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Information on EC 2.4.1.186 - glycogenin glucosyltransferase and Organism(s) Oryctolagus cuniculus and UniProt Accession P13280

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EC Tree
     2 Transferases
         2.4 Glycosyltransferases
             2.4.1 Hexosyltransferases
                2.4.1.186 glycogenin glucosyltransferase
IUBMB Comments
The first reaction of this enzyme is to catalyse its own glucosylation, normally at Tyr-194 of the protein if this group is free. When Tyr-194 is replaced by Thr or Phe, the enzyme's Mn2+-dependent self-glucosylation activity is lost but its intermolecular transglucosylation ability remains . It continues to glucosylate an existing glucosyl group until a length of about 5--13 residues has been formed. Further lengthening of the glycogen chain is then carried out by EC 2.4.1.11, glycogen (starch) synthase. The enzyme is not highly specific for the donor, using UDP-xylose in addition to UDP-glucose (although not glucosylating or xylosylating a xylosyl group so added). It can also use CDP-glucose and TDP-glucose, but not ADP-glucose or GDP-glucose. Similarly it is not highly specific for the acceptor, using water (i.e. hydrolysing UDP-glucose) among others. Various forms of the enzyme exist, and different forms predominate in different organs. Thus primate liver contains glycogenin-2, of molecular mass 66 kDa, whereas the more widespread form is glycogenin-1, with a molecular mass of 38 kDa.
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Oryctolagus cuniculus
UNIPROT: P13280
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The taxonomic range for the selected organisms is: Oryctolagus cuniculus
The expected taxonomic range for this enzyme is: Eukaryota, Bacteria
Synonyms
glycogenin, glycogenin-1, glycogenin-2, priming glucosyltransferase, glycogenin 1, glycogenin 2, m-glycogenin, glycogenin glucosyltransferase, proglycogen synthase, udp-glucose protein transglucosylase, more
SYNONYM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glucosyltransferase, uridine diphosphoglucose-protein
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glucosyltransferase, uridine diphosphoglucose-protein 4-alpha-
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glycogen initiator synthase
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glycogenin
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glycogenin glycosyltransferase
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glyogenin
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priming glucosyltransferase
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proglycogen synthase
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UDP-glucose protein transglucosylase
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UDP-glucose-protein glucosyltransferase
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UDP-glucose:protein glucosyltransferase
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UDPGlc:protein transglucosylase
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UPTG
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uridine diphosphate glucose-protein transglucosylase I
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uridine diphosphoglucose protein transglucosylase I
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uridine diphosphoglucose-protein 4-alpha-glucosyltransferase
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uridine diphosphoglucose-protein glucosyltransferase
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REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
UDP-alpha-D-glucose + glycogenin = UDP + alpha-D-glucosylglycogenin
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexosyl group transfer
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PATHWAY SOURCE
PATHWAYS
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SYSTEMATIC NAME
IUBMB Comments
UDP-alpha-D-glucose:glycogenin alpha-D-glucosyltransferase
The first reaction of this enzyme is to catalyse its own glucosylation, normally at Tyr-194 of the protein if this group is free. When Tyr-194 is replaced by Thr or Phe, the enzyme's Mn2+-dependent self-glucosylation activity is lost but its intermolecular transglucosylation ability remains [7]. It continues to glucosylate an existing glucosyl group until a length of about 5--13 residues has been formed. Further lengthening of the glycogen chain is then carried out by EC 2.4.1.11, glycogen (starch) synthase. The enzyme is not highly specific for the donor, using UDP-xylose in addition to UDP-glucose (although not glucosylating or xylosylating a xylosyl group so added). It can also use CDP-glucose and TDP-glucose, but not ADP-glucose or GDP-glucose. Similarly it is not highly specific for the acceptor, using water (i.e. hydrolysing UDP-glucose) among others. Various forms of the enzyme exist, and different forms predominate in different organs. Thus primate liver contains glycogenin-2, of molecular mass 66 kDa, whereas the more widespread form is glycogenin-1, with a molecular mass of 38 kDa.
CAS REGISTRY NUMBER
COMMENTARY hide
117590-73-5
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SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
UDP-alpha-D-glucose + glycogenin
UDP + alpha-D-glucosylglycogenin
show the reaction diagram
autoglucosylation by glycogenin-1
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-
?
UDP-glucose + glycogenin
UDP + glucosylated glycogenin
show the reaction diagram
UDP-glucose + glycogenin
UDP + glucosylglycogenin
show the reaction diagram
self-glucosylation
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-
?
UDP-glucose + maltose
UDP + maltotriose
show the reaction diagram
trans-glucosylation. 93% of the transferred glucose molecules appears in maltotriose, 6% are attached to glycogenin, and 1% is liberated as free glucose
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-
?
UDP-xylose + glycogenin
UDP + xylosylated glycogenin
show the reaction diagram
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-
-
?
UDP-galactose + glycogenin
UDP + galactosylated glycogenin
show the reaction diagram
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autoglycosylation reaction
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?
UDP-glucose + glycogenin
UDP + glucosylated glycogenin
show the reaction diagram
UDP-glucose + N-(maltosyl-alpha-1,4-(1-deoxyglucitol))-peptide
UDP + glucosylated N-(maltosyl-alpha-1,4-(1-deoxyglucitol))-peptide
show the reaction diagram
UDP-glucose + n-dodecyl-beta-D-maltoside
UDP + n-dodecyl-beta-D-maltotriose
show the reaction diagram
additional information
?
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NATURAL SUBSTRATE
NATURAL PRODUCT
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
UDP-alpha-D-glucose + glycogenin
UDP + alpha-D-glucosylglycogenin
show the reaction diagram
autoglucosylation by glycogenin-1
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-
?
UDP-glucose + glycogenin
UDP + glucosylated glycogenin
show the reaction diagram
essential for the formation of glycogen granules, binds a chain of 5-13 glucose molecules at a specific tyrosine residue by autoglycosylation, catalyzes two chemically different autoglucosylation reactions, the glucosylation of a tyrosine hydroxyl group and the formation of alpha-1,4 glucosidic linkages by subsequent glucosylations
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-
?
UDP-glucose + glycogenin
UDP + glucosylated glycogenin
show the reaction diagram
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Mg2+
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less effective than Mn2+
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
ADP
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skeletal muscle enzyme, allosteric inhibition of autoglucosylation
ATP
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skeletal muscle enzyme, allosteric inhibition of autoglucosylation, complete inhibition at 5 mM, possible role as natural regulator
UDP
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inhibits both autoglucosylation and glucosylation of exogenous acceptor
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.002
UDP-glucose
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-
additional information
additional information
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Km-values for UDP-glucose in different tissues
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TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0053 - 0.0183
UDP-glucose
additional information
additional information
-
the catalytic effectiveness of glycogenin falls off dramatically as the average glucosyl chain length increases
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SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
additional information
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pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
additional information
-
enzyme activity does not depend on physiological state of the organism
Manually annotated by BRENDA team
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
additional information
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not membrane-bound
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Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
the glycogenin-1 mutation T82M causes glycogenosis. Substitution of Thr82 for serine but not for valine restores the maximum extent of autoglucosylation as well as transglucosylation and UDP-glucose hydrolysis rate, structure analysis, overview
physiological function
Asp162 is the residue involved in the activation step of the glucose transfer reaction mechanism
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION?
GLYG_RABIT
333
0
37397
Swiss-Prot
other Location (Reliability: 4)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
103000
wild-type enzyme in glycosylated and nonglycosylated form, full-length mutant enzymes D162S and D159S, gel filtration
34000
non-glycosylated truncated protein, gel filtration of a 0.0005 mM solution
38000
58000
truncation mutant enzymes DELTA270-332/D159S and DELTA270-332/D162S, gel filtration
59000
non-glycosylated truncated protein, gel filtration of a 0.02 mM solution
124000
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gel filtration, heterodimeric glycogen synthase complex
200000
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gel filtration, active proteoglycogen
37280
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amino acid sequence determination
38000
47000
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a glycogenin species of average molecular weight 47000 Da is isolated from proteoglycogen isoamylolyzed for 4.5 h, SDS-PAGE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
dimer
monomer
tetramer
mutant enzymes D159N, D162N and DELTA270-332/D162N exist as both tetrameric and dimeric species
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
glycoprotein
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
purified recombinant glycogenin-1 truncation mutant DELTA-270, hanging drop vapor diffusion method, the protein in 12% w/v, PEG monomethyl ester 5000, 0.1 M MES, pH 6.5, and 0.2 M ammonium sulfate, is mixed with 1 M ammonium sulfate and 0.1 M sodium phosphate, pH 6.7, 4°C, crystals are soaked in mother liquor containing 1 mM MnSO4 and 1 mM UDP or UDP-glucose, X-ray diffraction structure determination and analysis
the DELTA270 enzyme crystallizes at 7.5 mg/ml from solutions containing 1 mM UDP-glucose, 1 mM MnCl2, 100 mM sodium acetate, pH 4.5–4.7, and 10–13% (w/v) PEG 4000 in the space group P64, with cell dimensions a = b = 75.0, c = 233.4, gamma = 120° and a dimer in the asymmetric unit. All crystals are grown using hanging drop vapor diffusion methods at 23°C
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
D159N
exists as both tetrameric and dimeric species, compared to wild-type enzyme which exists to more than 95% as dimer, self-glucosylation activities below the limit of detection of the assay. Ability to catalyze the transglucosylation of maltose is reduced by 260fold, hydrolysis of UDP-glucose is reduced 12fold
D159S
stable enzyme, self-glucosylation activities below the limit of detection of the assay. Transglucosylation activity of the mutant enzyme is reduced to undetectable levels, activity for the hydrolysis of UDP-glucose is reduced 14fold
D162N
exists as both tetrameric and dimeric species, compared to wild-type enzyme which exists to more than 95% as dimer, self-glucosylation activities below the limit of detection of the assay, undetectable activity for the transglucosylation of maltose and the hydrolysis of UDP-glucose to free glucose
D162S
stable enzyme, self-glucosylation activities below the limit of detection of the assay. 30fold less active for the trans-glucosylation of maltose and 340fold less active for the hydrolysis of UDP-glucose
DELTA270-332
mutant enzyme is fully active, specific activity for self- or transglucosylation is indistinguishable from the full-length enzyme
DELTA270-332/D159S
inactive mutant enzyme
DELTA270-332/D162N
exists as both tetrameric and dimeric species, compared to wild-type enzyme which exists to more than 95% as dimer
DELTA270-332/D162S
18fold less active for the transglucosylation of maltose and 190fold less active for the hydrolysis of UDP-glucose than wild-type enzyme, activity for the hydrolysis of UDP-glucose is reduced 4fold
T82M
inactive mutant, the mutation is equivalent to T83M according to previous authors amino acid numbering, it causes glycogenosis showing the loss of Thr82 hydrogen bond to Asp162, the residue involved in the activation step of the glucose transfer reaction mechanism. Autoglucosylation, maltoside transglucosylation and UDP-glucose hydrolyzing activities are abolished
T83S
site-directed mutagenesis, the mutant is catalytically active
T83V
site-directed mutagenesis, inactive mutant
Y194F
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exchange of glucose attachment site, no autoglucosylation activity
Y194X
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mutation at Y194 leads to a protein unable to attach glucose to itself
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
concentrated to 8 mg/ml, and 100 microl aliquots are flash frozen in liquid nitrogen and stored at -80°C. Glycogenin stored in this manner is stable for at least 2 years.
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PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant proteins
recombinant wild-type gycogenin-1, its truncated mutant DELTA-270, and of glycogenin-1 point mutants from Escherichia coli strain CGSC 4997
wild-type and mutant enzymes
Ni-Sepharose column chromatography and Q Sepharose column chromatography
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nickel-NTA column and Q-Sepharose column
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separation from glycogen synthase, EC 2.4.1.11, by LiBr
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separation from proteoglycogen
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CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expression of wild-type and mutant enzymes in Saccharomyces cerevisiae
expression of wild-type gycogenin-1, its truncated mutant DELTA-270, and of glycogenin-1 point mutants in Escherichia coli strain CGSC 4997
preparation of a truncated already glucosylated His-tag fusion protein in Escherichia coli ER2566, expression of the truncated non-glucosylated His-tag fusion protein in Escherichia coli CGSC 4997
expressed in Escherichia coli strain Rosetta (DE3)
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expression in COS cells
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expression of mutant Y194F in Escherichia coli
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expresssion of recombinant protein in the CGSC Escherichia coli cell line 4997, which lacks UDP-glucose pyrophosphorylase activity
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functional expression of the wild-type enzyme in Escherichia coli
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APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
analysis
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development of assay method with n-dodecyl-beta-D-maltoside as substrate
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Pitcher, J.; Smythe, C.; Cohen, P.
Glycogenin is the priming glucosyltransferase required for the initiation of glycogen biogenesis in rabbit skeletal muscle
Eur. J. Biochem.
176
391-395
1988
Oryctolagus cuniculus
Manually annotated by BRENDA team
Meezan, E.; Manzella, S.; Roden, L.
Menage a trois: glycogenin, proteoglycan core protein xylosyltransferase and UDP-xylose
Trends Glycosci. Glycotechnol.
7
303-332
1995
Bos taurus, Coturnix sp., Gallus gallus, Mus musculus, Oryctolagus cuniculus, Rattus norvegicus
-
Manually annotated by BRENDA team
Pitcher, J.; Smythe, C.; Campell, D.G.; Cohen, P.
Identification of the 38-kDa subunit of rabbit skeletal muscle glycogen synthase as glycogenin
Eur. J. Biochem.
169
497-502
1987
Oryctolagus cuniculus
Manually annotated by BRENDA team
Carrizo, M.E.; Miozzo, M.C.; Goldraij, A.; Curtino, J.A.
Purification of rabbit skeletal muscle proteoglycogen: studies on the glucosyltransferase activity of polysaccharide-free and -bound glycogenin
Glycobiology
7
571-578
1997
Oryctolagus cuniculus
Manually annotated by BRENDA team
Viskupic, E.; Cao, Y.; Zhang, W.; Cheng, C.; DePaoli-Roach, A.A.; Roach, P.J.
Rabbit skeletal muscle glycogenin. Molecular cloning and production of fully functional protein in Escherichia coli
J. Biol. Chem.
267
25759-25763
1992
Bos taurus, Oryctolagus cuniculus, Rattus norvegicus
Manually annotated by BRENDA team
LOmako, J.; LOmako, W.M.; Whelan, W.J.
A self-glucosylating protein is the primer for rabbit muscle glycogen synthesis
FASEB J.
2
3097-3103
1988
Oryctolagus cuniculus
Manually annotated by BRENDA team
Cao, Y.; Mahrenholz, A.M.; DePaoli-Roach, A.A.; Roach, P.J.
Characterization of rabbit skeletal muscle glycogenin. Tyrosine 194 is essential for function
J. Biol. Chem.
268
14687-14693
1993
Oryctolagus cuniculus
Manually annotated by BRENDA team
Hurley, T.D.; Stout, S.; Miner, E.; Zhou, J.; Roach, P.J.
Requirements for catalysis in mammalian glycogenin
J. Biol. Chem.
280
23892-23899
2005
Oryctolagus cuniculus (P13280)
Manually annotated by BRENDA team
Gibbons, B.J.; Roach, P.J.; Hurley, T.D.
Crystal structure of the autocatalytic initiator of glycogen biosynthesis, glycogenin
J. Mol. Biol.
319
463-477
2002
Oryctolagus cuniculus
Manually annotated by BRENDA team
Hurley, T.D.; Walls, C.; Bennett, J.R.; Roach, P.J.; Wang, M.
Direct detection of glycogenin reaction products during glycogen initiation
Biochem. Biophys. Res. Commun.
348
374-378
2006
Oryctolagus cuniculus
Manually annotated by BRENDA team
Bazan, S.; Issoglio, F.M.; Carrizo, M.E.; Curtino, J.A.
The intramolecular autoglucosylation of monomeric glycogenin
Biochem. Biophys. Res. Commun.
371
328-332
2008
Oryctolagus cuniculus (P13280)
Manually annotated by BRENDA team
Romero, J.; Issoglio, F.; Carrizo, M.; Curtino, J.
Evidence for glycogenin autoglucosylation cessation by inaccessibility of the acquired maltosaccharide
Biochem. Biophys. Res. Commun.
374
704-708
2008
Oryctolagus cuniculus
Manually annotated by BRENDA team
Carrizo, M.; Romero, J.; Issoglio, F.; Curtino, J.
Structural and biochemical insight into glycogenin inactivation by the glycogenosis-causing T82M mutation
FEBS Lett.
586
254-257
2012
Oryctolagus cuniculus (P13280), Oryctolagus cuniculus
Manually annotated by BRENDA team