Phosphorylation state of postsynaptic density proteins.
The postsynaptic density (PSD) is an electron-dense structure located at the synaptic contacts between neurons. Its considerable complexity includes cytoskeletal and scaffold proteins, receptors, ion channels and signaling molecules, in line with the role of PSDs in signal transduction and processing. The phosphorylation state of components of the PSD is central to synaptic transmission and is known to play a role in synaptic plasticity, learning and memory. The presence of a range of kinases and phosphatases in the PSD defines potential key players in this context. However, the substrates that these enzymes target have not been fully identified to date. We analyzed the protein composition of purified PSD samples from adult mouse brains by strong cation exchange chromatography fractionation of a tryptic digest followed by nano-reverse phase liquid chromatography coupled with electrospray ionization-quadrupole time of flight tandem mass spectrometry. This led to the identification of 244 proteins. To gain an insight into the phosphoproteome of the PSD we then purified phosphorylated tryptic peptides by immobilized metal ion affinity chromatography. This approach for the specific enrichment of phosphopeptides resulted in the identification of 42 phosphoproteins in the PSD preparation, 39 of which are known PSD components. Here we present a total of 83 in vivo phosphorylation sites.
Trinidad
J C
JC
Mass Spectrometry Facility, Department of Pharmaceutical Chemistry, University of California, 513 Parnassus Avenue, San Francisco, CA 94143, USA.
Thalhammer
A
A
Specht
C G
CG
Schoepfer
R
R
Burlingame
A L
AL
D000818:Animals; D001923:Brain Chemistry; D002852:Chromatography, Ion Exchange; D004798:Enzymes / Q000737:chemistry / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry* / Q000378:metabolism; D051379:Mice; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000378:metabolism; D010455:Peptides / Q000737:chemistry / Q000378:metabolism; D010750:Phosphoproteins / Q000737:chemistry* / Q000378:metabolism; D010766:Phosphorylation; D020543:Proteome / Q000737:chemistry; D021241:Spectrometry, Mass, Electrospray Ionization; D013347:Subcellular Fractions / Q000737:chemistry; D013570:Synaptic Membranes / Q000737:chemistry* / Q000378:metabolism; D009435:Synaptic Transmission / Q000502:physiology
D004798:Enzymes; D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D010455:Peptides; D010750:Phosphoproteins; D020543:Proteome