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Literature summary for 2.4.99.18 extracted from

  • Huang, C.; Mohanty, S.; Banerjee, M.
    A novel method of production and biophysical characterization of the catalytic domain of yeast oligosaccharyl transferase (2010), Biochemistry, 49, 1115-1126.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 2.4.99.18

Cloned(Commentary)

Cloned (Comment) Organism
PCR-amplification, expression of the pure C-terminal domain of the subunit Stt3p in Escherichia coli BL21 (DE3) Saccharomyces cerevisiae
subcloning, overexpression, and a method of production of the pure C-terminal domain of Stt3p at 60-70 mg/l in Escherichia coli Saccharomyces cerevisiae

Protein Variants

Protein Variants Comment Organism
D518E the replacement of a key residue, Asp518, located within the WWDYG signature motif (residues 516-520), leads to a distinct tertiary structure, even though both proteins have similar overall secondary structures, as demonstrated by CD, fluorescence and NMR spectroscopies. Asp518 plays a critical structural role, in addition to the catalytic role. The activity of the protein is confirmed by saturation transfer difference and nuclear magnetic resonance titration studies Saccharomyces cerevisiae
D518E within WWDYG signature motif leads to distinct tertiary structure interfering with function (complete loss of N-linked glycosylation activity), lethal Saccharomyces cerevisiae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
 additional information dissociation constant KD=9.97 mM Saccharomyces cerevisiae

Localization

Localization Comment Organism GeneOntology No. Textmining
endoplasmic reticulum
-
 Saccharomyces cerevisiae 5783
-

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
31500
-
 MALDI-TOF result in accordance with calculated results for histidine-tagged protein subunit Stt3p Saccharomyces cerevisiae

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
dolichyl diphosphooligosaccharide + protein L-asparagine Saccharomyces cerevisiae
-
 dolichyl disphosphate + protein with oligosaccharide attached to protein L-asparagine
-
 ?
dolichyl diphosphooligosaccharide + [protein]-L-asparagine Saccharomyces cerevisiae oligosaccharyl transferase (OT) is a multisubunit enzyme that catalyzes N-linked glycosylation of nascent polypeptides in the lumen of the endoplasmic reticulum dolichyl diphosphate + [protein]-N-oligosaccharidyl-L-asparagine
-
 ?

Organism

Organism UniProt Comment Textmining
Saccharomyces cerevisiae
-
-
-
Saccharomyces cerevisiae P39007
-
-

Purification (Commentary)

Purification (Comment) Organism
C-terminal domain of Stt3p, wild-type and mutant D518E Saccharomyces cerevisiae
dissolution in denaturing buffer (6 M guanidine hydrochloride, 500 mM Na Cl, 25 mM imidazole, 20 mM phosphate buffer, pH 7.4), cenrtrifugation, supernatant loaded onto nickel-NTA column for affinity chromatography with 20 mM phosphate buffer, pH 6.5, SDS-PAGE Saccharomyces cerevisiae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
dolichyl diphosphooligosaccharide + asparagine-asparagine-threonine-NH2 acceptor peptide
-
 Saccharomyces cerevisiae dolichyl disphosphate + protein with oligosaccharide attached to protein L-asparagine
-
 ?
dolichyl diphosphooligosaccharide + protein L-asparagine
-
 Saccharomyces cerevisiae dolichyl disphosphate + protein with oligosaccharide attached to protein L-asparagine
-
 ?
dolichyl diphosphooligosaccharide + [protein]-L-asparagine oligosaccharyl transferase (OT) is a multisubunit enzyme that catalyzes N-linked glycosylation of nascent polypeptides in the lumen of the endoplasmic reticulum Saccharomyces cerevisiae dolichyl diphosphate + [protein]-N-oligosaccharidyl-L-asparagine
-
 ?

Synonyms

Synonyms Comment Organism
yeast oligosaccharyl transferase
-
 Saccharomyces cerevisiae

General Information

General Information Comment Organism
malfunction congenital disorders of glycosylation (CDG) have severe effects in humans, complete loss is lethal for eukaryots Saccharomyces cerevisiae
physiological function N-linked glycosylation of nascent polypeptides in the lumen of the endoplasmic reticulum Saccharomyces cerevisiae