Abstract
BackgroundCerebrospinal fluid (CSF) is an important source of potential biomarkers that affect the brain. Biomarkers for neurodegenerative disorders are needed to assist in diagnosis, monitoring disease progression and evaluating efficacy of therapies. Recent studies have demonstrated the involvement of tyrosine kinases in neuronal cell death. Thus, neurodegeneration in the brain is related to altered tyrosine phosphorylation of proteins in the brain and identification of abnormally phosphorylated tyrosine peptides in CSF has the potential to ascertain candidate biomarkers for neurodegenerative disorders.MethodsIn this study, we used an antibody-based tyrosine phosphopeptide enrichment method coupled with high resolution Orbitrap Fusion Tribrid Lumos Fourier transform mass spectrometer to catalog tyrosine phosphorylated peptides from cerebrospinal fluid. The subset of identified tyrosine phosphorylated peptides was also validated using parallel reaction monitoring (PRM)-based targeted approach.ResultsTo date, there are no published studies on global profiling of phosphotyrosine modifications of CSF proteins. We carried out phosphotyrosine profiling of CSF using an anti-phosphotyrosine antibody-based enrichment and analysis using high resolution Orbitrap Fusion Lumos mass spectrometer. We identified 111 phosphotyrosine peptides mapping to 66 proteins, which included 24 proteins which have not been identified in CSF previously. We then validated a set of 5 tyrosine phosphorylated peptides in an independent set of CSF samples from cognitively normal subjects, using a PRM-based targeted approach.ConclusionsThe findings from this deep phosphotyrosine profiling of CSF samples have the potential to identify novel disease-related phosphotyrosine-containing peptides in CSF.
Highlights
Cerebrospinal fluid (CSF) is an important source of potential biomarkers that affect the brain
Lumbar CSF samples The sample group consisted of subjects with normal pressure hydrocephalus (NPH) and cognitively normal individuals seen as part of the BIOCARD study [27]
LC–MS/MS analysis of tyrosine phosphorylated peptides in CSF The CSF proteins were digested with trypsin, phosphotyrosine peptides were enriched, and LC–MS/ MS analysis was conducted as described in Methods [25, 30, 31] (Fig. 1)
Summary
Cerebrospinal fluid (CSF) is an important source of potential biomarkers that affect the brain. Cerebrospinal fluid (CSF) is a colorless, clear liquid that surrounds the brain and spinal cord [1] and conventionally acts as a shock absorber in the central nervous system by maintaining electrolytes as well as acid–base balance [2] It plays a crucial role in neurodevelopment and waste clearance in addition. Heegaard et al in a previous study carried out phosphoproteomic profiling of cerebrospinal fluid using TiO2 based enrichment. They identified 56 novel phosphorylation sites on 38 proteins [19]. This lead to identification of 47 phosphopeptides corresponding to 24 proteins [21]. All earlier mass spectrometry studies were focused on global phosphorylation of CSF
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
