Dolichol Monophosphate Glucose: An Intermediate in Glucose Transfer in Liver

Behrens, Nicolas; Leloir, Luis F.

doi:10.1073/pnas.66.1.153

Cited by 243 publications

(62 citation statements)

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“…Dol-P was isolated and purified up to the DEAE-cellulose step [5]. D o~-P -[~~C ] G~C was prepared by incubation of UDP-[14C]Glc with liver microsomes and purified by DEAE-cellulose chromatography [5,6]. Dol-P-P-Glc-oligosaccharide labelled in the glucose or in the mannose was obtained by incubation of microsomes with UDP-['4C]G1~ [7] Abbreviations.…”

Section: Methodsmentioning

confidence: 99%

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Addition of Glucose to Dolichyl Diphosphate Oligosaccharide and Transfer to Protein

Staneloni

Ugalde

Leloir

1980

European Journal of Biochemistry

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The glycosylation of asparagine residues in proteins is known to occur by transfer from a dolichyl diphosphate oligosaccharide containing glucose. Paper chromatography allowed the separation of oligosaccharides (obtained by acid hydrolysis of the dolichyl diphosphate derivative) containing 1, 2 and 3 glucose residues. Using this procedure it was found that the addition of all three glucoses to the dolichyl diphosphate oligosaccharide occur with dolichyl phosphate glucose as donor. Furthermore only the compound with three glucoses was used as donor in the transfer to protein. The addition of glucose to exogenous dolichyldiphosphate oligosaccharide labelled by transfer from radioactive guanosine diphosphate mannose was detected.There is evidence showing that the donor for the glycosylation of asparagine residues in proteins is a dolichyl diphosphate oligosaccharide [l -21. The evidence shows that this oligosaccharide, whose structure is known [3], is built up by the transfer of acetylglucosamine phosphate to dolichyl phosphate followed by the addition of one more acetylglucosamine, nine mannoses and one to three glucoses. The oligosaccharide is then transferred to protein and the glucose residues are removed. The complex oligosaccharides may then be formed by removal of six mannoses and addition of acetylglucosamine, galactose and sialic acid. Another pathway consisting in removal or addition of mannose residues would lead to the formation of the high-mannose-type oligosaccharides.This paper reports studies on some aspects of the process of glucose addition to the oligosaccharide and of transfer of the latter to protein. MATERIALS AND METHODSRat liver microsomes were prepared as described previously [4]. Dol-P was isolated and purified up to the DEAE-cellulose step [5]. D o~-P -[~~C ] G~C was prepared by incubation of UDP-[14C]Glc with liver microsomes and purified by DEAE-cellulose chromatography [5,6]. Dol-P-P-Glc-oligosaccharide labelled in the glucose or in the mannose was obtained by incubation of microsomes with UDP-['4C]G1~ [7] Abbreviations. The glucose-containing oligosaccharides with three, two and one glucose residues are referred to as Glc3, Glcz and Glcl oligosaccharides. Dol, dolichyl; Glc, glucose; Man, mannose; P, phosphate.or GDP-['4C]Man [8] respectively. Dol-P-P-Glc-oligosaccharide labelled in mannose and glucose was obtained from oviduct slices incubated with ['4C]-mannose [9,10]. Free oligosaccharides were obtained from the dolichyl diphosphate derivatives by mild acid hydrolysis [lo].Paper chromatography of the oligosaccharides was carried out with the following solvent: 1-propanol/ nitromethane/water (5 : 2 : 4). This mixture is referred to as the propanol-nitromethane solvent. RESULTSPreparation of Dolichyl-Diphosphate-Glucose-Containing Oligosaccharides. Incubation of UDP-[14C]G1~ with liver microsomes leads to the formation of labelled Dol-P-P oligosaccharides. Mild acid hydrolysis of these compounds results in the liberation of free oligosaccharides which can be separated by paper chroma...

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Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Addition of Glucose to Dolichyl Diphosphate Oligosaccharide and Transfer to Protein

Staneloni

Ugalde

Leloir

1980

European Journal of Biochemistry

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“…On the other hand, dolichol is more widely distributed within the cell and in different tissues and organs. Dolichol was first identified in 1960 by Pennock, Hemming & Morton [1], and it was established by Behrens & Leloir in 1970 [2] that its phosphorylated derivative is an obligatory intermediate in the synthesis of certain types of glycoproteins. Of all lipids, this compound has thus been studied for the shortest period of time and, despite detailed investigations in many laboratories in recent years, our knowledge about the biosynthesis, distribution and function of dolichol is still fragmentary.…”

Section: Introductionmentioning

confidence: 99%

The biological role of dolichol

Chojnacki

Dallner²

1988

Biochemical Journal

286

191

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“…In the first paper on the formation of dolichol monophosphate glucose (dolichol-P-Glc) it was reported that the glucosyl residue of this substance was transferred to an endogenous acceptor on further incubation with liver microsomes (1). The product thus formed was believed to be a glycoprotein because of its insolubility in certain organic solvents and in trichloroacetic acid.…”

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The Role of Polyprenol-Bound Saccharides as Intermediates in Glycoprotein Synthesis in Liver

Parodi¹,

Behrens²,

Leloir³

et al. 1972

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

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149

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It has been reported that liver microsomes catalyze the transfer of glucose from uridine diphosphate glucose to dolichol monophosphate so as to produce dolichol monophosphate glucose. Dolichol is a polyprenol containing about 20 isoprene units. The glucosyl residue of dolichol monophosphate glucose is transferred to an endogenous acceptor on further incubation with liver microsomes. The glucosylated endogenous acceptor appears to be an oligosaccharide of about 20 monosaccharide units bound to dolichol through a phosphate or pyrophosphate bridge. In this paper it is reported that liver microsomes catalyze the transfer of the oligosaccharide from the glucosylated endogenous acceptor to an endogenous protein. This transfer reaction requires the presence of bivalent cations,-manganese being more effective than magnesium. The presence of deoxycholate is also required. Besides the glycoprotein, several watersoluble products are also formed. Preliminary evidence indicates that they are glucose, oligosaccharides of different size, and possibly oligosaccbarides bound to amino acids.In the first paper on the formation of dolichol monophosphate glucose (dolichol-P-Glc) it was reported that the glucosyl residue of this substance was transferred to an endogenous acceptor on further incubation with liver microsomes (1). The product thus formed was believed to be a glycoprotein because of its insolubility in certain organic solvents and in trichloroacetic acid. However, further work (2, 3) showed that most of the radioactivity transferred from glucoselabeled dolichol-P-Glc under the conditions described was recovered in an amphipatic substance, the hydrophilic residue of which appears to be an' oligosaccharide of about 20 monosaccharide units. The lipophilic residue had some properties in common with dolichol-P, and evidence was obtained indicating that the bridge between the lipid moiety and the oligosaccharide is a pyrophosphate. The substance was referred to as glucosylated endogenous acceptor (Glc-acceptor). On the basis of present evidence its structure can be written as dolichol-P-P-glycoseg-Glc. Dolichol (4) is a polyprenol containing about 20 isoprene units and glycosel9 stands for an oligosaccharide of about 19 unidentified monosaccharide residues. It should be emphasized that this is a tentative formula for the Glc-acceptor. Several aspects of its structure have not been fully clarified, especially the number and identity of the monosaccharide residues and the number of phosphates in the bond between the lipophilic and hydrophilic moieties.In this paper we report that under appropriate conditions, liver microsomes catalyze the transfer of the hydrophilic moiety of the Glc-acceptor to an endogenous protein. The reactions of glucose transfer under study can be schematically written as follows:UDP-Glc -dolichol-P-Glc dolichol-P-P-glycoseis-Glc protein-glycoseig-Glc MATERIALS AND METHODSRadioactive Glc-acceptor labeled in the glucose residue was obtained directly from UDP-Glc as described (5).Enzymes. Papain 2 X ...

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Dolichol Monophosphate Glucose: An Intermediate in Glucose Transfer in Liver

Abstract: Abstract. The microsomal fraction of liver has been found to catalyze glucose transfer from UDPG to a lipid acceptor which appears to be identical to the compound obtained by chemical

Cited by 243 publications

References 14 publications

Addition of Glucose to Dolichyl Diphosphate Oligosaccharide and Transfer to Protein

Addition of Glucose to Dolichyl Diphosphate Oligosaccharide and Transfer to Protein

The biological role of dolichol

The Role of Polyprenol-Bound Saccharides as Intermediates in Glycoprotein Synthesis in Liver

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