The impact of specific oxidized amino acids on protein turnover in J774 cells.
Oxidized protein deposition and accumulation have been implicated in the aetiology of a wide variety of age-related pathologies. Protein oxidation in vivo commonly results in the in situ modification of amino acid side chains, generating new oxidized amino acid residues in proteins. We have demonstrated previously that certain oxidized amino acids can be (mis)incorporated into cell proteins in vitro via protein synthesis. In the present study, we show that incorporation of o- and m-tyrosine resulted in increased protein catabolism, whereas dopa incorporation generated proteins that were inefficiently degraded by cells. Incorporation of higher levels of L-dopa into proteins resulted in an increase in the activity of lysosomal cathepsins, increased autofluorescence and the generation of high-molecular-mass SDS-stable complexes, indicative of protein aggregation. These effects were due to proteins containing incorporated L-dopa, since they were not seen with the stereoisomer D-dopa, which enters the cell and generates the same reactive species as L-dopa, but cannot be incorporated into proteins. The present study highlights how the nature of the oxidative modification to the protein can determine the efficiency of its removal from the cell by proteolysis. Protection against the generation of dopa and other species that promote resistance to proteolysis might prove to be critical in preventing toxicity from oxidative stress in pathologies associated with protein deposition, such as atherosclerosis, Alzheimer's disease and Parkinson's disease.
Dunlop
Rachael A
RA
The Cell Biology Research Group, Heart Research Institute, 114 Pyrmont Bridge Road, Camperdown, NSW 2050, Australia. dunlopr@hri.org.au
Dean
Roger T
RT
Rodgers
Kenneth J
KJ
D000596:Amino Acids / Q000378:metabolism*; D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002851:Chromatography, High Pressure Liquid; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D006868:Hydrolysis; D051379:Mice; D010084:Oxidation-Reduction; D011506:Proteins / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D000596:Amino Acids; D017931:DNA Primers; D011506:Proteins