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Taxonomic group: fungi / Ascomycota (Phylum: Ascomycota)

NCBI PubMed ID: 7018901
Journal NLM ID: 0107600
Publisher: Oxford, UK: Blackwell Science Ltd. on behalf of the Federation of European Biochemical Societies
Institutions: Institut für Botanik, Fakultät für Biologie und Vorklinische Medizin, Universität Regensburg

The enzyme transferring the oligosaccharide from DolPP-(GlcNAc)2(Glc)3 to asparagine residues of glycoproteins has been solubilized from yeast membranes by extraction with detergents. Enzyme activity was tested by measuring transfer of the glycosyl moiety from DolPP-[14C]saccharides to the hexapeptide Tyr-Asn-Leu-Thr-Ser-Val. The rate of transfer was linear for 20 min, with about 40% of dolichyl-diphosphate-bound radioactivity transferred to the peptide. The solubilized enzyme has been characterized as follows: 1. The enzyme is most efficiently solubilized (60% of the membrane-associated activity) by 0.5% Nonidet P40 at a protein/detergent ratio of 2. Octylglucoside solubilizes one third of the activity, but strongly inhibits the reaction if present in the test at a concentration of 1%. 2. Divalent cations are absolutely required. 1 mM Mn2+ is optimal; Mg2+ at a concentration of 10mM yields only one third the activity observed with Mn2+. 3. The enzyme transfers besides dolichyl-diphosphate-bound (GlcNAc)2(Man)9(Glc)3 also (GlcNAc)2(Man)1 and (GlcNAc)2; the rate decreases in this order. No transfer is observed from DolPP-(GlcNAc)2(Man)9 and from DolPP-GlcNAc. 4. The Km value for DolPP-(GlcNAc)2(Man)9(Glc)3 of 0.5 microM does not differ significantly from that for DolPP-(GlcNAc)2 of 1.2 microM. A broad pH-optimum for the reaction with both substrates was found between 6.5 and 7.7. 5. However, a clear difference in Km values for the hexapeptide was observed with difference dolichol-linked sugar derivatives. With DolPP-(GlcNAc)2 a peptide concentration of 0.6 mM resulted in half-maximal transfer rate, whereas 0.05 mM peptide were sufficient with DolPP-(GlcNAc)2(Man)9(Glc)3 as donor.

Structure type: oligomer
Location inside paper: DolPP-(GlcNAc)2(Man)1
Aglycon: dolichol
Compound class: N-glycan
Contained glycoepitopes: IEDB_135813,IEDB_137340,IEDB_137485,IEDB_141807,IEDB_144983,IEDB_150077,IEDB_151531,IEDB_152206,IEDB_983930,SB_44,SB_72,SB_74,SB_85

Methods: gel filtration, enzyme assay, radio labeling
Comments, role: structure is not well-defined in paper

Related record ID(s): 108968, 122540, 122541, 122542
NCBI Taxonomy refs (TaxIDs): 4932
Reference(s) to other database(s): CCSD:9173, CBank-STR:8685
Show glycosyltransferases

There is only one chemically distinct structure:

CC(=O)N[C@H]1[C@H](O[C@@H]2[C@@H](CO)O[C@H](OP(=O)(O)OP(=O)(O)O)[C@H](NC(C)=O)[C@H]2O)O[C@H](CO)[C@@H](O[C@@H]2O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]2O)[C@@H]1O

SVG MW SMILES 3D mol Ionize

Structure type: oligomer
Location inside paper: DolPP-(GlcNAc)2(Man)9
Aglycon: dolichol
Compound class: N-glycan
Contained glycoepitopes: IEDB_123886,IEDB_130701,IEDB_135813,IEDB_136104,IEDB_137340,IEDB_137485,IEDB_140116,IEDB_140942,IEDB_141793,IEDB_141807,IEDB_141828,IEDB_141829,IEDB_141830,IEDB_141831,IEDB_143632,IEDB_144983,IEDB_150077,IEDB_151079,IEDB_151531,IEDB_152206,IEDB_153220,IEDB_164174,IEDB_429156,IEDB_540671,IEDB_548907,IEDB_857734,IEDB_983930,SB_136,SB_191,SB_196,SB_197,SB_198,SB_33,SB_44,SB_53,SB_67,SB_72,SB_73,SB_74,SB_77,SB_85

Methods: gel filtration, enzyme assay, radio labeling
Comments, role: structure is not well-defined in paper

Related record ID(s): 122539, 122541, 122542, 133588
NCBI Taxonomy refs (TaxIDs): 4932
Reference(s) to other database(s): CCSD:11522, CBank-STR:20384
Show glycosyltransferases

There is only one chemically distinct structure:

CC(=O)N[C@H]1[C@H](O[C@@H]2[C@@H](CO)O[C@H](OP(=O)(O)OP(=O)(O)O)[C@H](NC(C)=O)[C@H]2O)O[C@H](CO)[C@@H](O[C@@H]2O[C@H](CO[C@H]3O[C@H](CO[C@H]4O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]4O[C@H]4O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]4O)[C@@H](O)[C@H](O[C@H]4O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]4O[C@H]4O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]4O)[C@@H]3O)[C@@H](O)[C@H](O[C@H]3O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]3O[C@H]3O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]3O[C@H]3O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]3O)[C@@H]2O)[C@@H]1O

SVG MW SMILES 3D mol Ionize

Structure type: oligomer
Location inside paper: DolPP-(GlcNAc)2(Man)9(Glc)3
Compound class: N-glycan
Contained glycoepitopes: IEDB_123886,IEDB_130701,IEDB_135813,IEDB_136104,IEDB_137340,IEDB_137485,IEDB_140116,IEDB_140942,IEDB_141793,IEDB_141807,IEDB_141828,IEDB_141829,IEDB_141830,IEDB_141831,IEDB_142488,IEDB_143632,IEDB_144983,IEDB_144998,IEDB_146664,IEDB_151079,IEDB_151531,IEDB_152206,IEDB_153220,IEDB_164174,IEDB_232584,IEDB_429156,IEDB_540671,IEDB_548907,IEDB_857734,IEDB_983930,IEDB_983931,SB_136,SB_191,SB_192,SB_196,SB_197,SB_198,SB_33,SB_44,SB_53,SB_67,SB_72,SB_73,SB_74,SB_77,SB_85

Methods: gel filtration, enzyme assay, radio labeling
Comments, role: primary structure is not characterized in the paper. ST1 imported from Carbbank

Related record ID(s): 122539, 122540, 122542, 133535
NCBI Taxonomy refs (TaxIDs): 4932
Reference(s) to other database(s): GTC:G72079JG, CCSD:7002, CBank-STR:20905
Show glycosyltransferases

There is only one chemically distinct structure:

CC(=O)N[C@H]1[C@@H](O)O[C@H](CO)[C@@H](O[C@@H]2O[C@H](CO)[C@@H](O[C@@H]3O[C@H](CO[C@H]4O[C@H](CO[C@H]5O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]5O[C@H]5O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]5O)[C@@H](O)[C@H](O[C@H]5O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]5O[C@H]5O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]5O)[C@@H]4O)[C@@H](O)[C@H](O[C@H]4O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]4O[C@H]4O[C@H](CO)[C@@H](O)[C@H](O)[C@@H]4O[C@H]4O[C@H](CO)[C@@H](O)[C@H](O[C@H]5O[C@H](CO)[C@@H](O)[C@H](O[C@H]6O[C@H](CO)[C@@H](O)[C@H](O)[C@H]6O[C@H]6O[C@H](CO)[C@@H](O)[C@H](O)[C@H]6O)[C@H]5O)[C@@H]4O)[C@@H]3O)[C@H](O)[C@H]2NC(C)=O)[C@@H]1O

SVG MW SMILES 3D mol Ionize

Structure type: oligomer
Location inside paper: DolPP-(GlcNAc)2-DolPP-GlcNAc
Aglycon: dolichol
Contained glycoepitopes: IEDB_135813,IEDB_137340,IEDB_141807,IEDB_150077,IEDB_151531,SB_74,SB_85

Methods: gel filtration, enzyme assay, radio labeling
Comments, role: structure is not well-defined in paper

Related record ID(s): 122539, 122540, 122541
NCBI Taxonomy refs (TaxIDs): 4932
Reference(s) to other database(s): CCSD:29333, CBank-STR:3314
Show glycosyltransferases

There is only one chemically distinct structure:

CC(=O)N[C@H]1[C@H](O[C@@H]2[C@@H](CO)O[C@H](OP(=O)(O)OP(=O)(O)O)[C@H](NC(C)=O)[C@H]2O)O[C@H](CO)[C@@H](O)[C@@H]1O

SVG MW SMILES 3D mol Ionize