Physiological Characterization of a Heme-Deficient Mutant of Staphylococcus aureus by a Proteomic Approach
AUTOR(ES)
Kohler, Christian
FONTE
American Society for Microbiology
RESUMO
The high-resolution two-dimensional (2D) protein gel electrophoresis technique combined with matrix-assisted laser desorption ionization-time of flight mass spectrometry was used for identification of proteins whose levels were changed by a mutation in hemB. Cytoplasmic protein extracts obtained from the mutant and the wild type (strain COL) at different stages of growth in tryptone soya broth (exponential, transitional, and stationary growth phases) were separated on 2D protein gels. Comparison of the 2D patterns of the protein extracts of the two strains revealed major differences. Because the electron transport chain of the mutant is interrupted due to the deficiency of heme, this organism should be unable to use oxygen or nitrate as a terminal electron acceptor. Consistent with this hypothesis, proteins involved in the glycolytic pathway and related pathways (glyceraldehyde-3-phosphate dehydrogenase, enolase, and phosphoglycerate kinase) and in fermentation pathways (lactate dehydrogenase, alcohol dehydrogenase, and pyruvate formate lyase) were induced in exponentially growing cells of the mutant. These results strongly indicate that the hemB mutant generates ATP from glucose or fructose only by substrate phosphorylation. Analyses of the fermentation reactions showed that the main product was lactate. Although pyruvate formate lyase (Pfl) and pyruvate dehydrogenase were present, neither ethanol nor acetate was detected in significant amounts. Presumably, Pfl was not activated in the presence of oxygen, and pyruvate dehydrogenase might have very low activity. Transcriptional analysis of citB, encoding the aconitase, revealed that the activity of the citrate cycle enzymes was down-regulated in the hemB mutant. The arginine deiminase pathway was also induced, and it could provide ATP as well. Furthermore, the amounts of most of the extracellular virulence factors were significantly reduced by a mutation in hemB, which is consistent with previous reports.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=262702Documentos Relacionados
- Nitrate reductase activity in heme-deficient mutants of Staphylococcus aureus.
- Fate of reversing factor during restoration of protein synthesis by hemin or GTP in heme-deficient reticulocyte lysates.
- Roles of a 67-kDa polypeptide in reversal of protein synthesis inhibition in heme-deficient reticulocyte lysate.
- The role of hemoglobin heme loss in Heinz body formation: studies with a partially heme-deficient hemoglobin and with genetically unstable hemoglobins
- Protein synthesis in rabbit reticulocytes: characteristics of a ribosomal factor that reverses inhibition of protein synthesis in heme-deficient lysates.