Chlorination of bacterial and neutrophil proteins during phagocytosis and killing of Staphylococcus aureus.
Myeloperoxidase is proposed to play a central role in bacterial killing by generating hypochlorous acid within neutrophil phagosomes. However, it has yet to be demonstrated that these inflammatory cells target hypochlorous acid against bacteria inside phagosomes. In this investigation, we treated Staphylococcus aureus with varying concentrations of reagent hypochlorous acid and found that even at sublethal doses, it converted some tyrosine residues in their proteins to 3-chlorotyrosine and 3,5-dichlorotyrosine. To determine whether or not ingested bacteria were exposed to hypochlorous acid in neutrophil phagosomes, we labeled their proteins with [(13)C(6)]tyrosine and used gas chromatography with mass spectrometry to identify the corresponding chlorinated isotopes after the bacteria had been phagocytosed. Chlorinated tyrosines were detected in bacterial proteins 5 min after phagocytosis and reached levels of approximately 2.5/1000 mol of tyrosine at 60 min. Inhibitor studies revealed that chlorination was dependent on myeloperoxidase. Chlorinated neutrophil proteins were also detected and accounted for 94% of total chlorinated tyrosine residues formed during phagocytosis. We conclude that hypochlorous acid is a major intracellular product of the respiratory burst. Although some reacts with the bacteria, most reacts with neutrophil components.
Chapman
Anna L P
AL
Free Radical Research Group, Department of Pathology, Christchurch School of Medicine and Health Sciences, Post Office Box 4345, Christchurch, New Zealand.
Hampton
Mark B
MB
Senthilmohan
Revathy
R
Winterbourn
Christine C
CC
Kettle
Anthony J
AJ
D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002713:Chlorine / Q000378:metabolism; D002849:Chromatography, Gas; D004305:Dose-Response Relationship, Drug; D006997:Hypochlorous Acid / Q000494:pharmacology*; D013058:Mass Spectrometry; D009504:Neutrophils / Q000737:chemistry / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology; D009195:Peroxidase / Q000378:metabolism*; D010587:Phagocytosis; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013211:Staphylococcus aureus / Q000187:drug effects / Q000378:metabolism*; D013696:Temperature; D013997:Time Factors; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism*
D000900:Anti-Bacterial Agents; D016877:Oxidants; D014443:Tyrosine; D002713:Chlorine; D006997:Hypochlorous Acid; D009195:Peroxidase