The Pasteurella haemolytica 35 kDa iron-regulated protein is an FbpA homologue.
In a previous investigation, a 35 kDa iron-regulated protein was identified from total cellular proteins of Pasteurella haemolytica grown under iron-depleted conditions. This study reports identification of the gene (fbpA) encoding the 35 kDa protein based on complementation of an entA Escherichia coli strain transformed with a plasmid derived from a P. haemolytica lambda ZAP II library. Cross-reactivity was demonstrated between an anti-35 kDa mAb and a 35 kDa protein expressed in this strain. Furthermore, a translated ORF identified on the recombinant plasmid corresponded with the N-terminal amino acid sequence of the intact and a CNBr-cleaved fragment of the 35 kDa iron-regulated protein. Nucleotide sequence analysis of the gene encoding the 35 kDa protein demonstrated homology with the cluster 1 group of extracellular solute-binding proteins, especially to the iron-binding proteins of this family. Complete sequence analysis of the recombinant plasmid insert identified three other predominant ORFs, two of which appeared to be in an operonic organization with fbpA. These latter components (fbpB and fbpC) showed homology to the transmembrane and ATPase components of ATP-binding cassette (ABC)-type uptake systems, respectively. Based on amino acid/DNA sequencing, citrate competition assay of iron affinity and visible wavelength spectra, it was concluded that the P. haemolytica 35 kDa protein functions as an FbpA homologue (referred to as PFbpA) and that the gene encoding this protein is part of an operon comprising a member of the FbpABC family of iron uptake systems. Primary sequence analysis revealed rather surprisingly that PFbpA is more closely related to the intracellular Mn/Fe-binding protein IdiA found in cyanobacteria than to any of the homologous FbpA proteins currently known in commensal or pathogenic members of the Pasteurellaceae or Neisseriaceae.
Kirby
S D
SD
Department of Microbiology and Infectious Diseases, University of Calgary,Calgary, Alberta,Canada T2N 4N1.
Lainson
F A
FA
Moredun Research Institute, International Research Centre,Pentlands Science Park, Bush Loan, Penicuik, Midlothian EH26 0PZ,UK.
Donachie
W
W
Moredun Research Institute, International Research Centre,Pentlands Science Park, Bush Loan, Penicuik, Midlothian EH26 0PZ,UK.
Okabe
A
A
Department of Microbiology3and First Department of Physiology.
Tokuda
M
M
Kagawa Medical University, Kita-gun, Kagawa, 76107,Japan.
Hatase
O
O
Kagawa Medical University, Kita-gun, Kagawa, 76107,Japan.
Schryvers
A B
AB
Department of Microbiology and Infectious Diseases, University of Calgary,Calgary, Alberta,Canada T2N 4N1.
D000595:Amino Acid Sequence; D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D004269:DNA, Bacterial / Q000032:analysis; D007501:Iron / Q000378:metabolism*; D033862:Iron-Binding Proteins; D016978:Mannheimia haemolytica / Q000145:classification / Q000235:genetics*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009876:Operon; D033902:Periplasmic Binding Proteins; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D004269:DNA, Bacterial; D033862:Iron-Binding Proteins; D033902:Periplasmic Binding Proteins; D007501:Iron