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    Use of split-dihydrofolate reductase for the detection of protein-protein interactions and simultaneous selection of multiple plasmids in Plasmodium falciparum

    Osborne, Andrew (2020) Use of split-dihydrofolate reductase for the detection of protein-protein interactions and simultaneous selection of multiple plasmids in Plasmodium falciparum. Molecular and Biochemical Parasitology 238 , p. 111292. ISSN 0166-6851.

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    Abstract

    Defining protein-protein interactions is fundamental to the understanding of gene function. Protein-fragment complementation assays have been used for the analysis of protein-protein interactions in various organisms. The split-dihydrofolate reductase (DHFR) protein-fragment complementation assay utilises two complementary fragments of the enzyme fused to a pair of potentially interacting proteins. If these proteins interact, the DHFR fragments associate, fold into their native structure, reconstitute their function and confer resistance to antifolate drugs. We show that murine DHFR fragments fused to interacting proteins reconstitute a functional enzyme and confer resistance to the antifolate drug WR99210 in Plasmodium falciparum. These data demonstrate that the split-DHFR method can be used to detect in vivo protein-protein interactions in the parasite. Additionally, we show that split-DHFR fragments can be used as selection markers, permitting simultaneous selection of two plasmids in the presence of a single antifolate drug. Taken together, these experiments show that split-DHFR represents a valuable tool for the characterisation of Plasmodium protein function and genetic manipulation of the parasite.

    Metadata

    Item Type: Article
    Keyword(s) / Subject(s): Malaria, Split-Dihydrofolate reductase, Protein-Fragment complementation assay, Plasmodium transfection, Co-Transfection, Selection marker
    School: Birkbeck Schools and Departments > School of Science > Biological Sciences
    Depositing User: Andrew Osborne
    Date Deposited: 19 Jun 2020 14:18
    Last Modified: 28 Jun 2020 00:31
    URI: http://eprints.bbk.ac.uk/id/eprint/32306

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