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Information on EC 2.4.1.132 - GDP-Man:Man1GlcNAc2-PP-dolichol alpha-1,3-mannosyltransferase and Organism(s) Saccharomyces cerevisiae and UniProt Accession P43636
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2.4.1.132 GDP-Man:Man1GlcNAc2-PP-dolichol alpha-1,3-mannosyltransferaseIUBMB Comments
The biosynthesis of asparagine-linked glycoproteins utilizes a dolichyl diphosphate-linked glycosyl donor, which is assembled by the series of membrane-bound glycosyltransferases that comprise the dolichol pathway. Alg2 mannosyltransferase from Saccharomyces cerevisiae carries out an alpha1,3-mannosylation of D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol, followed by an alpha1,6-mannosylation (cf. EC 2.4.1.257), to form the first branched pentasaccharide intermediate of the dolichol pathway [1,2].
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Word Map
- 2.4.1.132
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alpha-1,2-mannosyltransferase
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lipid-linked
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alpha-1,3-linked
- mannoses
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beta-glcnac
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oligosaccharide-lipids
- mannans
- pharmacology
- medicine
The taxonomic range for the selected organisms is: Saccharomyces cerevisiae
The expected taxonomic range for this enzyme is: Eukaryota, Archaea, Bacteria
The expected taxonomic range for this enzyme is: Eukaryota, Archaea, Bacteria
Reaction Schemes
Synonyms
alpha-1,3-mannosyltransferase, mannosyltransferase ii, halg2, asparagine-linked glycosylation 2, scalg2, gdp-man:dol-pp-glcnac2man2 alpha-1,3-mannosyltransferase, gdp-man:man1glcnac2-pp-dolichol mannosyltransferase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
alpha-1,3-mannosyltransferase
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GDP-mannose-oligosaccharide-lipid mannosyltransferase II
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mannosyltransferase II
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mannosyltransferase, guanosine diphosphomannose-oligosaccharide-lipid II
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REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hexosyl group transfer
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PATHWAY SOURCE
PATHWAYS
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SYSTEMATIC NAME
IUBMB Comments
GDP-D-mannose:D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol 3-alpha-mannosyltransferase
The biosynthesis of asparagine-linked glycoproteins utilizes a dolichyl diphosphate-linked glycosyl donor, which is assembled by the series of membrane-bound glycosyltransferases that comprise the dolichol pathway. Alg2 mannosyltransferase from Saccharomyces cerevisiae carries out an alpha1,3-mannosylation of D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol, followed by an alpha1,6-mannosylation (cf. EC 2.4.1.257), to form the first branched pentasaccharide intermediate of the dolichol pathway [1,2].
CAS REGISTRY NUMBER
COMMENTARY
81181-76-2
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SUBSTRATE
PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
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-
-
?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
GDP-alpha-D-mannose + Man-(beta1,4)-Gn-(beta1,4)-Gn-PP-phytanyl
GDP + Man-(alpha1,3)[Man-(alpha1,6)]-Man1Gn2-PPhy
synthesis of acceptor phytanyl oligosaccharide, Man1Gn2-PPhy, from Gn-(beta1,4)-Gn-PP-phytanyl (Gn2-PPhy) using yeast Alg1. Recombinant scAlg2 transfers 2 Man residues to the beta1,4-Man of the Man1Gn2-PPhy substrate with alpha1,6 and alpha1,3-linkages, yielding Man-(alpha1,3)[Man-(alpha1,6)]-Man1Gn2-PPhy, cf. EC 2.4.1.257
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-
?
GDPmannose + tetrasaccharide-diphosphoryl-lipid
GDP + mannosyl-alpha-1,3-tetrasaccharide-diphosphoryl-lipid
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-
-
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?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
in eukaryotes, biosynthesis of N-glycans starts with the assembly of the common core oligosaccharide precursor Glc3Man9 GlcNAc2-PP-Dol, the glycan moiety of which is subsequently transferred onto selected Asn-Xaa-(Ser/Thr) acceptor sites of the nascent polypeptide chain by the oligosaccharyl-transferase complex
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-
?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
the biosynthesis of asparagine-linked glycoproteins utilizes a dolichylpyrophosphate-linked glycosyl donor, which is assembled by the series of membrane-bound glycosyltransferases that comprise the dolichol pathway. Alg2 carries out an alpha1,3-mannosylation of D-Man-beta-(1-4)-D-GlcNAc-beta-(1-4)-D-GlcNAc-diphosphodolichol, followed by an alpha1,6-mannosylation, to form the first branched pentasaccharide intermediate of the dolichol pathway
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?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
Alg2 carries out an alpha1,3-mannosylation of Man-beta1,4-GlcNAc-beta1,4-GlcNAc-diphosphodolichol, followed by an alpha1,6-mannosylation, to form the first branched pentasaccharide intermediate of the dolichol pathway
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-
?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
Alg2 is able to catalyze both the addition of the alpa1,3- and alpha1,6-linked mannose residue to Man1GlcNAc2-PP-Dol, forming Man2GlcNAc2-PP-Dol (cf EC 2.4.1.132) and subsequently to Man3GlcNAc2-PP-Dol
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-
?
additional information
?
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Alg2 shows no activity with D-Man-beta-(1-4)-D-GlcNAc-beta-(1-4)-D-GlcNAc-diphosphodolichol
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-
?
additional information
?
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unique bifunctionality of Alg2 during lipid-linked oligosaccharide (LLO) synthesis
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additional information
?
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Mnn1p has 2 conserved aspartate residues necessary for activity
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-
?
additional information
?
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Mnn1p family of alpha-1,3-mannosyltransferases is responsible for adding the terminal mannose residues of O-linked oligosaccharides, MNT2 and MNT3 genes in combination with MNN1 have overlapping roles in the addition of the fourth and fifth alpha-1,3-linked mannose residues to form Man4 and Man5 oligosaccharides
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?
additional information
?
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primary and secondary structure of the ALG2 protein
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?
additional information
?
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Mnn1p is required for the complex glycosylation of secreted proteins
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?
NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate)
(Substrate)
LITERATURE
(Substrate)
(Substrate)
COMMENTARY
(Product)
(Product)
LITERATURE
(Product)
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
r=reversible
ir=irreversible
?=not specified
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
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-
-
?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
GDPmannose + tetrasaccharide-diphosphoryl-lipid
GDP + mannosyl-alpha-1,3-tetrasaccharide-diphosphoryl-lipid
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-
-
-
?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
in eukaryotes, biosynthesis of N-glycans starts with the assembly of the common core oligosaccharide precursor Glc3Man9 GlcNAc2-PP-Dol, the glycan moiety of which is subsequently transferred onto selected Asn-Xaa-(Ser/Thr) acceptor sites of the nascent polypeptide chain by the oligosaccharyl-transferase complex
-
-
?
GDP-alpha-D-mannose + D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-D-GlcNAc-diphosphodolichol
GDP + D-Man-alpha-(1->3)-D-Man-beta-(1->4)-D-GlcNAc-beta-(1->4)-GlcNAc-diphosphodolichol
the biosynthesis of asparagine-linked glycoproteins utilizes a dolichylpyrophosphate-linked glycosyl donor, which is assembled by the series of membrane-bound glycosyltransferases that comprise the dolichol pathway. Alg2 carries out an alpha1,3-mannosylation of D-Man-beta-(1-4)-D-GlcNAc-beta-(1-4)-D-GlcNAc-diphosphodolichol, followed by an alpha1,6-mannosylation, to form the first branched pentasaccharide intermediate of the dolichol pathway
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-
?
additional information
?
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Mnn1p is required for the complex glycosylation of secreted proteins
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-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY
GeneOntology No.
LITERATURE
SOURCE
-
physical interactions between the Alg1, Alg2, and Alg11 mannosyltransferases
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
malfunction
cells deleted for ALG2 are inviable. Mutant alg2 alleles display intraallelic complementation
metabolism
the fourth and fifth steps of lipid-linked oligosaccharide (LLO) synthesis are catalyzed by Alg2, an unusual mannosyltransferase (MTase) with two different MTase activities
physiological function
asparagine (N)-linked glycosylation requires the ordered, stepwise synthesis of lipid-linked oligosaccharide (LLO) precursor Glc3Man9GlcNAc2-diphosphate-dolichol (Glc3Man9Gn2-PDol) on the endoplasmic reticulum. The fourth and fifth steps of LLO synthesis are catalyzed by Alg2, an unusual mannosyltransferase (MTase) with two different MTase activities. Alg2 adds both an alpha1,3- and alpha1,6-mannose ontoManGlcNAc2-PDol to form the trimannosyl core Man3GlcNAc2-PDol. Alg2-dependent Man3GlcNAc2-PDol production relies on net-neutral lipids with a propensity to form bilayers
additional information
the conserved C-terminal EX7E motif, N-terminal cytosolic tail, and 3G-rich loop motifs in Alg2 play crucial roles for these activities, both in vitro and in vivo. Alg2 immunoprecipitates from extracts of yeast microsomal membranes also displays both alpha1,3- and alpha1,6-mannosyltransferase (MTase) activities. The conserved Val62 residue is required for yeast Alg2 function. The first E (E335) and His-336 are partially required for alpha1,6-mannosylation, and importance of both E335 and E343 of the EX7E domain for Alg2 function in vivo. Identification of three conserved G-rich motifs in scAlg2, located in the N-terminal cytosolic short tail, in the middle of Alg2, and in the C-terminal domain. Residues G17, G19, and G20 are within the N-terminal cytosolic tail of Alg2, importance of this domain for Alg2 function
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
glycoprotein
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
E335A
E335A/E343A
significant lower level of product formation, identical to that of the E335A mutant
E343A
F337A
site-directed mutagenesis, Trx-scAlg2F337A produces 26% Man3Gn2 product compared to wild-type enzyme
G377R
site-directed mutagenesis, a temperature-sensitive alg2-1 mutant containing a single missense mutation, catalytically inactive
H336A
V62G
site-directed mutagenesis, Trx-scAlg2V62G produces 25% Man3Gn2 product compared to wild-type enzyme. The HA-tagged mutant allele (3HAscAlg2V62G) fails to complement the lethality of the alg2DELTA LSY2 when grown on 5-FOA
additional information
E335A
site-directed mutagenesis, Trx-scAlg2E335A produces only no final product and only 32% of intermediate Man2Gn2 compared to wild-type enzyme
H336A
site-directed mutagenesis, Trx-scAlg2H336A produces 8% Man3Gn2 product compared to wild-type enzyme
additional information
mutational analysis of Alg2 and identification of amino acids required for its activity. None of the four domains (predicted as transmembrane-spanning helices) is essential for transferase activity because truncated Alg2 variants can exert their function as long as Alg2 is associated with the endaplasmic reticulum by either its N- or C-terminal hydrophobic regions
additional information
site-directed mutagenesis of conserved EX7E motif. Trx-scAlg2E335A, mutated in the first E, has significantly decreased activity, producing no final product and only 32% of intermediate Man2Gn2. Trx-scAlg2E343A, mutated in the second E, has no detectable activity. The intervening amino acids of the EX7E are also important, though less than either E335 or E343. Trx-scAlg2H336A and Trx-scAlg2F337A produce 8% and 26% of Man3Gn2 product, respectively, compared to wild-type. Cells deleted for ALG2 are inviable, a plasmid shuffling technique is used to measure complementation. Mutant alg2 alleles display intraallelic complementation. Mutations (changed to proline) in five of the glycines (G19, G20, G256, G357, G358) result in complete loss of activity, while two of them (G17, G257) are significantly decreased
additional information
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ALG2 mutant with severely reduced enzyme activity
additional information
ALG2 mutant with severely reduced enzyme activity
additional information
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mutagenesis of Mnn1p by altering either of the 2 conserved aspartates eliminates all enzymic activity, but does not affect the overall folding and assembly of Mnn1p
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant N-terminal thioredoxin (Trx)-His6-tagged wild-type and mutant ALG2s from Escherichia coli strain DE3 membranes, preparation of proteoliposomes
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
gene ALG2, cloning from Saccharomyces cerevisiae strain W303a, recombinant expression of wild-type and mutant N-terminal thioredoxin (Trx)-His6-tagged enzymes in Escherichia coli strain DE3
overexpression in Escherichia coli. Two Alg2 constructs are expressed and isolated, one with the N-terminal TRX domain and C-terminal His and V5 epitope tags and the other with only an N-terminal His tag
ALG2 gene encodes GDP-Man:Dol-PP-GlcNAc2Man2 alpha-1,3-mannosyltransferase, amino acid sequence
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MNN1, MNT2 and MNT3 genes encode alpha-1,3-mannosyltransferases, wild type and mutants carrying single and multiple combinations of the disrupted gene
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REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Graham, T.R.; Krasnov, V.A.
Sorting of yeast alpha 1,3 mannosyltransferase is mediated by a lumenal domain interaction, and a transmembrane domain signal that can confer clathrin-dependent Golgi localization to a secreted protein
Mol. Biol. Cell
6
809-824
1995
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Graham, T.R.; Seeger, M.; Payne, G.S.; MacKay, V.L.; Emr, S.D.
Clathrin-dependent localization of alpha 1,3 mannosyltransferase to the Golgi complex of Saccharomyces cerevisiae
J. Cell. Biol.
127
667-678
1994
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Romero, P.A.; Lussier, M.; Veronneau, S.; Sdicu, A.M.; Herscovics, A.; Bussey, H.
Mnt2p and Mnt3p of Saccharomyces cerevisiae are members of the Mnn1p family of alpha-1,3-mannosyltransferases responsible for adding the terminal mannose residues of O-linked oligosaccharides
Glycobiology
9
1045-1051
1999
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Thiel, C.; Schwarz, M.; Peng, J.; Grzmil, M.; Hasilik, M.; Braulke, T.; Kohlschuetter, A.; von Figura, K.; Lehle, L.; Koerner, C.
A new type of congenital disorders of glycosylation (CDG-Ii) provides new insights into the early steps of dolichol-linked oligosaccharide biosynthesis
J. Biol. Chem.
278
22498-22505
2003
Homo sapiens, Homo sapiens (Q9H553), Saccharomyces cerevisiae, Saccharomyces cerevisiae (P43636), Saccharomyces cerevisiae A
Yip, C.L.; Welch, S.K.; Klebl, F.; Gilbert, T.; Seidel, P.; Grant, F.; O'Hara, P.J.; MacKay, V.L.
Cloning and analysis of the Saccharomyces cerevisiae MNN9 and MNN1 genes required for complex glycosylation of secreted proteins
Proc. Natl. Acad. Sci. USA
91
2723-2727
1994
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Reynolds, T.B.; Hopkins, B.D.; Lyons, M.R.; Graham, T.R.
The high osmolarity glycerol response (HOG) MAP kinase pathway controls localization of a yeast Golgi glycosyltransferase
J. Cell Biol.
143
935-946
1998
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Wiggins, C.A.R.; Munro, S.
Activity of the yeast MNN1 alpha-1,3-mannosyltransferase requires a motif conserved in many other families of glycosyltransferases
Proc. Natl. Acad. Sci. USA
95
7945-7940
1998
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Shpakov, A.O.; Derkach, K.V.
Yeast dolichol-coupled mannosyltransferases. Theoretical analysis of primary structure and identification of sites homologous to other enzymes of the dolichol cycle
Zh. Evol. Biokhim. Fiziol.
32
3-18
1996
Saccharomyces cerevisiae, Saccharomyces cerevisiae A
Gao, X.D.; Nishikawa, A.; Dean, N.
Physical interactions between the Alg1, Alg2, and Alg11 mannosyltransferases of the endoplasmic reticulum
Glycobiology
14
559-570
2004
Saccharomyces cerevisiae
O'Reilly, M.K.; Zhang, G.; Imperiali, B.
In vitro evidence for the dual function of Alg2 and Alg11: essential mannosyltransferases in N-linked glycoprotein biosynthesis
Biochemistry
45
9593-9603
2006
Saccharomyces cerevisiae (P43636)
Kmpf, M.; Absmanner, B.; Schwarz, M.; Lehle, L.
Biochemical characterization and membrane topology of Alg2 from Saccharomyces cerevisiae as a bifunctional alpha1,3- and 1,6-mannosyltransferase involved in lipid-linked oligosaccharide biosynthesis
J. Biol. Chem.
284
11900-11912
2009
Saccharomyces cerevisiae (P43636)
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