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Molecular constituents of the postsynaptic density fraction revealed by proteomic analysis using multidimensional liquid chromatography‐tandem mass spectrometry
Author(s) -
Yoshimura Yoshiyuki,
Yamauchi Yoshio,
Shinkawa Takashi,
Taoka Masato,
Donai Hitomi,
Takahashi Nobuhiro,
Isobe Toshiaki,
Yamauchi Takashi
Publication year - 2004
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.2003.02136.x
Subject(s) - postsynaptic density , chemistry , protein mass spectrometry , proteomics , bottom up proteomics , tandem mass spectrometry , electrospray ionization , biochemistry , membrane protein , mass spectrometry , chromatography , postsynaptic potential , receptor , membrane , gene
Protein constituents of the postsynaptic density (PSD) fraction were analysed using an integrated liquid chromatography (LC)‐based protein identification system, which was constructed by coupling microscale two‐dimensional liquid chromatography (2DLC) with electrospray ionization (ESI) tandem mass spectrometry (MS/MS) and an automated data analysis system. The PSD fraction prepared from rat forebrain was solubilized in 6 m guanidium hydrochloride, and the proteins were digested with trypsin after S‐carbamoylmethylation under reducing conditions. The tryptic peptide mixture was then analysed with the 2DLC‐MS/MS system in a data‐dependent mode, and the resultant spectral data were automatically processed to search a genome sequence database for protein identification. In triplicate analyses, the system allowed assignments of 5264 peptides, which could finally be attributed to 492 proteins. The PSD contained various proteins involved in signalling transduction, including receptors, ion channel proteins, protein kinases and phosphatases, G‐protein and related proteins, scaffold proteins, and adaptor proteins. Structural proteins, including membrane proteins involved in cell adhesion and cell–cell interaction, proteins involved in endocytosis, motor proteins, and cytoskeletal proteins were also abundant. These results provide basic data on a major protein set associated with the PSD and a basis for future functional studies of this important neural machinery.