Journal of Lipid Research, Vol 34, 905-914, Copyright © 1993 by Lipid Research, Inc.
Effect of membrane lipids on the lactosylceramide molecular species specificity of CMP-N-acetylneuraminate:lactosylceramide sialyltransferase
H Kadowaki, MA Grant and LA Williams
Department of Medicine, Boston University School of Medicine, MA 02118.
It has previously been shown that when the molecular species specificity of
rat liver Golgi CMP-N-acetylneuraminate:lactosylceramide alpha
2,3-sialyltransferase was determined, using as the substrate
lactosylceramide (LacCer) incorporated into liposomes prepared with rat
liver Golgi lipids, the enzyme showed a pronounced variation in activity
towards the various molecular species of LacCer (J. Lipid Res. 1989. 30:
1789-1797). In this paper, -the LacCer molecular species specificity of
sialyltransferase from neuroblastoma NB2a cells was examined using five
naturally occurring and three synthetic molecular species of LacCer. The
enzyme activity was determined by following the formation of [14C]GM3 from
CMP-[14C]neuraminic acid and individual molecular species of LacCer
incorporated into liposomes. Nonspecific lipid transfer protein was
included in the enzyme assay to facilitate the transfer of LacCer and other
lipids between the liposomes and the membrane where sialyltransferase is
located. In these enzyme assays the liposomes contained approximately 10
times more lipid phosphorus than either the microsomal fraction of NB2a
cells or the Golgi fraction of rat liver. Thus, in the presence of
nonspecific lipid transfer protein, the lipid composition of the membrane
where sialyltransferase is located was modified to resemble the lipid
composition of the liposomes. When the molecular species specificity of
NB2a cell sialyltransferase was determined with LacCer incorporated into
liposomes prepared with NB2a cell lipids, the enzyme showed no specificity
towards the various molecular species of LacCer. However, when the
molecular species specificity of NB2a cell sialyltransferase was determined
with LacCer incorporated into liposomes prepared with rat liver Golgi
lipids, the enzyme showed a variation in activity towards the various
LacCer molecular species similar to that observed with the liver Golgi
enzyme using liposomes prepared with liver Golgi lipids. Likewise, when the
molecular species specificity of rat liver Golgi sialyltransferase was
determined with LacCer incorporated into liposomes prepared with NB2a cell
lipids, the liver enzyme then showed no specificity towards the various
molecular species of LacCer. These results indicate that the lipid
environment of the membrane can alter the molecular species specificity of
sialyltransferase towards its lipid substrate, LacCer.
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