@misc{oai:niigata-u.repo.nii.ac.jp:00004444,
 author = {Ishizuka, Toru},
 month = {Mar},
 note = {Two monoclonal antibodies (SPM-1 and SPM-2) immunoprecipitate brain N-type calcium channels. On immunoaffinity chromatography of digitonin or Triton X-100 extracts of bovine brain membranes on SPM-1- and SPM-2-Sepharose, proteins of 36 kDa (syntaxins), 28 kDa (SNAP-25 and rab3A), and 19 kDa are specifically retained by both columns. Here I show that the 19 kDa protein contains VAMP/synaptobrevin-2 and novel cytosolic proteins termed synaphins. We have isolated and cloned a bovine and two rat brain cDNAs for synaphins. Synaphins are very hydrophilic proteins consisting of 134 amino acid residues rich in glutamic acid and lysine residues. Bovine synaphin and rat synaphin 1 is 100% identical, and rat synaphin 2 exhibits 84% identity with bovine and rat synaphins 1. Immunoblot and Northern blot analyses in various rat tissues showed that synaphins predominantly exist in brain. In situ hybridization. with synthetic 50 meric oligonucleotide probe specific to synaphins 1 or 2 demonstrated different localizations of the two mRNAs in the rat brain. Subcellular fractionation studies revealed that synaphins exist in both soluble and insoluble fractions. A Triton X-100 extract of the rat brain LP2 fraction was subjected to a glycerol density gradient, and the resultant fractions were immunoprecipitated with monoclonal antibodies that recognize syntaxin 1, SNAP-25 or synaphins. Synaphins fractionated in insoluble fraction were mostly present in the 7S complex together with synaptotagmin, syntaxin, SNAP-25, and VAMP/synaptobrevin. These results strongly suggest that the synaptic vesicle proteins, VAMP/synaptobrevin and synaptotagmin, the plasma membrane proteins, syntaxin and SNAP-25, and cytosolic protein, synaphin, form a tight complex probably essential to neurotransmitter release in brain presynaptic terminals. Functional relevance of two synaphin isoforms is unknown, but it is likely that specific sets of protein variants forming the complex will confer different properties on the docking/fusion complex and thereby contribute the distinct functions onto individual neurons., 学位の種類: 博士（理学）. 報告番号: 甲第1268号. 学位記番号: 新大院博（理）甲第72号. 学位授与年月日: 平成9年3月25日, 新大院博（理）甲第72号},
 title = {Isolation and characterization of synaphins, novel cytosolic proteins associated with the docking/fusion complex for neurotransmitter release},
 year = {1997}
}