A synthetic system for expression of components of a bacterial microcompartment

Frank Sargent, Fordyce A. Davidson, Ciarán L. Kelly, Rachelle Binny, Natasha Christodoulides, David Gibson, Emelie Johansson, Katarzyna Kozyrska, Lucia Licandro Lado, Jane MacCallum, Rachel Montague, Brian Ortmann, Richard Owen, Sarah J. Coulthurst, Lionel Dupuy, Alan R. Prescott, Tracy Palmer

    Research output: Contribution to journalArticle

    23 Citations (Scopus)

    Abstract

    In general, prokaryotes are considered to be single-celled organisms that lack internal membrane-bound organelles. However, many bacteria produce proteinaceous microcompartments that serve a similar purpose; that is to concentrate specific enzymatic reactions together or to shield the wider cytoplasm from toxic metabolic intermediates. In this work, a synthetic operon encoding the key structural components of a microcompartment was designed based on the genes for the Salmonella propanediol utilisation (Pdu) microcompartment. The genes chosen included pduA, -B, -J, -K, -N, -T, and -U, and each were shown to produce protein in an Escherichia coli chassis. In parallel, a set of compatible vectors designed to express non-native cargo proteins were also designed and tested. Engineered hexa-Histidine tags allowed isolation of the components of the microcompartments together with co-expressed, untagged, cargo proteins. Finally, an in vivo protease accessibility assay suggested that a PduD-GFP fusion could be protected from proteolysis when co-expressed with the synthetic microcompartment operon. This work gives encouragement that it may be possible to harness the genes encoding a non-native microcompartment for future biotechnological applications.
    Original languageEnglish
    Pages (from-to)2427-2436
    Number of pages10
    JournalMicrobiology
    Volume159
    Issue number11
    DOIs
    Publication statusPublished - Nov 2013

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    Operon
    Propylene Glycols
    Genes
    Poisons
    Escherichia coli Proteins
    Histidine
    Salmonella
    Organelles
    Proteolysis
    Cytoplasm
    Proteins
    Peptide Hydrolases
    Bacteria
    Membranes

    Cite this

    Sargent, Frank ; Davidson, Fordyce A. ; Kelly, Ciarán L. ; Binny, Rachelle ; Christodoulides, Natasha ; Gibson, David ; Johansson, Emelie ; Kozyrska, Katarzyna ; Licandro Lado, Lucia ; MacCallum, Jane ; Montague, Rachel ; Ortmann, Brian ; Owen, Richard ; Coulthurst, Sarah J. ; Dupuy, Lionel ; Prescott, Alan R. ; Palmer, Tracy. / A synthetic system for expression of components of a bacterial microcompartment. In: Microbiology. 2013 ; Vol. 159, No. 11. pp. 2427-2436.
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    abstract = "In general, prokaryotes are considered to be single-celled organisms that lack internal membrane-bound organelles. However, many bacteria produce proteinaceous microcompartments that serve a similar purpose; that is to concentrate specific enzymatic reactions together or to shield the wider cytoplasm from toxic metabolic intermediates. In this work, a synthetic operon encoding the key structural components of a microcompartment was designed based on the genes for the Salmonella propanediol utilisation (Pdu) microcompartment. The genes chosen included pduA, -B, -J, -K, -N, -T, and -U, and each were shown to produce protein in an Escherichia coli chassis. In parallel, a set of compatible vectors designed to express non-native cargo proteins were also designed and tested. Engineered hexa-Histidine tags allowed isolation of the components of the microcompartments together with co-expressed, untagged, cargo proteins. Finally, an in vivo protease accessibility assay suggested that a PduD-GFP fusion could be protected from proteolysis when co-expressed with the synthetic microcompartment operon. This work gives encouragement that it may be possible to harness the genes encoding a non-native microcompartment for future biotechnological applications.",
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    Sargent, F, Davidson, FA, Kelly, CL, Binny, R, Christodoulides, N, Gibson, D, Johansson, E, Kozyrska, K, Licandro Lado, L, MacCallum, J, Montague, R, Ortmann, B, Owen, R, Coulthurst, SJ, Dupuy, L, Prescott, AR & Palmer, T 2013, 'A synthetic system for expression of components of a bacterial microcompartment', Microbiology, vol. 159, no. 11, pp. 2427-2436. https://doi.org/10.1099/mic.0.069922-0

    A synthetic system for expression of components of a bacterial microcompartment. / Sargent, Frank; Davidson, Fordyce A.; Kelly, Ciarán L.; Binny, Rachelle; Christodoulides, Natasha; Gibson, David; Johansson, Emelie; Kozyrska, Katarzyna; Licandro Lado, Lucia; MacCallum, Jane; Montague, Rachel; Ortmann, Brian; Owen, Richard; Coulthurst, Sarah J.; Dupuy, Lionel; Prescott, Alan R.; Palmer, Tracy.

    In: Microbiology, Vol. 159, No. 11, 11.2013, p. 2427-2436.

    Research output: Contribution to journalArticle

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    AU - Gibson, David

    AU - Johansson, Emelie

    AU - Kozyrska, Katarzyna

    AU - Licandro Lado, Lucia

    AU - MacCallum, Jane

    AU - Montague, Rachel

    AU - Ortmann, Brian

    AU - Owen, Richard

    AU - Coulthurst, Sarah J.

    AU - Dupuy, Lionel

    AU - Prescott, Alan R.

    AU - Palmer, Tracy

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    Sargent F, Davidson FA, Kelly CL, Binny R, Christodoulides N, Gibson D et al. A synthetic system for expression of components of a bacterial microcompartment. Microbiology. 2013 Nov;159(11):2427-2436. https://doi.org/10.1099/mic.0.069922-0