

Research output: Contribution to journal › Article › peer-review

Abstract

The mupirocin trans-AT polyketide synthase pathway, provides a model system for manipulation of antibiotic biosynthesis. Its final phase involves removal of the tertiary hydroxyl group from pseudomonic acid B, PA-B, producing the fully active PA-A in a complex series of steps. To further clarify requirements for this conversion, we fed extracts containing PA-B to mutants of the producer strain singly deficient in each mup gene. This additionally identified mupM and mupN as required plus the sequence but not enzymic activity of mupL and ruled out need for other mup genes. A plasmid expressing mupLMNOPVCFU + macpE together with a derivative of the producer P. fluorescens strain NCIMB10586 lacking the mup cluster allowed conversion of PA-B to PA-A. MupN converts apo-mAcpE to holo-form while MupM is a mupirocin-resistant isoleucyl tRNA synthase, preventing self-poisoning. Surprisingly, the expression plasmid failed to allow the closely related P. fluorescens strain SBW25 to convert PA-B to PA-A.

Original language	English
Article number	1542
Number of pages	9
Journal	Scientific Reports
Volume	9
Early online date	7 Feb 2019
DOIs	https://doi.org/10.1038/s41598-018-38038-9
Publication status	E-pub ahead of print - 7 Feb 2019

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Connolly, J. A., Wilson, A., Macioszek, M., Song, Z., Wang, L., Mohammad, H. H., Yadav, M., di Martino, M., Miller, C. E., Hothersall, J., Haines, A. S., Stephens, E. R., Crump, M. P., Willis, C. L., Simpson, T. J., Winn, P. J., & Thomas, C. M. (2019). Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586. Scientific Reports, 9, Article 1542. Advance online publication. https://doi.org/10.1038/s41598-018-38038-9

@article{500b24c018a04aa09878ffd58725d449,

title = "Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586",

abstract = "The mupirocin trans-AT polyketide synthase pathway, provides a model system for manipulation of antibiotic biosynthesis. Its final phase involves removal of the tertiary hydroxyl group from pseudomonic acid B, PA-B, producing the fully active PA-A in a complex series of steps. To further clarify requirements for this conversion, we fed extracts containing PA-B to mutants of the producer strain singly deficient in each mup gene. This additionally identified mupM and mupN as required plus the sequence but not enzymic activity of mupL and ruled out need for other mup genes. A plasmid expressing mupLMNOPVCFU + macpE together with a derivative of the producer P. fluorescens strain NCIMB10586 lacking the mup cluster allowed conversion of PA-B to PA-A. MupN converts apo-mAcpE to holo-form while MupM is a mupirocin-resistant isoleucyl tRNA synthase, preventing self-poisoning. Surprisingly, the expression plasmid failed to allow the closely related P. fluorescens strain SBW25 to convert PA-B to PA-A.",

author = "Connolly, {Jack A.} and Amber Wilson and Malgorzata Macioszek and Zhongshu Song and Luoyi Wang and Mohammad, {Hadi H.} and Mukul Yadav and {di Martino}, Maura and Miller, {Claire E.} and Joanne Hothersall and Haines, {Anthony S.} and Stephens, {Elton R.} and Crump, {Matthew P.} and Willis, {Christine L.} and Simpson, {Thomas J.} and Winn, {Peter J.} and Thomas, {Christopher M.}",

note = "This work was funded by BBSRC grants BB/I014373/1 and BB/I014039/1, by BBSRC and EPSRC through the Bristol Centre for Synthetic Biology (BB/L01386X) and NPRONET PoC10. JAC and MdiM were funded by BBSRC MIBTP (BB/J014532/1) studentships. MY was funded by an Indian State Scholarship. HHM was funded by an Iraqi Government Scholarship (HCED).",

year = "2019",

month = feb,

day = "7",

doi = "10.1038/s41598-018-38038-9",

language = "English",

volume = "9",

journal = "Scientific Reports",

issn = "2045-2322",

publisher = "Nature publishing group",

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Connolly, JA, Wilson, A, Macioszek, M, Song, Z, Wang, L, Mohammad, HH, Yadav, M, di Martino, M, Miller, CE, Hothersall, J, Haines, AS, Stephens, ER, Crump, MP, Willis, CL, Simpson, TJ, Winn, PJ & Thomas, CM 2019, 'Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586', Scientific Reports, vol. 9, 1542. https://doi.org/10.1038/s41598-018-38038-9

TY - JOUR

T1 - Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586

AU - Connolly, Jack A.

AU - Wilson, Amber

AU - Macioszek, Malgorzata

AU - Song, Zhongshu

AU - Wang, Luoyi

AU - Mohammad, Hadi H.

AU - Yadav, Mukul

AU - di Martino, Maura

AU - Miller, Claire E.

AU - Hothersall, Joanne

AU - Haines, Anthony S.

AU - Stephens, Elton R.

AU - Crump, Matthew P.

AU - Willis, Christine L.

AU - Simpson, Thomas J.

AU - Winn, Peter J.

AU - Thomas, Christopher M.

N1 - This work was funded by BBSRC grants BB/I014373/1 and BB/I014039/1, by BBSRC and EPSRC through the Bristol Centre for Synthetic Biology (BB/L01386X) and NPRONET PoC10. JAC and MdiM were funded by BBSRC MIBTP (BB/J014532/1) studentships. MY was funded by an Indian State Scholarship. HHM was funded by an Iraqi Government Scholarship (HCED).

PY - 2019/2/7

Y1 - 2019/2/7

N2 - The mupirocin trans-AT polyketide synthase pathway, provides a model system for manipulation of antibiotic biosynthesis. Its final phase involves removal of the tertiary hydroxyl group from pseudomonic acid B, PA-B, producing the fully active PA-A in a complex series of steps. To further clarify requirements for this conversion, we fed extracts containing PA-B to mutants of the producer strain singly deficient in each mup gene. This additionally identified mupM and mupN as required plus the sequence but not enzymic activity of mupL and ruled out need for other mup genes. A plasmid expressing mupLMNOPVCFU + macpE together with a derivative of the producer P. fluorescens strain NCIMB10586 lacking the mup cluster allowed conversion of PA-B to PA-A. MupN converts apo-mAcpE to holo-form while MupM is a mupirocin-resistant isoleucyl tRNA synthase, preventing self-poisoning. Surprisingly, the expression plasmid failed to allow the closely related P. fluorescens strain SBW25 to convert PA-B to PA-A.

AB - The mupirocin trans-AT polyketide synthase pathway, provides a model system for manipulation of antibiotic biosynthesis. Its final phase involves removal of the tertiary hydroxyl group from pseudomonic acid B, PA-B, producing the fully active PA-A in a complex series of steps. To further clarify requirements for this conversion, we fed extracts containing PA-B to mutants of the producer strain singly deficient in each mup gene. This additionally identified mupM and mupN as required plus the sequence but not enzymic activity of mupL and ruled out need for other mup genes. A plasmid expressing mupLMNOPVCFU + macpE together with a derivative of the producer P. fluorescens strain NCIMB10586 lacking the mup cluster allowed conversion of PA-B to PA-A. MupN converts apo-mAcpE to holo-form while MupM is a mupirocin-resistant isoleucyl tRNA synthase, preventing self-poisoning. Surprisingly, the expression plasmid failed to allow the closely related P. fluorescens strain SBW25 to convert PA-B to PA-A.

U2 - 10.1038/s41598-018-38038-9

DO - 10.1038/s41598-018-38038-9

M3 - Article

SN - 2045-2322

VL - 9

JO - Scientific Reports

JF - Scientific Reports

M1 - 1542

ER -

Defining the genes for the final steps in biosynthesis of the complex polyketide antibiotic mupirocin by Pseudomonas fluorescens NCIMB10586

Abstract

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