Pore-forming pyocin S5 utilizes the FptA ferripyochelin receptor to kill Pseudomonas aeruginosa.
Pyocins are toxic proteins produced by some strains of Pseudomonas aeruginosa that are lethal for related strains of the same species. Some soluble pyocins (S2, S3 and S4) were previously shown to use the pyoverdine siderophore receptors to enter the cell. The P. aeruginosa PAO1 pore-forming pyocin S5 encoding gene (PAO985) was cloned into the expression vector pET15b, and the affinity-purified protein product tested for its killing activity against different P. aeruginosa strains. The results, however, did not show any correlation with a specific ferripyoverdine receptor. To further identify the S5 receptor, transposon mutants were generated. Pooled mutants were exposed to pyocin S5 and the resistant colonies growing in the killing zone were selected. The majority of S5-resistant mutants had an insertion in the fptA gene encoding the receptor for the siderophore pyochelin. Complementation of an fptA transposon mutant with the P. aeruginosa fptA gene in trans restored the sensitivity to S5. In order to define the receptor-binding domain of pyocin S5, two hybrid pyocins were constructed containing different regions from pyocin S5 fused to the C-terminal translocation and DNase killing domains of pyocin S2. Only the protein containing amino acid residues 151 to 300 from S5 showed toxicity, indicating that the pyocin S5 receptor-binding domain is not at the N-terminus of the protein as in other S-type pyocins. Pyocin S5 was, however, unable to kill Burkholderia cenocepacia strains producing a ferripyochelin FptA receptor, nor was the B. cenocepacia fptA gene able to restore the sensitivity of the resistant fptA mutant P. aeruginosa strain.
Department of Genetics, Faculty of Agriculture, Assiut University, Assiut, Egypt.|VIB Department of Structural Biology, Department of Bioengineering Sciences, Research Group Microbiology, Vrije Universiteit Brussel, Pleinlaan 2, Brussels B-1050, Belgium.
Ameer
A
Elfarash
VIB Department of Structural Biology, Department of Bioengineering Sciences, Research Group Microbiology, Vrije Universiteit Brussel, Pleinlaan 2, Brussels B-1050, Belgium.
Jozef
J
Dingemans
VIB Department of Structural Biology, Department of Bioengineering Sciences, Research Group Microbiology, Vrije Universiteit Brussel, Pleinlaan 2, Brussels B-1050, Belgium.
Lumeng
L
Ye
VIB Department of Structural Biology, Department of Bioengineering Sciences, Research Group Microbiology, Vrije Universiteit Brussel, Pleinlaan 2, Brussels B-1050, Belgium.
Ahmed Amir
AA
Hassan
VIB Department of Structural Biology, Department of Bioengineering Sciences, Research Group Microbiology, Vrije Universiteit Brussel, Pleinlaan 2, Brussels B-1050, Belgium.
Michael
M
Craggs
Département de Microbiologie Fondamentale, Université de Lausanne, Lausanne, Switzerland.
Cornelia
C
Reimmann
Department of Infection and Immunity, School of Medicine, University of Sheffield, Beech Hill Road, Sheffield S10 2RX, UK.
Mark S
MS
Thomas
VIB Department of Structural Biology, Department of Bioengineering Sciences, Research Group Microbiology, Vrije Universiteit Brussel, Pleinlaan 2, Brussels B-1050, Belgium.
Pierre
P
Cornelis
D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism*; D057508:Burkholderia cenocepacia / Q000187:drug effects / Q000235:genetics; D004252:DNA Mutational Analysis; D004251:DNA Transposable Elements; D055786:Gene Knockout Techniques; D005816:Genetic Complementation Test; D050296:Microbial Viability / Q000187:drug effects*; D016254:Mutagenesis, Insertional; D025941:Protein Interaction Mapping; D011550:Pseudomonas aeruginosa / Q000187:drug effects* / Q000235:genetics / Q000502:physiology; D011709:Pyocins / Q000302:isolation & purification / Q000378:metabolism* / Q000633:toxicity*; D011956:Receptors, Cell Surface / Q000378:metabolism*
D001425:Bacterial Outer Membrane Proteins; D004251:DNA Transposable Elements; C075523:FPTA protein, Pseudomonas; D011709:Pyocins; D011956:Receptors, Cell Surface