Isoosmotic isolation of rat brain synaptic vesicles, some of which contain tyrosine hydroxylase.
Rat brain synaptic vesicles were isoosmotically isolated and examined for Mg(2+)-ATPase [EC 3.6.1.3.] and tyrosine hydroxylase [EC 1.14.16.2.] associated with the synaptic vesicles. Synaptosomes in 0.32 M sucrose were disrupted by freezing and thawing treatment, and the cytosol fraction was fractionated on a Sephacryl S-500 column with a mean exclusion size of 200 nm. Peak I at the void volume was a mixture of large vesicular membranes, small amounts of synaptic vesicles and coated vesicles, etc. Peak II consisted of non- and granulated synaptic vesicles of 35-40 nm diameter, and peak III of soluble proteins. The synaptic vesicles in peak II reacted with antibodies against the H(+)-ATPase A-subunit, vesicular acetylcholine transporter, and vesicular monoamine transporter. However, they showed little Mg(2+)-ATPase activity. Tyrosine hydroxylase was observed in either peak II or III on blotting with an anti-tyrosine hydroxylase antibody. These results imply that tyrosine hydroxylase exists in soluble and bound forms to synaptic vesicles in nerve terminals.
Tsudzuki
Toshihiro
T
Department of Biochemistry I, Nagoya City University Medical School, Kawasumi 1, Mizuho-ku, Nagoya 467-8601.
Tsujita
Maki
M
D000109:Acetylcholine / Q000737:chemistry; D000251:Adenosine Triphosphatases / Q000737:chemistry; D000818:Animals; D001692:Biological Transport; D015153:Blotting, Western; D001921:Brain / Q000473:pathology*; D017301:Ca(2+) Mg(2+)-ATPase / Q000737:chemistry; D002850:Chromatography, Gel; D003600:Cytosol / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D008297:Male; D008854:Microscopy, Electron; D009995:Osmosis; D051381:Rats; D017208:Rats, Wistar; D013395:Sucrose / Q000494:pharmacology; D013569:Synapses / Q000378:metabolism*; D013572:Synaptic Vesicles / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000737:chemistry
D013395:Sucrose; D014446:Tyrosine 3-Monooxygenase; D000251:Adenosine Triphosphatases; D017301:Ca(2+) Mg(2+)-ATPase; D000109:Acetylcholine