Current Topics in Medicinal Chemistry - Volume 1, Issue 1, 2001

Bacteria need a sufficient supply of iron in ionic form for their metabolism. When living in an environment where this is not possible (as in the soil due to the presence of highly unsoluble ferric oxide hydrates, or in living organisms where iron is bound to peptidic chelators) Fe 3 complexing compounds, called siderophores, are produced. The siderophores of Pseudomonas aeruginosa, a dangerous opportunistic human pathogen, and of related potentially pathogenic species will be presented.

Numerous bacterial proteins are involved in microbial iron uptake and transport and considerable variation has been found in the uptake schemes used by different bacterial species. However, whether extracting iron from host proteins such as transferrin, lactoferrin or hemoglobin or importing low molecular weight iron-chelating compounds such as heme, citrate or siderophores, Gram-negative pathogenic bacteria typically employ a specific outer membrane receptor, a periplasmic binding protein and two inner membrane associated proteins: a transporter coupled with an ATP-hydrolyzing protein. Often, studies have shown that proteins with similar function but little amino acid sequence homology are structurally related. Elucidation of the structures of the Escherichia coli outer membrane siderophore transport proteins FepA and FhuA have provided the first insights into the conformational changes required for ligand transport through the bacterial outer membrane. The variations between the structures of the prototypical periplasmic ferric binding protein FbpA from Neisseria and Haemophilus influenzae and the unusual E. coli periplasmic siderophore binding protein FhuD reveal that the different periplasmic ligand binding proteins exercise distinct mechanisms for ligand binding and release. The structure of the hemophore HasA from Serratia marcescens shows how heme may be extracted and utilized by the bacteria. Other biochemical evidence also shows that the proteins that provide energy for iron transport at the outer membrane, such as the TonB-ExbB-ExbD system, are structurally very similar across bacterial species. Likewise, the iron-sensitive gene regulatory protein Fur is found in most bacteria. To date, no structural information is available for Fur, but the structure for the related protein DxtR has been determined. Together, these three-dimensional structures complement our knowledge of iron transport systems from other pathogenic bacteria, including Pseudomonas aeruginosa, which has a number of homologous iron uptake proteins. More importantly, the current structures for iron transport proteins provide rational starting points for design of novel antimicrobial agents.

Tools for the identification of bacteria are of great importance especially for taxonomical and medical purposes. In the case of fluorescent pseudomonads a quick and unambiguous identification is possible by methods that are referred to as siderotyping. All of them are based upon the characterization of the bacterial siderophores or the receptors expressed for the uptake of these compounds. Different microbiological and bioanalytical tests that are accurate, rapid and easy to use will be described.

Pseudomonas aeruginosa is an opportunistic human pathogen characterized by an intrinsic resistance to multiple antimicrobial agents and the ability to develop high-level (acquired) multidrug resistance during antibiotic therapy. Much of this resistance is promoted by highly homologous three-component efflux systems of broad substrate specificity, of which four have been identified to date. These include MexA-MexB-OprM and MexX-MexY-OprM, which are expressed constitutively in wild type cells and, thus, provide for intrinsic multidrug resistance, and MexC-MexD-OprJ and MexE-MexF-OprN, whose expression so far has only been seen in acquired multidrug resistant mutant strains. Additional homologues of these efflux systems are identifiable in the recently released genome sequence, though their roles, if any, in antimicrobial efflux are unknown. These tripartite pumps are composed of an integral cytoplasmic membrane drug-proton antiporter of the resistance-nodulation-cell division (RND) family of exporters, a channel-forming outer membrane efflux protein (or outer membrane factor (OMF) and a periplasmic membrane fusion protein (MFP) that links the other two. In addition to a number of antimicrobials of clinical significance, these pumps also export dyes, detergents, disinfectants, organic solvents and acylated homoserine lactones involved in quorum-sensing. While the natural functional of these pumps remains undefined, the fact that they contribute to antimicrobial resistance in P. aeruginosa makes them reasonable targets for therapeutic intervention.

Pseudomonas aeruginosa is a dangerous opportunistic bacterium responsible for frequently lethal hospital (nosocomial) infections. It endangers especially severely injured patients suffering from large wounds or severe burns, as well as persons whose immune system is weakened. An extremely critical situation exists for patients suffering from mucoviscidosis (cystic fibrosis), when P. aeruginosa infects the bronchial tubes. P. aeruginosa is resistant against many desinfecting agents and, more important, an increasing number of strains especially from hospital isolates have become highly resistant against most antibiotics. The low permeability of the outer membrane and an active export mechanism for low molecular weight substances are the main reasons for the resistance. In addition, b-lactamase activity affects treatment with b-lactam antibiotics. An approach to overcome the problem of resistance lies in the synthesis of antibiotics conjugated with compounds active as siderophores. In this way the transport ways for iron complexes into the cell can be used (Trojan Horse strategy), and the presence of large substituents reduces the export and the b-lactamase activity. The results obtained with natural (pyoverdins) and synthetic (mainly catecholate) siderophores will be reviewed.