Purification and proteomic analysis of synaptic vesicles.
Synaptic vesicles store and subsequently release neurotransmitters into the synaptic cleft thereby regulating chemical neurotransmission in the brain. Proteins present in synaptic vesicles vary greatly in structure and function and have been identified primarily by genetic knock-out analysis in C. elegans, Drosophila, and mice (1,2). However, knock-out methods are not useful for the identification of proteins when a detectable phenotype is not created. Further, certain knocked-out proteins have function(s) that could be compensated for by another protein or cause a lethal phenotype when deleted. Additionally, some transporters and enzymes that appear to copurify with synaptic vesicles have not been characterized and confirmed (3-5). We have determined the proteins associated with purified synaptic vesicles using 2-D polyacrylamide gel electrophoresis (PAGE) protein separation followed by identification by mass spectrometry (6). Some of the new proteins identified were evaluated by western blot and confocal immunofluorescence analysis.
Cox
Holly D
HD
Department of Pharmaceutical Sciences, University of Montana, Missoula, MT, USA.
Thompson
Charles M
CM
D000818:Animals; D015153:Blotting, Western / Q000379:methods; D001921:Brain / Q000648:ultrastructure; D002458:Cell Fractionation / Q000379:methods; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D004721:Endoplasmic Reticulum / Q000648:ultrastructure; D005990:Glycerol; D007525:Isoelectric Focusing / Q000379:methods; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification; D009419:Nerve Tissue Proteins / Q000737:chemistry* / Q000302:isolation & purification*; D020543:Proteome; D051381:Rats; D013572:Synaptic Vesicles / Q000737:chemistry* / Q000648:ultrastructure*
D008565:Membrane Proteins; D009419:Nerve Tissue Proteins; D020543:Proteome; D005990:Glycerol