Odile Gateau

Sialyl-transferase mitochondriale

Abstract A sialyl transferase activity is found in purified mitochondria. It is not due to residual contamination and this enzymatic system is located in the outer mitochondrial membrane. This proves mitochondrial autonomy in regard to glycoconjugate sialylation.

Effects of defined synthetic phospholipids on glycosylation processes. Modifications of membrane-bound N-acetylglucosaminyl-transferase and solubilized sialyl-transferase activities.

In this paper, we study effects of phosphatidylcholine synthetic derivatives on two different glycosyltransferasic systems, a membranous mitochondrial N-acetylglucosaminyl-transferase and a solubilized microsomal sialyl-transferase. From all our results, we proved the strong inhibitory effect of lysophosphatidylcholines in these glycosylation processes. We investigated then the relationship between this inhibitory effect and the chemical structure of the studied phospholipids. From this work, it became obvious that the most important inhibitory effect is obtained with a molecule containing a long aliphatic side chain in sn-1 of glycerol, a carbon atom devoid of any large group in sn-2 and an hydrophilic part in sn-3 (specially choline).

Galactosyltransferase activities in mitochondria outer membrane: biosynthesis of galactosylated proteins.

1. Mitochondria outer membranes prepared from mouse livers were purified on a discontinuous sucrose gradient. Control in electron microscopy and marker enzymes assays confirmed purity and homogeneity of this fraction. 2. Purified mitochondria outer membranes exhibited significant UDP-galactose: glycoprotein galactosyltransferase activities when incubated with endogenous or exogenous glycoprotein acceptors in presence of detergent (Nonidet P40). 3. Some properties of two distinct mitochondrial galactosyltransferases, acting respectively on ovomucoid and ovine asialo-mucin were investigated. 4. Transfer of galactose on ovomucoid was maximal for a pH of 7.6 at 33 degrees C whereas asialo-mucin galactosyltransferase exhibited an optimum pH of 5.6 for an optimal temperature of 46 degrees C. 5. These two distinct membrane-bound enzymes were both inhibited by diacylglycerophospholipids whereas lysophospholipids modulated both enzymes in a different way: at 5 mM lysophosphatidylcholine, asialo-mucin galactosyltransferase was slightly stimulated while ovomucoid galactosyltransferase was markedly activated. 6. The most important activating effect on ovomucoid galactosyltransferase was obtained with a phospholipid containing a long aliphatic side chain linked by an ester bond in sn-1 of glycerol, an hydroxyl group or hydrogen atoms in sn-2 and a phosphorylcholine head group in sn-3.

Solubilization of Lipid-Glycosyltransferases from Mitochondrial Outer Membranes

In the mitochondria, the enzymes which catalyze the incorporation of sugars onto lipidic acceptors are intimately associated with membranes. We have previously shown that mitochondrial outer membranes contain autonomous enzymatic systems which transfer mannose (Gateau et al., 1973) and glucose (Gateau et al., 1980) onto lipidic endogenous acceptors. Although these enzymes have been studied in mitochondrial membranes, no study has yet investigated the interactions of these enzymes with phospholipids.

Biosynthesis and Characterization of Lipid-Linked Sugars in Outer Membrane of Liver Mitochondria

Publisher Summary This chapter discusses the biosynthesis and characterization of lipid-linked sugars in outer membrane of liver mitochondria. In liver and in brain mitochondria, autonomous glycoprotein biosynthesis is possible. The transfer of mannose, from GDP-mannose to an endogenous protein-acceptor, is catalyzed by a mannosyltransferase, localized specifically in the inner membrane of mitochondria, and not detected in the outer membrane, intermembranous space, or matrix. Infection of the cell by Myxovirus or Arbovirus does not change. The biosynthesis of the mannose-N-acetylglucosamine-core oligosaccharide of many glycoproteins appears to be mediated through the participation of polyprenyl-linked saccharide intermediates. It is important to search for the presence in mitochondria of glycosyltransferases that are able to catalyze the transfer of carbohydrate to polyprenic acceptors. The study of the biosynthesis by the outer membrane shows that GDP is an inhibitor of the reaction. The reversibility of this reaction was tested by incubating the outer membrane containing preformed endogenous [14C] mannosyl-phosphate-polyprenol with GDP and measuring formation of GDP mannose. Autonomy of mitochondria for a part of glycoconjugate biosynthesis responds to classic concepts and by autonomy, which is limited to internal sugars of the core and not possible for other external sugars fucose, galactose, and sialic acid.