Ferric-coprogen receptor FhuE of Escherichia coli: processing and sequence common to all TonB-dependent outer membrane receptor proteins

Abstract

Iron transport via siderophores requires outer membrane receptor proteins and the TonB protein. The FhuE protein of Escherichia coli functions as the receptor for ferric coprogen and ferric-rhodotorulic acid. A chromosomal DNA fragment bearing the flauE gene was cloned into pACYC184. The gene was localized by insertion mutagenesis by using the transposon TnlOOO. Expression in minicells revealed a FhuE precursor with an apparent molecular weight of 82,000 and a FhuE protein with a molecular weight of 76,000. The transcription polarity of the JIuE gene was deduced from the size of truncated polypeptides derived from Tn1000 insertions, which were mapped by restriction analysis. The processing of truncated precursors that were synthesized by insertion mutants was strongly reduced even when the insertion site was close to the carboxy terminus of the FhuE protein. It is concluded that either the efficent insertion of proFhuE into the cytoplasmic membrane or the rate of cleavage of the signal peptide requires a particular conformation of the proFhuE protein, which is only formed by the complete primary structure. The amino-terminal amino acid sequence deduced from the nucleotide sequence was confirmed by gas-phase sequencing of the precursor and the mature form, which were separated by electrophoresis on polyacrylamide gels. The precursor contained an unusually long signal peptide of 36 amino acids. The amino-terminal end of the mature form contained the sequence Glu-Thr-Val-Ile-Val. A pentapeptide starting with either Glu or Asp, followed by Thr, and two uncharged residues ending with Val were found in all outer membrane receptor proteins that were constituents of TonB-dependent transport systems.From results of studies concerning the energy requirement and TonB dependence of the irreversible adsorption of phages Ti and 480 to Escherichia coli cells, we concluded in 1976 (14) that the function determined by the tonB region may be to couple the energized state of the cytoplasmic membrane to the TonA outer membrane receptor protein.Later it was shown that ligands did bind to their receptors in unenergized cells or in tonB mutants but remained there and were not translocated across the outer membrane (7,18,32). In some cases the requirement for the outer membrane receptor protein for uptake could be bypassed by osmotic shock (7) or by the preparation of spheroplasts (36). Under these conditions the TonB function was no longer needed, suggesting that it was indeed involved in the translocation of iron siderophores (12, 15), vitamin B12 (3, 32), and the B-group colicins (31) across the outer membrane. Despite this consistent evidence in favor of the original proposal, direct proof of a physical interaction between the TonB protein (25, 28-30) and outer membrane receptor proteins is lacking.Recently, the nucleotide sequences of the structural genes for the receptors of vitamin B12 (19), ferric aerobactin (21), ferrichrome (9; R. Burkhardt, unpublished data), and ferric enterochelin (23) were determined. They all possess a c...

A lipoprotein of Yersinia enterocolitica facilitates ferrioxamine uptake in Escherichia coli

“…In addition, a lack of PCPYe had no effect on ferrioxamine transport in Y. enterocolitica. A low level of cobalamine transport independent of TonB was the result of disruption of the outer membrane permeability barrier by insertion of elevated amounts of outer membrane proteins (29). In this case a 17-kDa polypeptide and a 84-kDa polypeptide were observed to complement the BtuB phenotype.…”

Identification and Characterization of an Outer Membrane Receptor Gene in <i>Acinetobacter baumannii</i> Required for Utilization of Desferricoprogen, Rhodotorulic Acid, and Desferrioxamine B as Xenosiderophores

“…34) In any event, it should be noted that these receptors for multiple ferric siderophores contain the conserved amino acid residues in the TonB boxes involved in the interaction of these receptors with the TonBExbB-ExbD complex as the energy supplying system. 13,30) In contrast, little is known about multiple receptors specific to an inherent siderophore, with the exception of the type 1 ferric pyoverdine receptors, FpvA and FpvB, in P. aeruginosa 35) and the ferric vibrioferrin receptors, PvuA1 and PvuA2, in Vibrio parahaemolyticus. 36) Considering that A. baumannii normally inhabits soil and water surroundings, utilization of desferricoprogen, rhodotorulic acid, and desferrioxamine B may represent one of the strategies that the bacterium has adopted to compete with other neighboring microorganisms for survival and proliferation within its various niches, and promote its opportunities of gaining access to human host.…”

“…In addition, the FhuE protein was found to possess the TonB box, QTIQV, characteristic of the TonB-dependent OMPs, where the invariant amino acids threonine and valine (underlined) are present at the same positions as in E. coli FhuE. 13,30) Iron-Repressible Transcription of fhuE To test the function of the putative Fur box for transcription of fhuE, RT-PCR was performed with total RNAs isolated from A. baumannii ATCC19606 grown under iron-replete and ironlimiting conditions. The total RNA isolated from the iron-limiting culture provided the anticipated 226-bp product, while that from the iron-replete culture did not (Fig.…”

FhuF, an iron‐regulated protein of Escherichia coli with a new type of [2Fe‐2S] center

“…However, it cannot be excluded that a minor fraction of FhuF is associated with the cytoplasmic membrane. Ferrioxamine B is transported across the outer membrane of E. coli K-12 mainly through the action of the FhuE protein [29], which transports the siderophore coprogen with high efficiency and ferrioxamine B with very low efficiency. When the specific ferrioxamine B receptor FoxA from Yersinia enterocolitica was cloned into E. coli, ferrioxamine B was transported with a high efficiency [4].…”