Identification of type II and type III pyoverdine receptors from Pseudomonas aeruginosa.
Pseudomonas aeruginosa produces, under conditions of iron limitation, a high-affinity siderophore, pyoverdine (PVD), which is recognized at the level of the outer membrane by a specific TonB-dependent receptor, FpvA. So far, for P. aeruginosa, three different PVDs, differing in their peptide chain, have been described (types I-III), but only the FpvA receptor for type I is known. Two PVD-producing P. aeruginosa strains, one type II and one type III, were mutagenized by a mini-TnphoA3 transposon. In each case, one mutant unable to grow in the presence of the strong iron chelator ethylenediaminedihydroxyphenylacetic acid (EDDHA) and the cognate PVD was selected. The first mutant, which had an insertion in the pvdE gene, upstream of fpvA, was unable to take up type II PVD and showed resistance to pyocin S3, which is known to use type II FpvA as receptor. The second mutant was unable to take up type III PVD and had the transposon insertion in fpvA. Cosmid libraries of the respective type II and type III PVD wild-type strains were constructed and screened for clones restoring the capacity to grow in the presence of PVD. From the respective complementing genomic fragments, type II and type III fpvA sequences were determined. When in trans, type II and type III fpvA restored PVD production, uptake, growth in the presence of EDDHA and, in the case of type II fpvA, pyocin S3 sensitivity. Complementation of fpvA mutants obtained by allelic exchange was achieved by the presence of cognate fpvA in trans. All three receptors posses an N-terminal extension of about 70 amino acids, similar to FecA of Escherichia coli, but only FpvAI has a TAT export sequence at its N-terminal end.
de Chial
Magaly
M
Flanders Interuniversity Institute of Biotechnology (VIB6), Laboratory of Microbial Interactions, Vrije Universiteit Brussel, Building E, room 6.6, Pleinlaan 2, B-1050 Brussels, Belgium.
Ghysels
Bart
B
Flanders Interuniversity Institute of Biotechnology (VIB6), Laboratory of Microbial Interactions, Vrije Universiteit Brussel, Building E, room 6.6, Pleinlaan 2, B-1050 Brussels, Belgium.
Beatson
Scott A
SA
MRC Functional Genetics Unit, Department of Human Anatomy and Genetics, University of Oxford, South Parks Road, Oxford OX1 3QX, UK.
Geoffroy
Valérie
V
Laboratoire de Microbiologie et de Génétique, Université Louis Pasteur, UPRES-A 7010, F-67000 Strasbourg, France.
Meyer
Jean Marie
JM
Laboratoire de Microbiologie et de Génétique, Université Louis Pasteur, UPRES-A 7010, F-67000 Strasbourg, France.
Pattery
Theresa
T
Flanders Interuniversity Institute of Biotechnology (VIB6), Laboratory of Microbial Interactions, Vrije Universiteit Brussel, Building E, room 6.6, Pleinlaan 2, B-1050 Brussels, Belgium.
Baysse
Christine
C
Flanders Interuniversity Institute of Biotechnology (VIB6), Laboratory of Microbial Interactions, Vrije Universiteit Brussel, Building E, room 6.6, Pleinlaan 2, B-1050 Brussels, Belgium.
Chablain
Patrice
P
Flanders Interuniversity Institute of Biotechnology (VIB6), Laboratory of Microbial Interactions, Vrije Universiteit Brussel, Building E, room 6.6, Pleinlaan 2, B-1050 Brussels, Belgium.
Parsons
Yasmin N
YN
Department of Medical Microbiology and Genitourinary Medicine, University of Liverpool, Duncan Building, Liverpool L69 3GA, UK.
Winstanley
Craig
C
Department of Medical Microbiology and Genitourinary Medicine, University of Liverpool, Duncan Building, Liverpool L69 3GA, UK.
Cordwell
Stuart J
SJ
Australian Proteome Analysis Facility, Sydney, Australia 2109.
Cornelis
Pierre
P
Flanders Interuniversity Institute of Biotechnology (VIB6), Laboratory of Microbial Interactions, Vrije Universiteit Brussel, Building E, room 6.6, Pleinlaan 2, B-1050 Brussels, Belgium.
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D003470:Culture Media; D003550:Cystic Fibrosis / Q000382:microbiology; D005029:Ethylenediamines; D005816:Genetic Complementation Test; D006801:Humans; D007501:Iron / Q000378:metabolism; D008969:Molecular Sequence Data; D009154:Mutation; D009842:Oligopeptides; D010860:Pigments, Biological / Q000378:metabolism*; D011552:Pseudomonas Infections / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000187:drug effects / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D011709:Pyocins / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017422:Sequence Analysis, DNA
D001425:Bacterial Outer Membrane Proteins; D003470:Culture Media; D005029:Ethylenediamines; C082326:FpvA protein, Pseudomonas aeruginosa; D009842:Oligopeptides; D010860:Pigments, Biological; D011709:Pyocins; D011956:Receptors, Cell Surface; C011813:ethylenediamine-N,N'-bis(2-hydroxyphenylacetic acid); C042453:pyoverdin; D007501:Iron