Peptide deformylase catalyzes the deformylation reaction of the amino terminal fMet residue of newly synthesized proteins in bacteria, and most likely in Plasmodium falciparum, and has therefore been identified as a potential antibacterial and antimalarial drug target. The structure of P. falciparum peptide deformylase, determined at 2.8 A resolution with ten subunits per asymmetric unit, is similar to the bacterial enzyme with the residues involved in catalysis, the position of the bound metal ion, and a catalytically important water structurally conserved between the two enzymes. However, critical differences in the substrate binding region explain the poor affinity of E. coli deformylase inhibitors and substrates toward the Plasmodium enzyme. The Plasmodium structure serves as a guide for designing novel antimalarials.

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An altered version of peptide deformylase from Plasmodium falciparum (PfPDF), the organism that causes the most devastating form of malaria, has been cocrystallized with a synthesized inhibitor that has submicromolar affinity for its target protein. The structure is solved at 2.2 Å resolution, an improvement over the 2.8 Å resolution achieved during the structural determination of unliganded PfPDF. This represents the successful outcome of modifying the protein construct in order to overcome adverse crystal contacts and other problems encountered in the study of unliganded PfPDF. Two molecules of PfPDF are found in the asymmetric unit of the current structure. The active site of each monomer of PfPDF is occupied by a proteolyzed fragment of the tripeptide-like inhibitor. Unexpectedly, each PfPDF subunit is associated with two nearly complete molecules of the inhibitor, found at a protein-protein interface. This is the first structure of a eukaryotic PDF protein, a potential drug target, in complex with a ligand.Keywords: crystal engineering; drug design; malaria; PDF; Plasmodium Prokaryotic protein synthesis begins with a formylmethionine residue. The resulting amino-terminal formyl group is subsequently removed by a metalloenzyme, peptide deformylase (PDF), during bacterial protein maturation. The resulting amino-terminal methionine is removed in many cases by methionine aminopeptidase, which has essentially no activity for N-formylated methionine substrates. Some proteins that fail to undergo this process of maturation are inactive. In support of this understanding, inhibitors of the first step of this process have been shown to be bacteriostatic in vitro (Apfel et al. 2001b). In recent years, inhibitors of bacterial peptide deformylases have made significant preclinical progress as novel antibacterial agents (Apfel et al. 2001a,b;Hackbarth et al. 2002;Wise et al. 2002;Roblin and Hammerschlag 2003), including very encouraging in vivo results (Clements et al. 2001). The unexpected finding of a human mitochondrial peptide deformylase has apparently not dampened enthusiasm for this potential new class of antibacterials. The lack of reported toxicity to human and other animal cells, despite evident antibacterial action, has prompted the suggestion that the human mitochondrial PDF protein may not be functional, or that the tested inhibitors may not be reaching the mitochondrion (Nguyen et al. 2003).Here, we focus on the peptide deformylase from a major human pathogen, a unicellular eukaryotic protozoan, Plasmodium falciparum. P. falciparum is the causative agent of the most deadly form of malaria, a disease that causes up to 2 million deaths annually, mostly of children under the age of five (WHO 2000). The P. falciparum genome was found to contain an ORF with homology to bacterial def genes that code for the peptide deformylase protein (Bracchi-Ricard et al. 2001 Article published online ahead of print. Article and publication date are at

Interactions involving hydrogen transfer were studied in a coculture of two hyperthermophilic microorganisms: Thermotoga maritima, an anaerobic heterotroph, and Methanococcus jannaschii, a hydrogenotrophic methanogen. Cell densities of T. maritima increased 10‐fold when cocultured with M. jannaschii at 85°C, and the methanogen was able to grow in the absence of externally supplied H2 and CO2. The coculture could not be established if the two organisms were physically separated by a dialysis membrane, suggesting the importance of spatial proximity. The significance of spatial proximity was also supported by cell cytometry, where the methanogen was only found in cell sorts at or above 4.5 μm in samples of the coculture in exponential phase. An unstructured mathematical model was used to compare the influence of hydrogen transport and metabolic properties on mesophilic and hyperthermophilic cocultures. Calculations suggest the increases in methanogenesis rates with temperature result from greater interactions between the methanogenic and fermentative organisms, as evidenced by the sharp decline in H2 concentration in the proximity of a hyperthermophilic methanogen. The experimental and modeling results presented here illustrate the need to consider the interactions within hyperthermophilic consortia when choosing isolation strategies and evaluating biotransformations at elevated temperatures. © 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 56: 268–278, 1997.

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