The formation of
reactive oxygen species (ROS) is linked to the
pathogenesis of neurodegenerative diseases. Here we have investigated
the effect of soluble and aggregated amyloid-β (Aβ) and
α-synuclein (αS), associated with Alzheimer’s and
Parkinson’s diseases, respectively, on the Cu2+-catalyzed
formation of ROS in vitro in the presence of a biological
reductant. We find that the levels of ROS, and the rate by which ROS
is generated, are significantly reduced when Cu2+ is bound
to Aβ or αS, particularly when they are in their oligomeric
or fibrillar forms. This effect is attributed to a combination of
radical scavenging and redox silencing mechanisms. Our findings suggest
that the increase in ROS associated with the accumulation of aggregated
Aβ or αS does not result from a particularly ROS-active
form of these peptides, but rather from either a local increase of
Cu2+ and other ROS-active metal ions in the aggregates
or as a downstream consequence of the formation of the pathological
amyloid structures.
Biomarkers in the cerebrospinal fluid (CSF) may be important for the diagnosis of chronic degenerative disorders in the central nervous system including dementia. Existing CSF biomarkers for dementia, however, are relatively nonspecific. More specific markers may be found by targeting investigations based on knowledge of the molecular pathology of the disease in question. In Alzheimer's disease, hyperphosphorylation of the tau protein is a characteristic feature and thus a comprehensive characterization of the phosphoproteome of the CSF may be pursued to obtain a complete picture of phosphorylation aberrations in health and disease. Toward that goal we here describe a method for a comprehensive isolation and identification of phosphorylated tryptic peptides derived from CSF proteins using a simple sample preparation step and titanium dioxide-affinity chromatography followed by MALDI-TOF or LC-MS/MS linear ion-trap-FT mass spectrometry. Whereas not all previously reported phosphoproteins were found in normal CSF, we detected 56 putative novel phosphorylation sites in 38 proteins in addition to known sites. The approach seems to be a promising foundation for the discovery of new biomarkers embedded in the CSF phosphoproteome.
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