Characterization of a
            ferric‐binding protein
            mutant in
            Haemophilus influenzae

ABSTRACT Horizontal gene transfer has been identified in only a small number of genes in Haemophilus influenzae, an organism which is naturally competent for transformation. This report provides evidence for the genetic transfer of the ftsI gene, which encodes penicillin-binding protein 3, in H. influenzae. Mosaic structures of the ftsI gene were found in several clinical isolates of H. influenzae. To identify the origin of the mosaic sequence, complete sequences of the corresponding gene from seven type strains of Haemophilus species were determined. Comparison of these sequences with mosaic regions identified a homologous recombination of the ftsI gene between H. influenzae and Haemophilus haemolyticus. Subsequently, ampicillin-resistant H. influenzae strains harboring identical ftsI sequences were genotyped by pulsed-field gel electrophoresis (PFGE). Divergent PFGE patterns among β-lactamase-nonproducing ampicillin-resistant (BLNAR) strains from different hospitals indicated the potential for the genetic transfer of the mutated ftsI gene between these isolates. Moreover, transfer of the ftsI gene from BLNAR strains to β-lactamase-nonproducing ampicillin-susceptible (BLNAS) H. influenzae strains was evaluated in vitro. Coincubation of a BLNAS strain (a rifampin-resistant mutant of strain Rd) and BLNAR strains resulted in the emergence of rifampin- and cefdinir-resistant clones at frequencies of 5.1 × 10−7 to 1.5 × 10−6. Characterization of these doubly resistant mutants by DNA sequencing of the ftsI gene, susceptibility testing, and genotyping by PFGE revealed that the ftsI genes of BLNAR strains had transferred to BLNAS strains during coincubation. In conclusion, horizontal transfer of the ftsI gene in H. influenzae can occur in an intraspecies and an interspecies manner.

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Functional characterization of the DNA mismatch binding protein MutS from Haemophilus influenzae

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2005.06.178 ◽

2005 ◽

Vol 334 (3) ◽

pp. 891-900 ◽

Cited By ~ 7

Author(s):

Nimesh Joseph ◽

Viswanadham Duppatla ◽

Desirazu N. Rao

Keyword(s):

Haemophilus Influenzae ◽

Binding Protein ◽

Functional Characterization ◽

Dna Mismatch

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DNA-binding proteins of Haemophilus influenzae: purification and characterization of a major intracellular binding protein.

Journal of Bacteriology ◽

10.1128/jb.155.1.246-253.1983 ◽

1983 ◽

Vol 155 (1) ◽

pp. 246-253 ◽

Cited By ~ 1

Author(s):

S L Sutrina ◽

J J Scocca

Keyword(s):

Dna Binding ◽

Haemophilus Influenzae ◽

Binding Proteins ◽

Binding Protein ◽

Dna Binding Proteins ◽

Purification And Characterization ◽

Intracellular Binding

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The role of vicinal tyrosine residues in the function of Haemophilus influenzae ferric-binding protein A

Biochemical Journal ◽

10.1042/bj20101043 ◽

2010 ◽

Vol 432 (1) ◽

pp. 57-67 ◽

Cited By ~ 13

Author(s):

Husain K. Khambati ◽

Trevor F. Moraes ◽

Jagroop Singh ◽

Stephen R. Shouldice ◽

Rong-hua Yu ◽

...

Keyword(s):

Haemophilus Influenzae ◽

Outer Membrane ◽

Binding Protein ◽

Protein A ◽

Membrane Receptor ◽

Receptor Complex ◽

Iron Binding ◽

Outer Membrane Receptor ◽

Tyrosine Residues ◽

Ferric Binding Protein

The periplasmic FbpA (ferric-binding protein A) from Haemophilus influenzae plays a critical role in acquiring iron from host transferrin, shuttling iron from the outer-membrane receptor complex to the inner-membrane transport complex responsible for transporting iron into the cytoplasm. In the present study, we report on the properties of a series of site-directed mutants of two adjacent tyrosine residues involved in iron co-ordination, and demonstrate that, in contrast with mutation of equivalent residues in the N-lobe of human transferrin, the mutant FbpAs retain significant iron-binding affinity regardless of the nature of the replacement amino acid. The Y195A and Y196A FbpAs are not only capable of binding iron, but are proficient in mediating periplasm-to-cytoplasm iron transport in a reconstituted FbpABC pathway in a specialized Escherichia coli reporter strain. This indicates that their inability to mediate iron acquisition from transferrin is due to their inability to compete for iron with receptor-bound transferrin. Wild-type iron-loaded FbpA could be crystalized in a closed or open state depending upon the crystallization conditions. The synergistic phosphate anion was not present in the iron-loaded open form, suggesting that initial anchoring of iron was mediated by the adjacent tyrosine residues and that alternate pathways for iron and anion binding and release may be considered. Collectively, these results demonstrate that the presence of a twin-tyrosine motif common to many periplasmic iron-binding proteins is critical for initially capturing the ferric ion released by the outer-membrane receptor complex.

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