Lack of expression of the global regulator OxyR in Haemophilus influenzae has a profound effect on growth phenotype

Isobel Maciver, Eric J. Hansen

Research output: Contribution to journalArticle

39 Scopus citations

Abstract

A pBR322-based library of chromosomal DNA from the nontypeable Haemophilus influenzae TN106 was screened for the expression of transferrin- binding activity in Escherichia coli. A recombinant clone expressing transferrin-binding activity contained a 3.7-kb fragment of nontypeable H. influenzae DNA. Nucleotide sequence analysis of this insert revealed the presence of two complete open reading frames encoding proteins of approximately 26 and 34 kDa. Mini-Tn10kan transposon mutagenesis at different sites within the open reading frame encoding the 34-kDa protein resulted in the abolition of transferrin-binding activity in the recombinant E. coli clone. The deduced amino acid sequence of the 34-kDa protein had 70% identity with the OxyR protein of E. coli; this latter macromolecule is a member of the LysR family of transcriptional activators. When a mutated H. influenzae oxyR gene was introduced into the chromosome of the wild-type H. influenzae strain by allelic exchange, the resulting oxyR mutant still exhibited wild- type levels of transferrin-binding activity but was unable to grow on media containing the heme precursor protoporphyrin IX (PPIX) in place of heme. This mutant also exhibited reduced growth around disks impregnated with heme sources. Supplementation of the PPIX-based growth media with catalase or sodium pyruvate resulted in normal growth of the H. influenzae oxyR mutant. Provision of the wild-type H. influenzae oxyR gene in trans also permitted the growth of this mutant on a PPIX-based medium. Exogenously supplied catalase restored the growth of this mutant with heme sources to nearly wild- type levels. These results indicate that expression of a wild-type OxyR protein by H. influenzae is essential to allow this organism to protect itself against oxidative stresses in vitro.

Original languageEnglish (US)
Pages (from-to)4618-4629
Number of pages12
JournalInfection and immunity
Volume64
Issue number11
StatePublished - Nov 5 1996

ASJC Scopus subject areas

  • Parasitology
  • Microbiology
  • Immunology
  • Infectious Diseases

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