Publications by Year: 1989

The ability of Shigella flexneri to bind Congo red or hemin is associated with virulence. A 101-kilodalton (kDa) protein responsible for this phenotype (Crb+) in S. flexneri was identified by a tetramethylbenzidine staining procedure which detects heme-protein complexes in polyacrylamide gels. Labeling of cell-surface polypeptides with 125I revealed that the 101-kDa heme-binding protein is expressed on the cell surface. Expression of the protein was regulated by growth temperature and was found to be encoded by the large virulence plasmid of S. flexneri. Deletion mutants and a Tn5 insertion mutant which were negative for Congo red binding (Crb-) did not express the 101-kDa protein. Enteroinvasive Escherichia coli strains that were Crb+, and whose plasmids shared homology with the S. flexneri virulence plasmid, also expressed the 101-kDa protein. Expression of the protein in S. flexneri and enteroinvasive E. coli correlated with the presence of a 9.2-kilobase EcoRI fragment of these plasmids.

Iron limitation, a condition encountered within mammalian hosts, induces the synthesis of a number of proteins in pathogenic Shigella species. These include several outer membrane proteins, Shiga toxin, and proteins involved in the biosynthesis and transport of high-affinity iron-binding compounds or siderophores. Although siderophores have been shown to play a major role in the virulence of some bacterial pathogens, these compounds do not appear to be essential for the virulence of Shigella species. Unlike those pathogens which are restricted to the extracellular compartments of the host, the Shigella species invade and multiply within host cells. Alternative iron-acquisition systems, such as the ability to utilize haem-iron, permit growth of the intracellular bacteria. Virulent shigellae also possess a cell-surface haem-binding protein, and synthesis of this protein correlates with infectivity and virulence. This protein does not appear to be involved in iron acquisition. Rather, it may allow the bacteria to coat themselves with haem compounds, thus enhancing their ability to interact with target host cells.

Syntheses of hexanediamine-based dihydroxamate (Hexamate), spermidine-based trihydroxamate (Spermexatins), and spermidine-based mixed siderophore analogues (Spermexatols) are described. Key intermediates include the N-hydroxysuccinimide esters of various hydroxamic acids, e.g., malonohydroxamate, succinohydroxamate, and glutarohydroxamate. These intermediates were synthesized, characterized, and incorporated as the ligating chains on spermidine. Also, mixed iron chelating compounds (Spermexatols) with both catechol and hydroxamic acid side chains were synthesized. The reagent carbobenzoxyimidazole was employed to distinguish between the primary and secondary amino groups of spermidine. The ability of these iron chelators to stimulate microbial growth is also described.