Protein Farnesyltransferase and Protein Prenylation inPlasmodium falciparum

Abstract

Comparison of the malaria parasite and mammalian protein prenyltransferases and their cellular substrates is important for establishing this enzyme as a target for developing antimalarial agents. Nineteen heptapeptides differing only in their carboxyl-terminal amino acid were tested as alternative substrates of partially purified Plasmodium falciparum protein farnesyltransferase. Only NRSCAIM and NRSCAIQ serve as substrates, with NRSCAIM being the best. Peptidomimetics, FTI-276 and GGTI-287, inhibit the transferase with IC 50 values of 1 and 32 nM, respectively. Incubation of P. falciparum-infected erythrocytes with [ 3 H]farnesol labels 50-and 22-28-kDa proteins, whereas [ 3 H]geranylgeraniol labels only 22-28-kDa proteins. The 50-kDa protein is shown to be farnesylated, whereas the 22-28-kDa proteins are geranylgeranylated, irrespective of the labeling prenol. Protein labeling is inhibited more than 50% by either 5 M FTI-277 or GGTI-298. The same concentration of inhibitors also inhibits parasite growth from the ring stage by 50%, decreases expression of prenylated proteins as measured with prenyl-specific antibody, and inhibits parasite differentiation beyond the trophozoite stage. Furthermore, differentiation specific prenylation of P. falciparum proteins is demonstrated. Protein labeling is detected predominantly during the trophozoite to schizont and schizont to ring transitions. These results demonstrate unique properties of protein prenylation in P. falciparum: a limited specificity of the farnesyltransferase for peptide substrates compared with mammalian enzymes, the ability to use farnesol to label both farnesyl and geranylgeranyl moieties on proteins, differentiation specific protein prenylation, and the ability of peptidomimetic prenyltransferase inhibitors to block parasite differentiation.Malaria continues to be a major disease in tropical areas of the world. Parasite resistance to current drugs used in the treatment of malaria has lead investigators to seek out new drug targets. Among those targets are proteins and enzymes, which are necessary for cellular division and differentiation. Such targets include the protein prenyltransferases, which are necessary for the post-translational modification of proteins involved in the signal transduction pathways and in regulation of DNA replication and cell cycling (1-3). These enzymes are currently a major focus of efforts to design drugs that inhibit unregulated cell growth in cancer (4, 5). Several candidate compounds show great promise as antitumor drugs and are currently being tested in clinical trials. The potential for the application of such drugs to inhibit malarial parasite division and differentiation motivates one to examine the properties of the protozoan enzyme with the goal of identifying unique features of this enzyme that would make it a target for the development of parasite-specific drugs.A variety of proteins including small G-proteins, such as Ras, Rac, Rap, Rho, Rab (6), heterotrimeric G protein ␥ subunits (7), nuclear lamins (8), prot...

Structurally Simple, Potent, Plasmodium Selective Farnesyltransferase Inhibitors That Arrest the Growth of Malaria Parasites

“…7,8,12 While inhibition of farnesyltransferase has been demonstrated to have limited toxicity to mammalian cells at concentrations required to elicit a therapeutic response, 8 selective inhibition of parasitic farnesyltransferase may yet be an essential goal in the development of safe and effective antimalarial PFT inhibitors. To examine the selectivity of this new series of PFT inhibitors, 12 compounds were chosen on the basis of structural diversity and in vitro PfPFT activity and their 50% inhibition concentrations (IC 50 ) against both PfPFT and rat PFT were determined (Table 5).…”

“…In an effort to reduce development costs and accelerate access to new antimalarials, recent attention has been directed toward identifying antimalarial activity from agents developed for the treatment of other diseases. [7][8][9][10][11] On recognizing the essential role of prenylation for cellular function in lower eukaryotes, 7,12 several groups have investigated the antimalarial potential of inhibitors of protein farnesyltransferase (PFT a),8,13-16 a recognized key target for the interception of aberrant Ras activity common to many (~30%) human cancers. 17 Nallan et al have recently surveyed a number of mammalian-cell-optimized PFT inhibitors that are in clinical and preclinical development, including those from Bristol-Myers Squibb (BMS) (e.g., 1 16 and 2 8 ) and from the Hamilton/Sebti group (e.g., 3a and 3b ),13 for their abilities to inhibit Plasmodium falciparum PFT (PfPFT a ) enzyme activity (as determined by IC 50 values) and to arrest the in vitro growth of the intraerythrocytic forms of P. falciparum (as determined by ED 50 values, Figure 1).…”

Discovery of new targets for antimalarial chemotherapy

“…A recent genomic and phylogenic comparison revealed that apicomplexa contain unusual calcium response pathways that need deeper investigations and could be exploited as new therapeutic agents (Nagamune & Sibley, 2006 (Gelb et al, 2003). Furthermore, interest was strengthened by the fact that peptidomimetics and prenyl analogues of PFT substrates were shown to inhibit in vitro parasite growth (Chakrabarti et al, 2002;Ohkanda et al, 2001). Although the PGGT-I activity was reported, searches in the P. falciparum genome database revealed an apparent absence of PGGT-I gene .…”

Protein geranylgeranyltransferase-I of Trypanosoma cruzi

“…These are consistent with the CaaX specificity of the T. cruzi PGGT-I (Table1). Alterations in the residues involving substrate interactions in PFT and PGGT-I subunits between protozoan and mammalian orthologs may cause substantial differences in binding specificity for the CaaX motif and smallmolecule inhibitors in protozoa and mammalian cells [15][16][17][18][19][20][21]40]. This suggests not only different selectivity of inhibitors against parasite PFT and PGGT-I from that of mammalian enzymes but also differential consequences of inhibitory effects of PGGT-I or PFT on cellular events in the parasites from those in mammalian cells.…”

“…PFT has been found in pathogenic protozoan parasites including trypanosmatids (T. cruzi, T. brucei, and Leishmania), the malaria parasite (Plasmodium falciparum), and the enteric parasite Entamoeba histolytica [15][16][17][18]. PGGT-I has recently been found in E. histolytica [19].…”