Iron transport ferrichromes

Mainly the outer membrane ferrichrome receptor and transporter FhuA will be discussed because most structural and functional studies have been performed with this protein. In fact, FhuA was the first outer membrane protein identified (called TonA), with known functions as a phage and colicin receptor, that are related to iron transport (for a historical account, see Braun and Hantke 1977). 

Relationships between the structure of the siderophores and the iron transport were investigated for the fungus Neurospora crassa (160, 160a). Apparently two different receptors exist for ferrichromes and for coprogenes. For the recognition and the binding to the cell surface the iron configuration and the nature of the acyl chains is of importance. However, the transport system seems to be the same for both siderophore types dependent on the peptide part of the molecules. 

Two fluorescent siderophore analogs, one based on ferrichrome 173 and the second on ferrioxamine 188, were used to study iron transport in the fungus Ustilago maydis that has an uptake system for ferrichrome but lacks a defined ferrioxamine receptor. Nevertheless, ferrioxamine can be utilized by the fungus albeit at a slower rate. 

The tonB mutation, which had already been connected to iron transport (see above), also affected the outer membrane. Inspection of the region of the E. coli linkage map analogous to the sid locus in S. typhimurium revealed that tonA, the structural gene for the T5 outer membrane receptor, maps near pan. These considerations raised the possibility that the tonA product could be related to ferrichrome and a sid function in E. coli. The characteristics of the ton mutations known at that time, with the exception of albomycin resistance, are recorded in Table I. 

Ferrichrome iron transport in the enteric bacteria affords an experimentally feasible model for mechanistic studies. The ligand is rugged and can be labeled to high specific activity by microwave discharge activation of tritium gas. The labile ferric ion can be replaced with chromium to yield the kinetically stable isostructural chromic complex, chromichrome (61). Assuming this analog has the transport properties of the iron complex, its 3H counts would be an index of the behavior of... 

Salmonella typhimurium. Ferrichrome. Ferrichrome iron transport in enb7 proceeds by two concurrent mechanisms (Figure 12) (62). In one, the iron is snatched out rapidly with effective accumulation of the free ligand in solution. With added iron, the latter enters the cell at a rate identical to that of chromichrome. Again the A-cis isomer is active without dissociation, and reductive release is virtually certain. 

While most workers report the outer membrane siderophore receptors to have molecular weights in the 75-95K range, some variation in the magnitude of these numbers may be attributed to the preparative and analytical methods as well as to the particular standards used. Since enterobactin will rapidly remove iron from ferrichrome, the transport of the latter must perforce be studied in mutants lacking the former. However, such mutants often display multiple lesions. Additionally, isogenic strains have seldom been used and variations in media and cultural conditions will further confound attempts to compare results reported from different laboratories. 

In times of iron deficiency, many bacteria and fungi release low molecular weight chelators called siderophores (see Iron Transport Siderophores). These molecules bind ferric iron tightly and the ferric-siderophore complexes are then transported into the cell by a system of uptake proteins. The first stage in the uptake process involves an outer membrane receptor specific to each siderophore. One of the best characterized of these receptors is FhuA, the ferrichrome uptake receptor of E. coli, and we will describe this in detail. However, though other ferric-siderophore complexes are taken up by cells, and their iron released by systems similar to those of ferrichrome, their mechanisms may vary from those of ferrichrome in some respects. FepA and FecA" are two of the outer membrane ferric-siderophore receptors that have recently been structurally characterized. 

A Figure 24.17 The iron-transport system of a bacterial cell. The iron-binding ligand, called a siderophore, is synthesized inside the cell and excreted into the surrounding medium. It reacts with Fe ion to form ferrichrome, which is then absorbed by the cell. Inside the cell the ferrichrome is reduced, forming Fe ", which is not tightly bound by the siderophore. Having released the iron for use in the cell, the siderophore may be recycled back into the medium. 

The majority of Fur-regulated gene products are involved in iron uptake. Genes for transport and biosynthesis of enterobactin have been studied in E. coli K-12 (Earhart, 1996). It is assumed that this system is found in nearly every E. coli strain. Also the ferrichrome transport system seems to have a very broad distribution. The ferric citrate transport system (fee), however, is only present in some E. coli strains and may be part of a pathogenicity island. The aerobactin and yersiniabactin biosynthesis and transport systems are not found in all E. coli strains and are integrated into pathogenicity islands (Schubert et al., 1999). The ability to utilize haem seems also to be a specific pathogenicity-related adaptation. Haem transport systems are used in the animal or human host, where transferrin and lactoferrin create an iron-poor environment for bacteria. 

Koster, W. and Braun, V. (1990). Iron(III)hydroxamate transport binding of ferrichrome to the periplasmic FhuD protein, J. Biol. Chem., 265, 21 407-21 410. 

On the other hand, desmethyl-retro-hydroxamate ferrichrome 15, where the terminal methyl groups were replaced by hydrogen, showed no signihcant growth promotion activity toward Arthrobacter flavescence, and only one third of the iron(III) transport efficiency toward U. sphaerogena, which confirms the importance of the methyl groups... 

Escherichia coli has at least five independent transport systems, one of which is the low affinity pathway described above. In addition, it synthesizes enterobactin as a siderophore it can take up the iron(III) complex of ferrichrome, a siderophore synthesized by certain fungi there is a citrate-induced system, and a less common process involving aerobactin. 

The kinetic lability of ferric siderophores requires that transport experiments be performed with molecules bearing separate radioactive labels in the metal and ligand moieties. As coordination compounds the siderophores are thermodynamically stable and kinetically labile. The formation constants are typically 1030. In the case of ferrichrome the exchange half time at pH 6.3 and 37° is about 10 min (57). Published work (58, 59) with doubly labeled ferric schizokinen in Bacillus mega-terium and ferric aerobactin in A. aerogenes as well as a study of ferric enterobactin in E. coli (60) in each instance suggests a synchronous uptake mechanism for iron and ligand. 

E. coli. Ferrichrome. The uptake rates of 55Fe-ferrichrome, 3H-ferrichrome, and A-cis-3H-chromichrome, all in separate cultures of RW193, are shown in Figure 11 (62). The essentially identical rates for uptake of the 55Fe and 3H of the chromichrome suggests rapid transport of the intact coordination compound with concomitant rejection of the ligand. The latter does slowly penetrate the cell via a second mechanism, apparently again as the iron complex since addition of excess iron to the... 

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