Biochemistry. 2012 Aug 29. [Epub ahead of print]
The N-terminus and the Chain-length Determination (CLD) Domain Play a Role in the Length of the Isoprenoid Product of the Bifunctional Toxoplasma gondii Farnesyl-diphosphate Synthase
Li ZH, Cintrón R, Koon NA, Moreno SN.
Toxoplasma gondii possesses a bifunctional farnesyl diphosphate (FPP)/geranylgeranyl diphosphate (GGPP) synthase (TgFPPS) that synthesizes C15 and C20 isoprenoid diphosphates from isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). This enzyme has a unique arrangement of the 4th and 5th amino acid upstream to the First Aspartic Rich Domain (FARM) where the 4th amino acid is aromatic and the 5th is a cysteine. We mutated these amino acids converting the enzyme to an absolute FPPS by changing the cysteine to a tyrosine. The enzyme could be converted to an absolute GGPPS by changing both the 4th and 5th amino acids to alanines. We also constructed four mutated TgFPPSs whose regions around the first aspartate-rich motif were replaced with the corresponding regions of FPP synthases from Arabidopsis thaliana or Saccharomyces cerevisiae or with the corresponding regions of GGPP synthases from Homo sapiens or S. cerevisiae. We determined that the presence of a cysteine at the 4th position is essential for the TgFPPS bifunctionality. We also found that the length of the N-terminal domain has a role in determining the specificity and the length of the isoprenoid product. Phylogenetic analysis supports the grouping of this enzyme with other Type I FPPSs but the biochemical data indicates that TgFPPS has unique characteristics that differentiate it from mammalian FPPSs and GGPPSs and is therefore an important drug target.