Cost-effective expression and purification of antimicrobial and host defense peptides in Escherichia coli

B. Bommarius, Håvard Jenssen, M. Elliott, J. Kindrachuk, Mukesh Pasupuleti, H. Gieren, K.-E. Jaeger, R.E.W. Hancock, D. Kalman

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    Resumé

    Cationic antimicrobial host defense peptides (HDPs) combat infection by directly killing a wide variety of microbes, and/or modulating host immunity. HDPs have great therapeutic potential against antibioticresistant bacteria, viruses and even parasites, but there are substantial roadblocks to their therapeutic application. High manufacturing costs associated with amino acid precursors have limited the delivery of inexpensive therapeutics through industrial-scale chemical synthesis. Conversely, the production of peptides in bacteria by recombinant DNA technology has been impeded by the antimicrobial activity of these peptides and their susceptibility to proteolytic degradation, while subsequent purification of
    recombinant peptides often requires multiple steps and has not been cost-effective. Here we have developed methodologies appropriate for large-scale industrial production of HDPs; in particular, we describe (i) a method, using fusions to SUMO, for producing high yields of intact recombinant HDPs in bacteria
    without significant toxicity and (ii) a simplified 2-step purification method appropriate for industrial use. We have used this method to produce seven HDPs to date (IDR1, MX226, LL37, CRAMP, HHC-10, E5 and E6). Using this technology, pilot-scale fermentation (10 L) was performed to produce large quantities of biologically active cationic peptides. Together, these data indicate that this new method represents a cost-effective means to enable commercial enterprises to produce HDPs in large-scale under Good Laboratory Manufacturing Practice (GMP) conditions for therapeutic application in humans.
    OriginalsprogEngelsk
    TidsskriftPeptides
    Vol/bind31
    Udgave nummer11
    Sider (fra-til)1957-1965
    ISSN0196-9781
    DOI
    StatusUdgivet - 2010

    Citer dette

    Bommarius, B., Jenssen, H., Elliott, M., Kindrachuk, J., Pasupuleti, M., Gieren, H., ... Kalman, D. (2010). Cost-effective expression and purification of antimicrobial and host defense peptides in Escherichia coli. Peptides, 31(11), 1957-1965. https://doi.org/10.1016/j.peptides.2010.08.008
    Bommarius, B. ; Jenssen, Håvard ; Elliott, M. ; Kindrachuk, J. ; Pasupuleti, Mukesh ; Gieren, H. ; Jaeger, K.-E. ; Hancock, R.E.W. ; Kalman, D. / Cost-effective expression and purification of antimicrobial and host defense peptides in Escherichia coli. I: Peptides. 2010 ; Bind 31, Nr. 11. s. 1957-1965.
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    abstract = "Cationic antimicrobial host defense peptides (HDPs) combat infection by directly killing a wide variety of microbes, and/or modulating host immunity. HDPs have great therapeutic potential against antibioticresistant bacteria, viruses and even parasites, but there are substantial roadblocks to their therapeutic application. High manufacturing costs associated with amino acid precursors have limited the delivery of inexpensive therapeutics through industrial-scale chemical synthesis. Conversely, the production of peptides in bacteria by recombinant DNA technology has been impeded by the antimicrobial activity of these peptides and their susceptibility to proteolytic degradation, while subsequent purification of recombinant peptides often requires multiple steps and has not been cost-effective. Here we have developed methodologies appropriate for large-scale industrial production of HDPs; in particular, we describe (i) a method, using fusions to SUMO, for producing high yields of intact recombinant HDPs in bacteria without significant toxicity and (ii) a simplified 2-step purification method appropriate for industrial use. We have used this method to produce seven HDPs to date (IDR1, MX226, LL37, CRAMP, HHC-10, E5 and E6). Using this technology, pilot-scale fermentation (10 L) was performed to produce large quantities of biologically active cationic peptides. Together, these data indicate that this new method represents a cost-effective means to enable commercial enterprises to produce HDPs in large-scale under Good Laboratory Manufacturing Practice (GMP) conditions for therapeutic application in humans.",
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    Bommarius, B, Jenssen, H, Elliott, M, Kindrachuk, J, Pasupuleti, M, Gieren, H, Jaeger, K-E, Hancock, REW & Kalman, D 2010, 'Cost-effective expression and purification of antimicrobial and host defense peptides in Escherichia coli', Peptides, bind 31, nr. 11, s. 1957-1965. https://doi.org/10.1016/j.peptides.2010.08.008

    Cost-effective expression and purification of antimicrobial and host defense peptides in Escherichia coli. / Bommarius, B.; Jenssen, Håvard; Elliott, M.; Kindrachuk, J.; Pasupuleti, Mukesh; Gieren, H.; Jaeger, K.-E.; Hancock, R.E.W.; Kalman, D.

    I: Peptides, Bind 31, Nr. 11, 2010, s. 1957-1965.

    Publikation: Bidrag til tidsskriftTidsskriftartikelForskningpeer review

    TY - JOUR

    T1 - Cost-effective expression and purification of antimicrobial and host defense peptides in Escherichia coli

    AU - Bommarius, B.

    AU - Jenssen, Håvard

    AU - Elliott, M.

    AU - Kindrachuk, J.

    AU - Pasupuleti, Mukesh

    AU - Gieren, H.

    AU - Jaeger, K.-E.

    AU - Hancock, R.E.W.

    AU - Kalman, D.

    PY - 2010

    Y1 - 2010

    N2 - Cationic antimicrobial host defense peptides (HDPs) combat infection by directly killing a wide variety of microbes, and/or modulating host immunity. HDPs have great therapeutic potential against antibioticresistant bacteria, viruses and even parasites, but there are substantial roadblocks to their therapeutic application. High manufacturing costs associated with amino acid precursors have limited the delivery of inexpensive therapeutics through industrial-scale chemical synthesis. Conversely, the production of peptides in bacteria by recombinant DNA technology has been impeded by the antimicrobial activity of these peptides and their susceptibility to proteolytic degradation, while subsequent purification of recombinant peptides often requires multiple steps and has not been cost-effective. Here we have developed methodologies appropriate for large-scale industrial production of HDPs; in particular, we describe (i) a method, using fusions to SUMO, for producing high yields of intact recombinant HDPs in bacteria without significant toxicity and (ii) a simplified 2-step purification method appropriate for industrial use. We have used this method to produce seven HDPs to date (IDR1, MX226, LL37, CRAMP, HHC-10, E5 and E6). Using this technology, pilot-scale fermentation (10 L) was performed to produce large quantities of biologically active cationic peptides. Together, these data indicate that this new method represents a cost-effective means to enable commercial enterprises to produce HDPs in large-scale under Good Laboratory Manufacturing Practice (GMP) conditions for therapeutic application in humans.

    AB - Cationic antimicrobial host defense peptides (HDPs) combat infection by directly killing a wide variety of microbes, and/or modulating host immunity. HDPs have great therapeutic potential against antibioticresistant bacteria, viruses and even parasites, but there are substantial roadblocks to their therapeutic application. High manufacturing costs associated with amino acid precursors have limited the delivery of inexpensive therapeutics through industrial-scale chemical synthesis. Conversely, the production of peptides in bacteria by recombinant DNA technology has been impeded by the antimicrobial activity of these peptides and their susceptibility to proteolytic degradation, while subsequent purification of recombinant peptides often requires multiple steps and has not been cost-effective. Here we have developed methodologies appropriate for large-scale industrial production of HDPs; in particular, we describe (i) a method, using fusions to SUMO, for producing high yields of intact recombinant HDPs in bacteria without significant toxicity and (ii) a simplified 2-step purification method appropriate for industrial use. We have used this method to produce seven HDPs to date (IDR1, MX226, LL37, CRAMP, HHC-10, E5 and E6). Using this technology, pilot-scale fermentation (10 L) was performed to produce large quantities of biologically active cationic peptides. Together, these data indicate that this new method represents a cost-effective means to enable commercial enterprises to produce HDPs in large-scale under Good Laboratory Manufacturing Practice (GMP) conditions for therapeutic application in humans.

    KW - Antimicrobial peptide

    KW - Host defense peptide

    KW - Bacterial expression system

    U2 - 10.1016/j.peptides.2010.08.008

    DO - 10.1016/j.peptides.2010.08.008

    M3 - Journal article

    VL - 31

    SP - 1957

    EP - 1965

    JO - Peptides

    JF - Peptides

    SN - 0196-9781

    IS - 11

    ER -