A novel post-translational incorporation of tyrosine into multiple proteins in activated human neutrophils. Correlation with phagocytosis and activation of the NADPH oxidase-mediated respiratory burst.
Activation of human neutrophils by PMA causes a post-translational incorporation of 14C-labeled tyrosine into multiple neutrophil (PMN) proteins, that is distinctly different from the enzymatic tyrosinolation of tubulin in FMLP-stimulated PMN. Post-translational incorporation of other radiolabeled amino acids, including the structurally similar amino acid phenylalanine, does not occur under identical conditions of neutrophil activation, suggesting an involvement of the phenolic hydroxyl group of tyrosine in the PMA-mediated reaction. Similar to the stimulation of PMN tubulin tyrosinolation by FMLP, the PMA-induced incorporation of tyrosine into multiple PMN proteins is closely associated with activation of the NADPH oxidase-mediated respiratory burst in stimulated PMN and can be inhibited by a variety of reducing agents, inhibitors of peroxidase-mediated reactions, and intracellular scavengers of oxygen radicals. Moreover, the PMA-induced post-translational incorporation of tyrosine does not occur in PMN from patients with chronic granulomatous disease and is significantly reduced (50%) in PMN of an individual with myeloperoxidase deficiency. A similar stimulus-induced incorporation of tyrosine into multiple PMN proteins is also observed in PMN exposed to various phagocytic stimuli, and the incorporated radioactivity in cells undergoing phagocytosis is substantially enriched (40- to 50-fold) in isolated PMN phagolysosomes. Consistent with this latter observation, HPLC fractionation of stimulated PMN proteins and analysis of the incorporated radioactivity reveal that the 14C label is primarily associated with PMN membrane proteins. Furthermore, this post-translational incorporation of tyrosine, like that associated with PMA stimulation, is associated with production of oxygen radicals and the generation of protein carbonyl derivatives, which are indicative of oxidative protein modifications via mixed function oxidases. Our findings indicate that tyrosine incorporation into membrane proteins of stimulated PMN is functionally relevant to the physiologic host-defense responses of human neutrophils undergoing phagocytosis.
Nath
J
J
Department of Hematology, Walter Reed Army Institute of Research, Washington, DC 20307.
Ohno
Y
Y
Gallin
J I
JI
Wright
D G
DG
D001798:Blood Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D004492:Edetic Acid / Q000494:pharmacology; D016166:Free Radical Scavengers; D006105:Granulomatous Disease, Chronic / Q000378:metabolism; D006801:Humans; D009247:NADH, NADPH Oxidoreductases / Q000502:physiology*; D019255:NADPH Oxidases; D009504:Neutrophils / Q000276:immunology / Q000378:metabolism*; D009195:Peroxidase / Q000172:deficiency; D010587:Phagocytosis; D010588:Phagosomes / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D016897:Respiratory Burst; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014443:Tyrosine / Q000378:metabolism*
D001798:Blood Proteins; D016166:Free Radical Scavengers; D014443:Tyrosine; D004492:Edetic Acid; D009195:Peroxidase; D009247:NADH, NADPH Oxidoreductases; D019255:NADPH Oxidases; D013755:Tetradecanoylphorbol Acetate