A mevalonate-independent pathway of isoprenoid biosynthesis present in Plasmodium falciparum was shown to represent an effective target for chemotherapy of malaria. This pathway includes 1-deoxy-D-xylulose 5-phosphate (DOXP) as a key metabolite. The presence of two genes encoding the enzymes DOXP synthase and DOXP reductoisomerase suggests that isoprenoid biosynthesis in P. falciparum depends on the DOXP pathway. This pathway is probably located in the apicoplast. The recombinant P. falciparum DOXP reductoisomerase was inhibited by fosmidomycin and its derivative, FR-900098. Both drugs suppressed the in vitro growth of multidrug-resistant P. falciparum strains. After therapy with these drugs, mice infected with the rodent malaria parasite P. vinckei were cured.

The gcpE and lytB gene products control the terminal steps of isoprenoid biosynthesis via the 2-C-methyl-D-erythritol 4-phosphate pathway in Escherichia coli. In lytB-deficient mutants, a highly immunogenic compound accumulates significantly, compared to wild-type E. coli, but is apparently absent in gcpE-deficient mutants. Here, this compound was purified from E. coli v vlytB mutants by preparative anion exchange chromatography, and identified by mass spectrometry, 1 H, 13 C and 31 P NMR spectroscopy, and NOESY analysis as (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate (HMB-PP). HMB-PP is 10 4 times more potent in activating human VQ Q9/VN N2 T cells than isopentenyl pyrophosphate. ß

Human VQ Q9/VN N2 T cells play a crucial role in the immune response to microbial pathogens, yet their unconventional reactivity towards non-peptide antigens has been enigmatic until recently. The break-through in identi¢cation of the speci¢c activator was only possible due to recent success in a seemingly remote ¢eld: the elucidation of the reaction steps of the newly discovered 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway of isoprenoid biosynthesis that is utilised by many pathogenic bacteria. Unexpectedly, the intermediate of the MEP pathway, (E)-4-hydroxy-3-methyl-but-2-enyl-pyrophosphate) (HMB-PP), turned out to be by far the most potent VQ Q9/VN N2 T cell activator known, with an EC 50 of 0.1 nM. ß

Approximately 40% of the world population live in areas with the risk of malaria. Each year, 300-500 million people suffer from acute malaria, and 0.5-2.5 million die from the disease. Although malaria has been widely eradicated in many parts of the world, the global number of cases continues to rise. The most important reason for this alarming situation is the rapid spread of malaria parasites that are resistant to antimalarial drugs, especially chloroquine, which is by far the most frequently used. The development of new antimalarial drugs has been neglected since the 1970s owing to the end colonialism, changes in the areas of military engagement, and the restricted market potential. Only in recent years, in part supported by public funding programs, has interest in the development of antimalarial drugs been renewed. New data available from the recently sequenced genome of the malaria parasite Plasmodium falciparum and the application of methods of modern drug design promise to bring significant development in the fight against this disease.

In all organisms studied so far, isoprenoids such as dolichol, ubiquinones, carotenoids, and sterols are synthesized from isopentenyl diphosphate (IPP) and its isomer, dimethylallyl diphosphate (DMAPP). However, there are two completely different biosynthetic pathways leading to these two precursor molecules. In animals, fungi, archaea, and some bacteria, IPP and DMAPP are synthesized via the well-known mevalonate pathway. In contrast, the vast majority of bacteria, and some parasitic protozoa of the phylum Apicomplexa, synthesize IPP and DMAPP via the 2-C-methyl-D-erythritol-4-phosphate (MEP) pathway [also known as the 1-deoxy-D-xylulose-5-phosphate (DOXP), or non-mevalonate pathway]. [1][2][3] Since the MEP pathway is not used by humans, it represents an attractive target for the development of new antimicrobial compounds and indeed, inhibitors of the second enzyme of the MEP pathway, DOXP reductoisomerase, have demonstrated good antibacterial as well as antimalarial activity, in clinical settings. [4][5][6] Both IPP and DMAPP are formed (in a ~5:1 ratio) in the last step of the MEP pathway from the substrate (E)-4-hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP) in the following reaction: catalyzed by the enzyme HMBPP reductase, also known as LytB or IspH. 7 The reaction is thought to involve an iron-sulfur cluster reducing HMBPP to an allylic anion, followed by protonation at either C2 or C4, to form IPP or DMAPP. 8 However, the three-dimensional structure of the LytB enzyme has not yet been reported, making mechanistic analyses more challenging. Here, we report the X-ray crystallographic structure of the LytB enzyme from Aquifex aeolicus and propose a structure-based model for catalysis.We first discuss several features of LytB sequences in general that might be expected to be of importance for substrate binding and catalysis by considering the sequence homology between 224 LytB enzymes as annotated by the JPRED3 program. 9 There are three totally conserved cysteines in the A. aeolicus sequence that anchor the catalytically active iron-sulfur cluster: Cys13, Cys96, and Cys193 (Figure 1 and Supporting Information, Figure S1). HMBPP can be expected to bind to the iron-sulfur cluster during catalysis via its O4 atom, but it also needs to bind to protein residues and related diphosphates typically bind to prenyl synthase enzymes either via a DDXXD motif or via electrostatic/hydrogen bond interactions with Lys, Arg, or His. On the basis of the sequence alignment (Supporting Information Figure S1) we find neither evidence for DDXXD clusters, consistent with the lack of a requirement for Mg 2+ , nor totally conserved Lys or Arg residues or even totally conserved Lys/Arg positions. There are, however, two highly conserved His residues, H42 and H124. These two conserved His were reported in 10 putative LytB sequences by Adam et al. 11 and are now seen in all 224 sequences. This strongly suggests that the diphosphate binds to H42 and H124. The third feature that would be expected for LytB is the presence of a totally...