Effect of bacitracin on the biosynthesis of dolichol derivatives in calf pancreas microsomes

Herscovics, Annetté; Bugge, Birgitte; Jeanloz, Roger W.

doi:10.1016/0014-5793(77)80587-5

Cited by 37 publications

(15 citation statements)

References 15 publications

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“…However maltose, which is or1 -4, also inhibited. The conclusion that three glucoses of (Glc)3(Man)q(GlcNAc)z have the x configuration agrees with the published evidence which is not all very convincing [6,16,171.…”

Section: Discussionsupporting

confidence: 77%

Microsomal Glucosidases of Rat Liver. Partial Purification and Inhibition by Disaccharides

Ugalde

Staneloni

Leloir

2005

European Journal of Biochemistry

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Further work on microsomal glucosidases of rat liver has confirmed that at least two enzymes are involved in the removal of glucose from the glucose-containing oligosaccharide. One acts on the oligosaccharide containing three glucose residues and another on the oligosaccharide which has one or two glucoses.The glucosidase which acts on (Glc),(Man),(GlcNAc), could be purified with a Concanavalin-A -Sepharose column followed by electrofocusing. This purified preparation was active on the oligosaccharide containing one or two glucoses. Heat inactivation and inhibition by disaccharides was parallel for both activities. Inhibition of the glucosidase active on (Glc),(Man),(GlcNAc), was obtained with kojibiose which has an a 1-2 linkage, while the glucosidase acting on (Glc), -,(Man),-(GlcNAc), was inhibited by nigerose (a 1-3 linkage), maltose (a 1-4 linkage) and glucose at a higher concentration. None of the / 3 anomers inhibited. These results are consistent with an a configuration of the three glucoses of the dolichyl-diphosphate-linked oligosaccharide. Kojibiose was found to inhibit glucosidase action not only on the free oligosaccharide but also on protein-bound one.The glycosylation of asparagine residues in proteins occurs by transfer of an oligosaccharide from its DolPP derivative [l, 21. The oligosaccharide which is transferred contains three glucose, nine mannose and two N-acetylglucosamine residues [3]. In order to be transferred the oligosaccharide has to have three glucoses and presumably these serve as signals for the transfer [4, 51. The step which immediately follows is the removal of the glucose residues after which other processing reactions may occur. These glucoses are linked as follows to the rest of the oligosaccharide : Glc(1-2)Glc(l-3)Glc(l-3)Man-R [3]. Glucosidases acting on (Glc)l-3(Man)g(GlcNAc)z were first detected in the microsomal fraction of liver [6]. It was found later that at least two different membrane-bound glucosidases could be detected using as substrates oligosaccharides of different glucose content [7 -91. One of these enzymes acts on the Glc(1-2)Glc linkage. It can be extracted with phosphate buffer and a nonionic detergent [S].Abbreuations. Glc, glucose; Man mannose; GlcNAc, N-acetylglucosamine; DolP, dolichyl phosphate; DolPP, dolichyl diphosphate; (Glc), , (Glc),, and (Glch glucosidases are the enzymes that act on the oligosaccharides containing one, two, and three glucose residues.Another glucosidase acts on the Glc(1-3)Glc linkages. It has been partially purified and some of its properties have been described [7 -91. Further studies on these enzymes are described in this paper. MATERIALS AND METHODSNigerose was prepared by partial hydrolysis of nigerian obtained from Koch Light Laboratories. Kojibiose and sophorose were supplied by Koch Light ; maltose, isomaltose and cellobiose by Sigma. Substrates

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Section: Discussionsupporting

confidence: 77%

Microsomal Glucosidases of Rat Liver. Partial Purification and Inhibition by Disaccharides

Ugalde

Staneloni

Leloir

2005

European Journal of Biochemistry

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“…The [19] for the same products. Biosynthesis of product Glb was increased by exogenous dolichyl monophosphate.…”

Section: Discussionmentioning

confidence: 99%

“…Formation of dolichylglucosyl phosphate has been reported for different membrane preparations [l, 2,15,19,201. In these, original experiments conducted with Acanthamoeha custellani cysts [21] have shown that these membrane preparations catalyse the transfer of labeled glucose into a dolichylglucosyl phosphate which has identical chromatographic behaviour to product Glb (on DEAE-cellulose column).…”

Section: Discussionmentioning

confidence: 99%

Glucosyltransferase Activities in Liver Mitochondria

Gateau¹,

Morelis²,

Louisot³

1980

European Journal of Biochemistry

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Mitochondria, and specially outer mitochondrial membranes, incorporate d‐[125C]glucose from UDP‐d‐[14C]glucose into products extracted with organic solvents and into a residual precipitate, with a pH optimum of about 6.5 in (2‐N‐morpholine‐ethane)‐sulfonic acid (MES) buffer. The chloroform/methanol (2:1, v/v) extract contains two products. The major [14C]glucolipid is stable to mild alkali, but releases [14C]glucose upon mild acid hydrolysis. It is retained on DEAE‐cellulose (acetate form) and is eluted with the same ionic strength as an hexosyldolichyl monophosphate diester. This [14C] glucolipid has the same chromatographic behaviour as dolichyl‐mannosylphosphate in neutral, acidic and basic solvent systems; and its biosynthesis is greatly increased by exogenous dolichylmonophosphate. the other [14C]glucolipid is stable upon mild acid hydrolysis and is not retained on DEAE‐cellulose. On silicic acid it is eluted with acetone. The biosynthesis of this compound is stimulated by exogenous ceramide. This glucolipid has the same chromatographic mobility in different solvent systems as glucosylceramide isolated from the liver of a patient with Gaucher's disease. Biosynthesis of these two glucolipids is inhibited by UDP, but only biosynthesis of dolichylglucosyl monophosphate is reversible with this nucleotide. The biosynthesis of these different glucosylated derivatives is stimulated by the addition of divalent cations (Mn2+, Mg2+). The effect of these two metal ions on dolichylglucosyl monophosphate and glucosylceramide formation is studied in different conditions.

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“…The glycosidase digestions and gel filtration studies strongly suggest a structure identical to a typical neutral oligosaccharide [Mann(GlcNAc)2] linked to asparagine with an estimate of 7-9 mannose residues. The possible presence of one or two glucose residues in the precursor oligosaccharide structures, as suggested elsewhere forlipid-and protein-linked oligosaccharides (23)(24)(25), was not directly examined and cannot be rigorously excluded. The a-mannosidase digestions, however, indicate that glucose was not present in a terminal, nonreducing position, which would have protected much of the radiolabeled mannose from removal.…”

Section: Methodsmentioning

confidence: 99%

Oligosaccharide chains are trimmed during synthesis of the envelope glycoprotein of vesicular stomatitis virus.

Hunt¹,

Etchison²,

Summers³

1978

Proc. Natl. Acad. Sci. U.S.A.

196

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The biosynthesis and maturation of the oligosaccharide moieties of the envelope glycoprotein of vesicular stomatitis virus were investigated in virus-infected HeLa and BHK21 cells after pulse labeling with [2-3Hlmannose. Two major forms of the virus glycoprotein were detected by polyacrylamide gel electrophoresis, which appear to correspond to the viral glycoprotein with either "precursor" or "mature" oligosaccharide chains. The precursor chains in both HeLa and BHK21 cells infected with vesicular stomatitis virus obtained after a 30-min pulse were large oligomannose structures containing approximately 7-9 mannose residues as estimated by gel filtration analysis. The size of the oligomannose structures initially transferred to the protein may have been even larger. Mature, virus-size oligosaccharide chains, which could be detected after a 20-to 30-min delay, contained only three mannose residues and, in addition, contained branch structures terminating in sialic acid. A precursor-product relationship of these two forms of oligosaccharide chains was demonstrated by pulse-chase labeling of virus-infected HeLa cells. These studies indicated that the large oligomannosyl core structures initially added to the glycoprotein were being "trimmed" by the removal of mannose residues prior to (and/or during) the addition of the branch chains terminating in sialic acid. Vesicular stomatitis virus (VSV) is an enveloped, RNA-containing virus of the rhabdovirus group which matures by budding through host cell membranes that have been modified by replacement of host proteins with two virus-specified proteins, one of which is a glycoprotein (1). The VSV-infected animal cell is an excellent experimental system for elucidating the molecular and subcellular mechanisms involved in the biosynthesis and maturation of both cell membrane and virus envelope glycoproteins (2-8).The VSV G polypeptide contains two major glycosylation sites with complex oligosaccharides that are linked N-glycosidically to asparagine (9-11). As expected for the biosynthesis of a membrane glycoprotein, mRNA coding for VSV G is localized on membrane-bound polyribosomes in infected cells (12, 13), and newly synthesized G is found predominantly in rough endoplasmic reticulum membranes (14). The sugar residues in the oligomannosyl core [Mann(GlcNAc)2] are apparently added to the G polypeptide in the rough endoplasmic reticulum, whereas the branch sugars (NeuNAc-Gal-GlcNAc) and fucose are apparently added in smooth internal membranes prior to the appearance of the mature glycoprotein at plasma membranes (15). The initial glycosylation event probably occurs by the en bloc transfer of a preformed oligomannosyl core structure from a lipid intermediate (16).We have examined the product of the initial glycosylation of the VSV G glycoprotein and followed its conversion to mature, virus-size oligosaccharide chains in VSV-infected HeLa and BHK21 cells. Our results indicate that a large oligomannosyl core structure [Man>7(GlcNAc)2] is initially added to the glycopro...

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Effect of bacitracin on the biosynthesis of dolichol derivatives in calf pancreas microsomes

Cited by 37 publications

References 15 publications

Microsomal Glucosidases of Rat Liver. Partial Purification and Inhibition by Disaccharides

Microsomal Glucosidases of Rat Liver. Partial Purification and Inhibition by Disaccharides

Glucosyltransferase Activities in Liver Mitochondria

Oligosaccharide chains are trimmed during synthesis of the envelope glycoprotein of vesicular stomatitis virus.

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