Iron scavenging by Neisseria gonorrhoeae is accomplished by the expression of receptors that are specific for host iron-binding proteins, such as transferrin and lactoferrin. Efficient transferrin-iron acquisition is dependent on the combined action of two proteins, designated TbpA and TbpB. TbpA is a TonB-dependent outer membrane receptor, whereas TbpB is lipid modified and serves to increase the efficiency of transferrin-iron uptake. Both proteins, together or separately, can be isolated from the gonococcal outer membrane by using affinity chromatography techniques. In the present study, we identified an additional protein in transferrinaffinity preparations, which had an apparent molecular mass of 45 kDa. The ability to copurify this protein by transferrin affinity was dependent upon the presence of TbpA and not TbpB. The amino-terminal sequence of the 45-kDa protein was identical to the amino terminus of gonococcal TonB, indicating that TbpA stably interacted with TonB, without the addition of chemical cross-linkers. Using immunoprecipitation, we could recover TbpA-TonB complexes without the addition of transferrin, suggesting that ligand binding was not a necessary prerequisite for TonB interaction. In contrast, a characterized TonB box mutant of TbpA did not facilitate interaction between these two proteins such that complexes could be isolated. We generated an in-frame deletion of gonococcal TonB, which removed 35 amino acids, including a Neisseria-specific, glycinerich domain. This mutant protein, like the parental TonB, energized TbpA to enable growth on transferrin. Consistent with the functionality of this deletion derivative, TbpA-TonB complexes could be recovered from this strain. The results of the present study thus begin to define the requirements for a functional interaction between gonococcal TbpA and TonB.Bacterial pathogens infecting humans must propagate in an environment that maintains a very small reservoir of soluble iron (38), and since iron is required for the growth of nearly all microorganisms, efficient iron scavenging systems are essential. Most bacteria acquire this scarce but necessary nutrient by synthesis of low-molecular-weight siderophores in addition to membrane-bound transporters for ferric-siderophore uptake (35). However, the pathogenic Neisseria species are not known to secrete iron-scavenging siderophores (46) but have evolved outer membrane receptors that directly bind to host iron sources, such as transferrin and lactoferrin, and relieve them of their bound iron. The transferrin receptor is composed of two proteins: an integral outer membrane, TonB-dependent receptor (TbpA), and a surface-exposed lipoprotein (TbpB) (4, 15, 17). Together, these proteins specifically bind human transferrin and remove and internalize the iron in an energy-dependent fashion (13, 16). By analogy with the well-defined activities of the TonB-dependent siderophore receptors, we proposed that receptor-dependent iron removal from transferrin is accomplished at the expense of the cytoplasmic memb...

We conclude that observation of asymptomatic splenic cysts is safe regardless of size and that aspiration should be reserved for those who are not surgical candidates or in cases of diagnostic uncertainty.

The transferrin iron acquisition system of Neisseria gonorrhoeae consists of two dissimilar transferrin binding proteins (Tbp) A and B. TbpA is a TonB dependent transporter while TbpB is a lipoprotein that makes iron acquisition from transferrin (Tf) more efficient. In an attempt to further define the individual roles of these receptors in the process of Tf-iron acquisition, the kinetics of the receptor proteins in regards to ligand association and dissociation were evaluated. Tf association with TbpB was rapid as compared to TbpA. Tf dissociation from the wild-type receptor occurred in a biphasic manner; an initial rapid release was followed by a slower dissociation over time. Both TbpA and TbpB demonstrated a two-phase release pattern; however TbpA required both TonB and TbpB for efficient Tf dissociation from the cell surface. The roles of TbpA and TbpB in Tf dissociation were further examined, utilizing previously created HA fusion proteins. Using a Tf-utilization deficient TbpA-HA mutant, we concluded that the slower rate of ligand dissociation demonstrated by the wild-type transporter was a function of successful iron internalization. Insertion into the Cterminus of TbpB decreased the rate of Tf dissociation, while insertion into the N-terminus had no effect on this process. From these studies, we propose that TbpA and TbpB function synergistically during the process of Tf iron acquisition and that TbpB makes the process of Tf-iron acquisition more efficient at least in part by affecting association and dissociation of Tf from the cell surface.

No abstract