Heterologous expression of plasmodial proteins for structural studies and functional annotation

Abstract

Malaria remains the world's most devastating tropical infectious disease with as many as 40% of the world population living in risk areas. The widespread resistance of Plasmodium parasites to the cost-effective chloroquine and antifolates has forced the introduction of more costly drug combinations, such as Coartem. In the absence of a vaccine in the foreseeable future, one strategy to address the growing malaria problem is to identify and characterize new and durable antimalarial drug targets, the majority of which are parasite proteins. Biochemical and structure-activity analysis of these proteins is ultimately essential in the characterization of such targets but requires large amounts of functional protein. Even though heterologous protein production has now become a relatively routine endeavour for most proteins of diverse origins, the functional expression of soluble plasmodial proteins is highly problematic and slows the progress of antimalarial drug target discovery. Here the status quo of heterologous production of plasmodial proteins is presented, constraints are highlighted and alternative strategies and hosts for functional expression and annotation of plasmodial proteins are reviewed.

Figure 1

Generalized strategy for the successful expression of plasmodial proteins. Extensive sequence analysis of a gene could facilitate genetic manipulation of the gene if needed. E. coli should remain the starting point for the expression of soluble proteins due to its ease of use and Xenopus remains the preferred choice for membrane protein expression. Several optimizations including codon adaptation, plasmodial chaperone co-expression and small sequence changes might be particularly necessary for the expression of the plasmodial protein of interest. Various other expression hosts including mammalian, baculovirus, yeasts, Toxoplasma, Dictyostelium and cell free systems have been used with various levels of success and might include an iterative process incorporating necessary optimizations.