Identification of an outer membrane protein involved in utilization of hemoglobin-haptoglobin complexes by nontypeable Haemophilus influenzae

Isobel Maciver, Jo L. Latimer, H. H. Liem, Ursula Muller-Eberhard, Zbynek Hrkal, Eric J. Hansen

Research output: Contribution to journalArticle

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Abstract

A recombinant plasmid containing a 6.5-kb fragment of nontypeable Haemophilus influenzae (NTHI) chromosomal DNA was shown to confer a hemoglobin-haptoglobin-binding phenotype on Escherichia coli. Use of a mini- Tn10kan transposon for random insertion mutagenesis of this recombinant plasmid allowed localization of the NTHI DNA responsible for this hemoglobin- haptoglobin-binding phenotype to a 3.5-kb PstI-XhoI fragment within the 6.5- kb NTHI DNA insert. When this mutagenized NTHI DNA fragment was used to transform the wild-type NTHI strain, the resultant kanamycin-resistant mutant exhibited significantly decreased abilities to bind hemoglobin-haptoglobin and utilize it as a source of heme for aerobic growth in vitro. This mutant also lacked expression of a 115-kDa outer membrane protein that was present in the wild-type parent strain. Transformation of this mutant with wild-type NTHI chromosomal DNA restored the abilities to bind and utilize hemoglobin- haptoglobin and to express the 115-kDa outer membrane protein. Nucleotide sequence analysis of the relevant NTHI DNA revealed the presence of a gene, designated hhuA, that encoded a predicted 117,145-Da protein. The HhuA protein exhibited features typical of a TonB-dependent outer membrane receptor and had significant identity with the hemoglobin receptors of both Haemophilus ducreyi and Neisseria meningitidis.

Original languageEnglish (US)
Pages (from-to)3703-3712
Number of pages10
JournalInfection and Immunity
Volume64
Issue number9
StatePublished - 1996

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Haemophilus influenzae
Membrane Proteins
Haptoglobins
Hemoglobins
DNA
Plasmids
Haemophilus ducreyi
Phenotype
Kanamycin
Neisseria meningitidis
haptoglobin-hemoglobin complex
Heme
Mutagenesis
Sequence Analysis
Proteins
Escherichia coli
Membranes
Growth
Genes

ASJC Scopus subject areas

  • Immunology

Cite this

Identification of an outer membrane protein involved in utilization of hemoglobin-haptoglobin complexes by nontypeable Haemophilus influenzae. / Maciver, Isobel; Latimer, Jo L.; Liem, H. H.; Muller-Eberhard, Ursula; Hrkal, Zbynek; Hansen, Eric J.

In: Infection and Immunity, Vol. 64, No. 9, 1996, p. 3703-3712.

Research output: Contribution to journalArticle

Maciver, Isobel ; Latimer, Jo L. ; Liem, H. H. ; Muller-Eberhard, Ursula ; Hrkal, Zbynek ; Hansen, Eric J. / Identification of an outer membrane protein involved in utilization of hemoglobin-haptoglobin complexes by nontypeable Haemophilus influenzae. In: Infection and Immunity. 1996 ; Vol. 64, No. 9. pp. 3703-3712.
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