The 100 kDa haem:haemopexin‐binding protein of Haemophilus Influenzae: structure and localization

Leslie D. Cope, Sharon E. Thomas, Jo L. Latimer, Clive A. Slaughter, Ursula Müller‐Eberhard, Eric J. Hansen

Research output: Contribution to journalArticle

56 Citations (Scopus)

Abstract

All Haemophilus influenzae strains have an absolute requirement for exogenously supplied haem for aerobic growth. A majority of strains of H. influenzae type b (Hib) produce a 100 kDa protein which binds haem: haemopexin complexes. This 100 kDa haem:haemopexin binding protein, designated HxuA, was originally detected on the Hib cell surface. Monoclonal antibody (mAb)‐based analyses revealed that the HxuA protein was also present in soluble form in Hib culture supernatants. This soluble HxuA protein exhibited haem:haemopexin‐binding activity in a direct binding assay. Nucleotide sequence analysis of the hxuA gene from Hib strain DL42, together with N‐terminal amino acid analysis of HxuA protein purified from Hib culture supernatant, revealed that this protein was synthesized as a 101 kDa precursor with a leader peptide that was removed to yield a 99kDa protein. Southern blot analysis of chromosomal DNA from four Hib and four non‐typeable H. influenzae (NTHI) strains detected the presence of a single band in each strain that hybridized a Hib hxuA gene probe. Subsequent analysis of these NTHI strains showed that all four strains released into culture supernatant a haem:haemopexin‐binding protein that migrated in SDS‐PAGE at a rate similar or identical to that of the Hib HxuA protein. A Hib hxuA mutant was used to screen an NTHI genomic DNA library and an NTHI gene was cloned that complemented the mutation in this Hib strain. Nucleotide sequence analysis of this NTHI gene revealed that it encoded a protein with 87% identity to the Hib HxuA protein. The expression of HxuA by both Hib and NTHI strains indicates that this particular haem acquisition system is conserved among H. influenzae strains.

Original languageEnglish (US)
Pages (from-to)863-873
Number of pages11
JournalMolecular Microbiology
Volume13
Issue number5
DOIs
StatePublished - Jan 1 1994
Externally publishedYes

Fingerprint

Haemophilus influenzae type b
Haemophilus influenzae
Heme
Proteins
Hemopexin
Genes
Sequence Analysis
Genomic Library
Protein Sorting Signals
Southern Blotting
Gene Library
Carrier Proteins

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

Cite this

The 100 kDa haem:haemopexin‐binding protein of Haemophilus Influenzae : structure and localization. / Cope, Leslie D.; Thomas, Sharon E.; Latimer, Jo L.; Slaughter, Clive A.; Müller‐Eberhard, Ursula; Hansen, Eric J.

In: Molecular Microbiology, Vol. 13, No. 5, 01.01.1994, p. 863-873.

Research output: Contribution to journalArticle

Cope, Leslie D. ; Thomas, Sharon E. ; Latimer, Jo L. ; Slaughter, Clive A. ; Müller‐Eberhard, Ursula ; Hansen, Eric J. / The 100 kDa haem:haemopexin‐binding protein of Haemophilus Influenzae : structure and localization. In: Molecular Microbiology. 1994 ; Vol. 13, No. 5. pp. 863-873.
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