Proteomics analysis of phosphotyrosyl-proteins in human lumbar cerebrospinal fluid.
Cerebrospinal fluid (CSF) is a secretion product of several different central nervous system (CNS) structures, including the choroid plexus in the ventricles. Pathological CNS processes are reflected in the protein composition of CSF. To elucidate the molecular events that occur in the homeostatic and pathological processes of the CNS, the high-throughput characterization of differentially expressed proteins, and of post-translationally modified proteins, is needed for proteomics studies of CSF. Among the post-translational modifications of proteins, phosphorylation is the most common and important mechanism for the reversible regulation of protein function. In this study, CSF phosphotyrosyl (p-Tyr)-proteins were detected with antibodies and were analyzed with proteomics methods. Three different combination methods--1D gel electrophoresis and Western blotting, immunoprecipitation and 2D gel electrophoresis, and 2D gel electrophoresis and Western blotting--were used to detect p-Tyr-proteins in human lumbar CSF samples. Six protein spots, representing four proteins on a 2D Western blot, were identified as p-Tyr-proteins with the 2D gel electrophoresis and Western blotting method. Those four p-Tyr-proteins are kallikrein-6 precursor, complement C4 gamma-chain, gelsolin, and ceruloplasmin precursor. Additionally, four other nonphosphorylated CSF proteins--beta-2-glycoprotein I precursor, fibulin-1 precursor, EGF-containing fibulin-like extracellur matrix protein 1 precursor, and angiotensinogen precursor--were characterized for the first time.
Charles B. Stout Neuroscience Mass Spectrometry Laboratory, Department of Neurology, University of Tennessee Health Science Center, 847 Monroe Avenue, Memphis, Tennessee 38163, USA.
Xianglin
X
Yuan
Dominic M
DM
Desiderio
D015153:Blotting, Western; D002556:Cerebrospinal Fluid Proteins / Q000032:analysis*; D002853:Chromatography, Liquid; D030562:Databases, Protein; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D013058:Mass Spectrometry; D010766:Phosphorylation; D011233:Precipitin Tests; D011506:Proteins / Q000032:analysis*; D040901:Proteomics / Q000379:methods*; D016622:Silver Staining; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D002556:Cerebrospinal Fluid Proteins; D011506:Proteins