Endogenous glycosphingolipid acceptor specificity of sialosyltransferase systems in intact Golgi membranes, synaptosomes, and synaptic plasma membranes from rat brain.
Preparations highly enriched in Golgi complex membranes, synaptosomes, and synaptic plasma membranes (SPM) by marker enzyme analysis and electron microscopic morphology were made from the brains of 28-day-old rats. These were incubated with cytidine 5'-monophosphate-N-acetyl[14C]neuraminic acid (CMP-NeuAc) in a physiologic buffer, without detergents. Glycolipid sialosyltransferase activities (SATs) were measured by analyzing incorporation of radiolabeled NeuAc into endogenous membrane gangliosides. Golgi SAT was diversified in producing all the various molecular species of labeled gangliosides [2.64 pmol of NeuAc transferred (mg of protein)-1 h-1]. Synaptosomal SAT exhibited a lower activity [0.66 pmol (mg of protein)-1 h-1], but it was highly specific in its labeling pattern, with a marked preference for labeling NeuAc alpha 2----8NeuAc alpha 2----3Gal beta 1----4Glc beta 1----1 Cer (GD3 ganglioside). SPM prepared from the synaptosomes retained the GD3-related SAT (or SAT-2), and the total specific activity increased [1.41 pmol (mg of protein)-1 h-1], which suggests that the location of the synaptosomal activity is in the SPM. These results indicate that SAT activity in Golgi membranes differs from that in synaptosomes with regard to endogenous acceptor substrate specificity and SAT activity of synaptosomes should be located in the synaptosomal plasma membrane. This SAT could function as an ectoenzyme in concert with ecto-sialidase to modulate the GD3 and other ganglioside population in situ at the SPM of the central nervous system.
Durrie
R
R
Division of Molecular Biology and Neural Regeneration, Nathan S. Kline Institute for Psychiatric Research, Orangeburg, New York 10962.
Saito
M
M
Rosenberg
A
A
D001921:Brain / Q000201:enzymology*; D002458:Cell Fractionation; D005732:Gangliosides / Q000032:analysis; D006028:Glycosphingolipids / Q000378:metabolism*; D006056:Golgi Apparatus / Q000201:enzymology* / Q000648:ultrastructure; D007700:Kinetics; D012799:Sialyltransferases / Q000378:metabolism*; D013379:Substrate Specificity; D013570:Synaptic Membranes / Q000201:enzymology* / Q000648:ultrastructure; D013574:Synaptosomes / Q000201:enzymology* / Q000648:ultrastructure
D005732:Gangliosides; D006028:Glycosphingolipids; D012799:Sialyltransferases