Characterization of the pColV-K30 encoded cloacin DF13/aerobactin outer membrane receptor protein ofEscherichia coli; isolation and purification of the protein and analysis of its nucleotide sequence and primary structure

ABSTRACT Eight genes have been identified that function in the regulation, biosynthesis, and transport of rhizobactin 1021, a hydroxamate siderophore produced under iron stress bySinorhizobium meliloti. The genes were sequenced, and transposon insertion mutants were constructed for phenotypic analysis. Six of the genes, named rhbABCDEF, function in the biosynthesis of the siderophore and were shown to constitute an operon that is repressed under iron-replete conditions. Another gene in the cluster, named rhtA, encodes the outer membrane receptor protein for rhizobactin 1021. It was shown to be regulated by iron and to encode a product having 61% similarity to IutA, the outer membrane receptor for aerobactin. Transcription of both therhbABCDEF operon and the rhtA gene was found to be positively regulated by the product of the eighth gene in the cluster, named rhrA, which has characteristics of an AraC-type transcriptional activator. The six genes in therhbABCDEF operon have interesting gene junctions with short base overlaps existing between the genes. Similarities between the protein products of the biosynthesis genes and other proteins suggest that rhizobactin 1021 is synthesized by the formation of a novel siderophore precursor, 1,3-diaminopropane, which is then modified and attached to citrate in steps resembling those of the aerobactin biosynthetic pathway. The cluster of genes is located on the pSyma megaplasmid of S. meliloti 2011. Reverse transcription-PCR with RNA isolated from mature alfalfa nodules yielded no products for rhbF or rhtA at a time when the nifH gene was strongly expressed, indicating that siderophore biosynthesis and transport genes are not strongly expressed when nitrogenase is being formed in root nodules. Mutants having transposon insertions in the biosynthesis or transport genes induced effective nitrogen-fixing nodules on alfalfa plants.

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Deletion of the proline-rich region of TonB disrupts formation of a 2:1 complex with FhuA, an outer membrane receptor ofEscherichia coli

Protein Science ◽

10.1110/ps.051342505 ◽

2005 ◽

Vol 14 (5) ◽

pp. 1266-1273 ◽

Cited By ~ 12

Author(s):

Cezar M. Khursigara ◽

Gregory De Crescenzo ◽

Peter D. Pawelek ◽

James W. Coulton

Keyword(s):

Outer Membrane ◽

Membrane Receptor ◽

Ofescherichia Coli ◽

Outer Membrane Receptor ◽

Rich Region ◽

Proline Rich Region

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Identification and characterization of an iron-regulated gene, chtA, required for the utilization of the xenosiderophores aerobactin, rhizobactin 1021 and schizokinen by Pseudomonas aeruginosa

Microbiology ◽

10.1099/mic.0.28552-0 ◽

2006 ◽

Vol 152 (4) ◽

pp. 945-954 ◽

Cited By ~ 19

Author(s):

Páraic Ó Cuív ◽

Paul Clarke ◽

Michael O'Connell

Keyword(s):

Pseudomonas Aeruginosa ◽

Outer Membrane ◽

Membrane Receptor ◽

Outer Membrane Receptor ◽

Significant Similarity ◽

Hydroxamate Siderophores ◽

Insertional Inactivation ◽

Titration Assay

Pseudomonas aeruginosa utilizes several xenosiderophores under conditions of iron limitation, including the citrate hydroxamate siderophore aerobactin. Analysis of the P. aeruginosa genome sequence revealed the presence of two genes, chtA (PA4675) and PA1365, encoding proteins displaying significant similarity to the aerobactin outer-membrane receptor, IutA, of Escherichia coli. The chtA and PA1365 genes were mutated by insertional inactivation and it was demonstrated that ChtA is the outer-membrane receptor for aerobactin. ChtA also mediated the utilization of rhizobactin 1021 and schizokinen, which are structurally similar to aerobactin. In contrast to the utilization of other xenosiderophores by P. aeruginosa, there was no apparent redundancy in the utilization of aerobactin, rhizobactin 1021 and schizokinen. The utilization of citrate hydroxamate siderophores by P. aeruginosa was demonstrated to be TonB1 dependent. A Fur box was identified in the region directly upstream of chtA and it was demonstrated by the in vivo Fur titration assay that this region is capable of binding Fur and accordingly that expression of chtA is iron regulated. The PA1365 mutant was unaffected in the utilization of citrate hydroxamate siderophores.

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Normal iron-enterochelin uptake in mutants lacking the colicin I outer membrane receptor protein of Escherichia coli.

Journal of Bacteriology ◽

10.1128/jb.130.3.1399-1401.1977 ◽

1977 ◽

Vol 130 (3) ◽

pp. 1399-1401 ◽

Cited By ~ 1

Author(s):

S Soucek ◽

J Konisky

Keyword(s):

Escherichia Coli ◽

Outer Membrane ◽

Membrane Receptor ◽

Receptor Protein ◽

Outer Membrane Receptor

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Characterization of the Outer Membrane Receptor ShuA from the Heme Uptake System ofShigella dysenteriae

Journal of Biological Chemistry ◽

10.1074/jbc.m611121200 ◽

2007 ◽

Vol 282 (20) ◽

pp. 15126-15136 ◽

Cited By ~ 34

Author(s):

Kimberly A. Burkhard ◽

Angela Wilks

Keyword(s):

Outer Membrane ◽

Membrane Receptor ◽

Uptake System ◽

Outer Membrane Receptor ◽

Heme Uptake

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Identification of rhtX and fptX, Novel Genes Encoding Proteins That Show Homology and Function in the Utilization of the Siderophores Rhizobactin 1021 by Sinorhizobium meliloti and Pyochelin by Pseudomonas aeruginosa, Respectively

Journal of Bacteriology ◽

10.1128/jb.186.10.2996-3005.2004 ◽

2004 ◽

Vol 186 (10) ◽

pp. 2996-3005 ◽

Cited By ~ 54

Author(s):

Páraic Ó Cuív ◽

Paul Clarke ◽

Damien Lynch ◽

Michael O'Connell

Keyword(s):

Pseudomonas Aeruginosa ◽

Outer Membrane ◽

Membrane Transport ◽

Sinorhizobium Meliloti ◽

Iron Acquisition ◽

Membrane Receptor ◽

Receptor Protein ◽

Outer Membrane Receptor ◽

E Coli ◽

Genes Encoding

ABSTRACT Rhizobactin 1021 is a hydroxymate siderophore produced by the soil bacterium Sinorhizobium meliloti 2011. A regulon comprising rhtA, encoding the outer membrane receptor protein for the ferrisiderophore; the biosynthesis operon rhbABCDEF; and rhrA, the Ara-C-like regulator of the receptor and biosynthesis genes has been previously described. We report the discovery of a gene, located upstream of rhbA and named rhtX (for “rhizobactin transport”), which is required, in addition to rhtA, to confer the ability to utilize rhizobactin 1021 on a strain of S. meliloti that does not naturally utilize the siderophore. Rhizobactin 1021 is structurally similar to aerobactin, which is transported in Escherichia coli via the IutA outer membrane receptor and the FhuCDB inner membrane transport system. E. coli expressing iutA and fhuCDB was found to also transport rhizobactin 1021. We demonstrated that RhtX alone could substitute for FhuCDB to transport rhizobactin 1021 in E. coli. RhtX shows similarity to a number of uncharacterized proteins which are encoded proximal to genes that are either known to be or predicted to be involved in iron acquisition. Among these is PA4218 of Pseudomonas aeruginosa, which is located close to the gene cluster that functions in pyochelin biosynthesis and outer membrane transport. PA4218 was mutated by allelic replacement, and the mutant was found to have a pyochelin utilization-defective phenotype. It is proposed that PA4218 be named fptX (for “ferripyochelin transport”). RhtX and FptX appear to be members of a novel family of permeases that function as single-subunit transporters of siderophores.

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