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Information on EC 2.4.1.40 - glycoprotein-fucosylgalactoside alpha-N-acetylgalactosaminyltransferase and Organism(s) Homo sapiens and UniProt Accession P16442
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2.4.1.40 glycoprotein-fucosylgalactoside alpha-N-acetylgalactosaminyltransferaseIUBMB Comments
Acts on blood group substance, and can use a number of 2-fucosyl-galactosides as acceptors.
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Word Map
- 2.4.1.40
- acetoacetate
- acetobutylicum
-
succinyl-coenzyme
-
3-oxoacids
-
2.8.3.5
-
solventogenesis
-
acetoacetyl-coenzyme
- synthesis
The taxonomic range for the selected organisms is: Homo sapiens
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
The expected taxonomic range for this enzyme is: Bacteria, Eukaryota
Reaction Schemes
Synonyms
a transferase, a-transferase, glycosyltransferases a, alpha-3-n-acetylgalactosaminyltransferase, blood group a glycosyltransferase, histo-blood group a transferase, 
more
topSYNONYM 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
A-transferase
-
-
-
-
alpha-3-N-acetylgalactosaminyltransferase
-
-
-
-
blood-group substance A-dependent acetylgalactosaminyltransferase
-
-
-
-
blood-group substance alpha-acetyltransferase
-
-
-
-
fucosylgalactose acetylgalactosaminyltransferase
-
-
-
-
histo-blood group A acetylgalactosaminyltransferase
-
-
-
-
histo-blood group A glycosyltransferase (Fucalpha1-2Galalpha1-3-N-acetylgalactosaminyltransferase)
-
-
-
-
histo-blood group A transferase
-
-
-
-
UDP-GalNAc:Fucalpha1-2Galalpha1-3-N-acetylgalactosaminyltransferase
-
-
-
-
UDP-N-acetyl-D-galactosamine:alpha-L-fucosyl-1,2-D-galactose 3-N-acetyl-D-galactosaminyltransferase
-
-
-
-
additional information
-
the A/B transferases, ABO, belong to the GT6 enzyme family
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
hexosyl group transfer
-
-
-
-
SYSTEMATIC NAME
IUBMB Comments
UDP-N-acetyl-D-galactosamine:glycoprotein-alpha-L-fucosyl-(1->2)-D-galactose 3-N-acetyl-D-galactosaminyltransferase
Acts on blood group substance, and can use a number of 2-fucosyl-galactosides as acceptors.
CAS REGISTRY NUMBER
COMMENTARY
9067-69-0
-
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
UDP-N-acetyl-alpha-D-galactosamine + alpha-Fuc-(1->2)-beta-D-Gal-(CH2)7CH3
UDP + N-acetyl-alpha-D-galactosaminyl-(1->3)-[alpha-Fuc-(1->2)]-beta-D-Gal-(CH2)7CH3
-
-
-
?
UDP-N-acetyl-D-galactosamine + alpha-L-fucosyl-1-2-D-galactosyl-O-R
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1-2]-D-galactosyl-O-R
UDP-N-acetyl-D-galactosamine + alpha-L-Fucp-(1,2)-beta-D-Galp-(1,3)-beta-DGlcNAcp-O(CH2)7CH3
?
41% of the activity with alpha-L-Fucp-(1,2)-beta-D-Galp-O(CH2)7CH3
-
-
?
UDP-N-acetyl-D-galactosamine + alpha-L-Fucp-(1,2)-beta-D-Galp-(1,4)-beta-D-GlcNAcp-O-(CH2)8CO2CH3
?
79% of the activity with alpha-L-Fucp-(1,2)-beta-D-Galp-O(CH2)7CH3
-
-
?
UDP-N-acetyl-D-galactosamine + alpha-L-Fucp-(1,2)-beta-D-Galp-O(CH2)7CH3
UDP + alpha-D-GalNAc-(1,3)-[alpha-(Fucp-(1,2)])-beta-D-Galp-O(CH2)7CH3
-
-
-
?
UDP-N-acetyl-D-galactosamine + L-fucosyl-alpha-1,2-beta-galactosyl-O(CH2)7CH3
UDP + alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-beta-D-galactosyl-O(CH2)7CH3
-
-
-
?
UDP-N-acetyl-D-glucosamine + L-fucosyl-alpha-1,2-beta-glucosyl-O(CH2)7CH3
UDP + N-acetyl-alpha-D-glucosaminyl-1,3-[alpha-L-fucosyl-1,2]-beta-D-glucosyl-O(CH2)7CH3
-
-
-
?
UDP-N-acetyl-alpha-D-galactosamine + alpha-Fuc-(1->2)-beta-D-Gal-(CH2)7CH3
UDP + alpha-D-GalNAc-(1->3)-[alpha-Fuc-(1->2)]-beta-D-Gal-(CH2)7CH3
-
-
-
?
UDP-N-acetyl-D-galactosamine + 2'-fucosyllactose
UDP + N-acetyl-alpha-D-galactosaminyl-2'-fucosyllactose
-
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
UDP-N-acetyl-D-galactosamine + glycoprotein-alpha-L-fucosyl-(1,2)-D-galactose
UDP + glycoprotein-N-acetyl-alpha-D-galactosaminyl-(1,3)-[alpha-L-fucosyl-(1,2)]-D-galactose
UDP-N-acetyl-D-galactosamine + L-fucosyl-alpha-1,2-beta-galactosyl-O(CH2)7CH3
UDP + alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-beta-D-galactosyl-O(CH2)7CH3
-
-
-
-
?
UDP-N-acetyl-D-galactosamine + lacto-N-fucopentaose I
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[lacto-N-fucopentaose I]
-
i.e. Fucalpha(1-2)Galbeta(1-3)GlcNAcbeta(1-3)Galbeta(1-4)Glc
-
-
?
UDP-N-acetyl-D-glucosamine + L-fucosyl-alpha-1,2-beta-glucosyl-O(CH2)7CH3
UDP + N-acetyl-alpha-D-glucosaminyl-1,3-[alpha-L-fucosyl-1,2]-beta-D-glucosyl-O(CH2)7CH3
-
-
-
-
?
UDP-N-acetylgalactosamine + octyl 3-O-methyl-alpha-L-fucopyranosyl-(1-2)-beta-D-galactopyranoside
UDP + N-acetyl-alpha-D-galactosamine-1,3-[3-O-methyl-alpha-L-fucosyl-1,2]-beta-D-galactosyl-O(CH2)7CH3
-
-
-
-
?
UDP-N-acetylgalactosamine + octyl 4-O-methyl-alpha-L-fucopyranosyl-(1-2)-beta-D-galactopyranoside
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[4-O-methyl-alpha-L-fucosyl-1,2]-beta-D-galactosyl-O(CH2)7CH3
-
-
-
-
?
UDP-N-acetylgalactosamine + octyl alpha-L-xylo-hexopyranosyl-(1-2)-beta-D-galactopyranoside
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-xylo-hexopyranosyl-1,2]-beta-D-galactosyl-O(CH2)7CH3
-
-
-
-
?
UDP-N-acetylgalactosamine + octyl beta-D-arabinopyranosyl-(1-2)-beta-D-galactopyranoside
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[beta-D-arabinopyranosyl-1,2-beta-D-galactosyl-O(CH2)7CH3]
-
-
-
-
?
UDP-N-acetyl-D-galactosamine + alpha-L-fucosyl-1-2-D-galactosyl-O-R
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1-2]-D-galactosyl-O-R
H-antigen disaccharide
-
-
?
UDP-N-acetyl-D-galactosamine + alpha-L-fucosyl-1-2-D-galactosyl-O-R
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1-2]-D-galactosyl-O-R
H-antigen disaccharide, Leu226 is responsible for substrate specificity for UDP-N-acetyl-D-galactosamine as donor substrate, acceptor substrate binding pocket structure, overview
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
-
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein alpha-L-fucosyl-1,2-D-galactose
UDP + glycoprotein N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1,2]-D-galactose
-
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein-alpha-L-fucosyl-(1,2)-D-galactose
UDP + glycoprotein-N-acetyl-alpha-D-galactosaminyl-(1,3)-[alpha-L-fucosyl-(1,2)]-D-galactose
-
substrate is the histo-blood H group antigen
product is the histo-blood A group antigen
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein-alpha-L-fucosyl-(1,2)-D-galactose
UDP + glycoprotein-N-acetyl-alpha-D-galactosaminyl-(1,3)-[alpha-L-fucosyl-(1,2)]-D-galactose
-
substrate is the histo-blood H group antigen. Arg176 of the enzyme is not important in determining the sugar specificity of enzyme, while Gly235 is influential and Leu266 and Gly268 are crucial in the determination of GalNAc/galactose specificity
product is the histo-blood A group antigen
-
?
additional information
?
-
transfers N-acetylgalactosamine from UDP-N-acetylgalactosamine to H-active structures to form A determinants
-
-
?
additional information
?
-
the enzyme catalyzes the final step in the enzymatic synthesis of the ABO(H) blood group A antigen, overview
-
-
?
additional information
?
-
substrate binding structure by wild-type and mutant GTA, overview
-
-
?
additional information
?
-
-
enzyme also catalyzes the transfer of galactose in alpha-linkage to 2'-fucosyllactose, the transfer rate of galactose is much lower than that of N-acetylgalactosamine
-
-
?
additional information
?
-
-
effective acceptor substrates contain a subterminal beta-galactosyl residue substituted at the O-2 position with fucose
-
-
?
additional information
?
-
-
transfers N-acetylgalactosamine from UDP-N-acetylgalactosamine to H-active structures to form A determinants
-
-
?
additional information
?
-
-
galactose is used as an acceptor analogue and UDP as a donor analogue in all soaking trials
-
-
?
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
UDP-N-acetyl-D-galactosamine + alpha-L-fucosyl-1-2-D-galactosyl-O-R
UDP + N-acetyl-alpha-D-galactosaminyl-1,3-[alpha-L-fucosyl-1-2]-D-galactosyl-O-R
H-antigen disaccharide
-
-
?
UDP-N-acetyl-D-galactosamine + glycoprotein-alpha-L-fucosyl-(1,2)-D-galactose
UDP + glycoprotein-N-acetyl-alpha-D-galactosaminyl-(1,3)-[alpha-L-fucosyl-(1,2)]-D-galactose
-
substrate is the histo-blood H group antigen
product is the histo-blood A group antigen
-
?
additional information
?
-
transfers N-acetylgalactosamine from UDP-N-acetylgalactosamine to H-active structures to form A determinants
-
-
?
additional information
?
-
the enzyme catalyzes the final step in the enzymatic synthesis of the ABO(H) blood group A antigen, overview
-
-
?
additional information
?
-
-
transfers N-acetylgalactosamine from UDP-N-acetylgalactosamine to H-active structures to form A determinants
-
-
?
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
Mn2+
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
alpha-L-fucosyl-1-2-beta-D-(3-amino)-galactosyl-O-R
complex mode inhibitor, binding structure, Leu226 is involved by interacting with the C3-position group
alpha-L-fucosyl-1-2-beta-D-(3-deoxy)-galactosyl-O-R
competitive inhibitor, binding structure, Leu226 is involved by interacting with the C3-position group
octyl 3'-amino-3'-deoxy-alpha-L-fucopyranosyl-(1-2)-beta-D-galactopyranoside
-
98% inhibition at 0.025 mM
octyl alpha-L-fucopyranosyl-(1-2)-beta-D-gulopyranoside
-
36% inhibition at 0.025 mM
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1.3
alpha-L-Fucp-(1,2)-beta-D-Galp-(1,3)-beta-DGlcNAcp-O(CH2)7CH3
37°C, pH 7.0
0.49
alpha-L-Fucp-(1,2)-beta-D-Galp-(1,4)-beta-D-GlcNAcp-O-(CH2)8CO2CH3
37°C, pH 7.0
additional information
additional information
kinetics of recombinant wild-type GTA 53-354 and of GTA/GTB mutant chimeric enzymes
-
0.167
L-fucosyl-alpha-1,2-beta-galactosyl-O(CH2)7CH3
G176R/P234S/S235G/M266L/A268G mutant
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
1.3
alpha-L-Fucp-(1,2)-beta-D-Galp-(1,3)-beta-DGlcNAcp-O(CH2)7CH3
37°C, pH 7.0
1.8
alpha-L-Fucp-(1,2)-beta-D-Galp-(1,4)-beta-D-GlcNAcp-O-(CH2)8CO2CH3
37°C, pH 7.0
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
IMAGE
0.0002
octyl 3'-amino-3'-deoxy-alpha-L-fucopyranosyl-(1-2)-beta-D-galactopyranoside
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
ORGANISM
COMMENTARY
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
489115, 659283, 674703, 693025, 718520, 756450, 756474, 756888, 756889, 756977, 757247, 758454, 758457
Uniprot
only four amino acids are responsible for B and A blood group enzymes R176G, G235S, L266M and G268A, the latter two amino acids are responsible for the difference in donor specificity and the first two have roles in acceptor binding and turnover
Uniprot
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SOURCE
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
physiological function
physiological function
the deletion of exon 3 or 4, but not of exon 2 or 5, of human GTA transcripts bestows moderate Forssman glycolipid synthase activity. Strong Forssman glycolipid synthase activity is attained, in addition to GTA and GTB activities, by cointroducing one of those deletions and the replacement of the LeuGlyGly tripeptide sequence at codons 266 to 268 by GlyGlyAla substitution, i.e. to the residues of Forssman glycolipid synthase
physiological function
when Von Willebrand factor expressing H-antigen (H-VWF) from blood group O and bovine serum albumin conjugated with H-antigen (H-BSA) are incubated with recombinant alpha1-3-N-acetylgalactosaminyltransferase (rA-transferase) and A-plasma and and UDP-GalNAc, both H-VWF and H-BSA acquire the A-antigen. Incubation with A-plasma very weakly converts the H-antigen on BSA and Von Willebrand factor only in the presence of supplemented UDP-GalNAc. This conversion is enhanced on desialylation of H-VWF
UNIPROT
ENTRY NAME
ORGANISM
NO. OF AA
NO. OF TRANSM. HELICES
MOLECULAR WEIGHT[Da]
SOURCE
SEQUENCE
LOCALIZATION PREDICTION
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable).
The reliability class of the localization prediction ranges from 1 (very reliable) to 5 (not reliable). BGAT_HUMAN
354
1
40934
Swiss-Prot
Secretory Pathway (Reliability: 2)
PDB
SCOP
CATH
UNIPROT
ORGANISM
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
SUBUNIT
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
additional information
structure analysis of GTA/GTB chimeric enzymes GTB/G176R and GTB/G176R/G235S bound to a panel of donor and acceptor analogue substrates, open, semi-closed, and closed conformations, overview
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
CRYSTALLIZATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
a large single crystal is subjected to H/D exchange prior to data collection and time-of-flight neutron diffraction data is collected to 2.5 A resolution at the Protein Crystallography Station to 85% overall completeness, with complementary X-ray diffraction data collected from a crystal from the same drop and extending to 1.85 A resolution
assignment of all methyl resonance signals in Ala, Ile, Leu, Met and Val labeled samples of GTA and GTB by lanthanide-induced pseudocontact shifts and methyl-methyl NOESY. The fully closed state is not adopted in the presence of lanthanide ions
crystals of purified native enzyme are soaked with various combinations of UDP-GalNAc, UDP-Gal, UDP, and acceptor analogues alpha-L-fucosyl-1-2-beta-D-(3-deoxy)-galactosyl-O-R or alpha-L-fucosyl-1-2-beta-D-(3-amino)-galactosyl-O-R, ligands are solved in 7.5% PEG 4000, 15% glycerol, 75 mM N-[2-acetamido]-2-iminodiacetic acid, pH 7.5, 10 mM MnCl2, and 10 mM inhibitor, 3-4 days, X-ray diffraction structure determination and analysis at 2.1 A resolution
enzyme adopts an open conformation in the absence of substrates. Binding of UDP induces a semiclosed conformation. In the presence of both donor and acceptor substrates, the enzymes shift towards a closed conformation with ordering of an internal loop and the C-terminal residues, which then completely cover the donor-binding pocket. The enzyme shows substantial plasticity and conformational flexibility. Residues Ile123 at the bottom of the UDP binding pocket, and Ile192 as part of the internal loop are significantly disturbed upon stepwise addition of UDP and H-disaccharide-O-CH3
enzyme soaked with acceptor analogs: galactose, lactose, N-acetyllactosamine, beta-D-Galp-O(CH2)8CO2CH3, alpha-L-Fucp-(1,2)-beta-D-Galp-O(CH2)7CH3, beta-D-Galp-(1,4)-beta-D-Glcp-OCH3, alpha-L-Fucp-(1,2)-beta-D-Galp-(1,3)-beta-D-GlcNAcp-O(CH2)7CH3, alpha-L-Fucp-(1,2)-beta-D-Galp-(1,4)-beta-D-GlcNAcp-O-(CH2)8CO2CH3
G176R/P234S/S235G/M266L/A268G-mutant with and without H-antigen, at 1.55 and 1.65 A resolution respectively
Methyl-TROSY-based titration experiments in combination with zz-exchange experiments show dramatic changes of binding kinetics associated with allosteric interactions between donor-type and acceptor-type ligands. Binding of the acceptor substrates H-disaccharide, H-type II trisaccharide, and H-type VI trisaccharide affects the chemical shifts of the 13C-methyl groups of Met 266, Val 299, Leu 324, and Leu 329, which belong to the acceptor substrate binding pocket. Depending on substrate concentrations in the Golgi apparatus an acceptor route and a donor route are possible. At high local concentrations of UDP-Gal or UDP-GalNAc binding of the nucleotide sugar to GTB or GTA would precede binding of the H-antigen. At low nucleotide sugar concentrations, it can be assumed that H-antigen binds first. In this latter case, the enzymes may discriminate between different types of H-antigens, preferring e. g. type-II over type-I H-antigens
of catalytic domain residues 63-354, with and without L-fucosyl-alpha-1,2-beta-galactosyl-O(CH2)7CH3-acceptor and UDP, at 1.35 and 1.8 A resolution respectively
purified recombinant GTA/GTB mutant chimeric enzymes, complexing with synthetic antigen disaccharides or UDP, hanging drop vapour diffusion method, different solutions for the different chimeric mutants, X-ray diffraction structure determinationand analysis at 1.41-1.75 A resolution, overview, structure modelling
structures of isoforms GTA and GTB in complex with their respective trisaccharide products. A conflict exists between the transferred sugar monosaccharide and the beta-phosphate of the UDP donor. The mechanism of product release shows monosaccharide transfer to the H-antigen acceptor induces active site disorder and ejection of the UDP leaving group prior to trisaccharide egress
structures of GTA, GTB and several chimeras determined by single-crystal X-ray diffraction demonstrate a range of susceptibility to the choice of cryoprotectant, in which the mobile polypeptide loops can be induced by glycerol to form the ordered closed conformation associated with substrate recognition and by MPD (hexylene glycol, 2-methyl-2,4-pentanediol) to hinder binding of substrate in the active site owing to chelation of the Mn2+ cofactor and thereby adopt the disordered open state
-
structures of wild-type and mutant D302C. Conserved active site residues Arg188 and Asp302 are critical for catalysis, and disruption of their hydrogen bond network through mutation can dramatically decrease enzymatic activity
PROTEIN VARIANTS
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
E303A
residue E303 plays a critical role in maintaining the stability of a strained double-turn in the active site through several hydrogen bonds
E303C
residue E303 plays a critical role in maintaining the stability of a strained double-turn in the active site through several hydrogen bonds. Mutant retains significant activity despite disrupted active site architecture
E303D
residue E303 plays a critical role in maintaining the stability of a strained double-turn in the active site through several hydrogen bonds. Mutant retains significant activity despite disrupted active site architecture
E303Q
residue E303 plays a critical role in maintaining the stability of a strained double-turn in the active site through several hydrogen bonds
G176R/P234S/S235G/M266L/A268G
expressed in Escherichia coli BL21, although 4 of the mutations correspond to a change from A- to B-blood group, the P234S-mutation shows a remarkable increase in A-donor specificity
additional information
additional information
construction of GTA/GTB chimeric enzymes GTB/G176R and GTB/G176R/G235S
additional information
amino acids at codons 266 and 268 of human isoforms GTA, EC 2.4.1.40, and GTB, EC 2.4.1.37, are crucial to their distinct sugar specificities. In vitro mutagenized GTAs/GTBs having any of 20 possible amino acids at those codons show that those codons determine the transferase activity and sugar specificity
additional information
expression of constructs, having either of AlaGlyGly, GlyGlyAla, HisAlaAla, LeuGlyGly, or MetGlyAla tripeptide sequence at codons corresponding to 266-268 of human GTA/GTB. The original human GTA construct with LeuGlyGly and the substitution construct with AlaGlyGly exhibit strong GTA. Human GTA constructs with GlyGlyAla, HisAlaAla, LeuGlyGly, or MetGlyAla exhibit GTA, GTA/GTB, GTB, GTA, or GTB activity, respectively
additional information
preparation of human GTA derivative constructs containing any of the 20 amino acids at codon 69 with and without a GlyGlyAla substitution of the LeuGlyGly tripeptide at codons 266-268. In presence of the Forssman glycolipid synthase substrate, all substitution constructs at codon 69 exhibit Forssman glycolipid synthase activity. The combination with tripeptide GlyGlyAla substitution significantly increases the activity. With increased globoside availability, the native human GTA with a methionine residue at codon 69 also synthesizes Forssman antigen
additional information
-
preparation of human GTA derivative constructs containing any of the 20 amino acids at codon 69 with and without a GlyGlyAla substitution of the LeuGlyGly tripeptide at codons 266-268. In presence of the Forssman glycolipid synthase substrate, all substitution constructs at codon 69 exhibit Forssman glycolipid synthase activity. The combination with tripeptide GlyGlyAla substitution significantly increases the activity. With increased globoside availability, the native human GTA with a methionine residue at codon 69 also synthesizes Forssman antigen
additional information
the enzymes GTA, EC 2.4.1.40, and GTB have nearly identical sequences, while the corresponding mutants of GTA/GTB have up to a 13fold difference in their residual activities relative to wild type.The mutated Cys, Asp and Gln functional groups are no more than 0.8 A further from the anomeric carbon of donor substrate compared to wild type
additional information
-
generation of a chimeric histo-blood group B transferase by replacing the N-acetyl-D-galactosamine recognition domain of human A transferase with the galactose-recognition domain of evolutionarily related murine alpha1,3-galactosyltransferase, the chimeric mutant shows a functional conversion from A to B transferase activity, overview. The AT-MluI-GT construct, which expresses the chimera between the long N-terminal sequence up to the MluI site of human A transferase, and the short C-terminal sequence up to the MluI site of murine a1-3Gal transferase, produces the antigens that react with both the anti-A and the Griffonia simplicifolia Lectin I, GSL-IA4, lectin
additional information
-
chimeric GTA and GTB enzymes are generated (AABB, ABBA and ABBB)
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY
LITERATURE
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
4°C, 0.05 M Tris-HCl, pH 7.4, 2 mM MnCl2, 1 mM EDTA, 0.01% w/v Triton X-100, 0.2 M NaCl, 1 mM dithiothreitol, 0.03% w/v NaN3, 10% loss of activity per month
-
PURIFICATION (Commentary)
ORGANISM
UNIPROT
LITERATURE
recombinant wild-type GTA 53-354 and of GTA/GTB mutant chimeric enzymes from Escherichia coli
using a two-step protocol consisting of ion-exchange chromatography followed by UDP-hexanolamine affinity chromatography
CLONED (Commentary)
ORGANISM
UNIPROT
LITERATURE
expressed in Escherichia coli BL21, P234S-mutant of blood-group substance B-dependent galactosyltransferase
expression of truncated wild-type GTA 53-354 and of GTA/GTB mutant chimeric enzymes in Escherichia coli
human GTA with the N-terminal transmembrane domain truncated at residue Met63 and codon-optimized for BL21 Escherichia coli is expressed and purified from cultures using a two-step protocol consisting of ion-exchange chromatography followed by UDP-hexanolamine affinity chromatography
REF.
AUTHORS
TITLE
JOURNAL
VOL.
PAGES
YEAR
ORGANISM (UNIPROT)
PUBMED ID
SOURCE
Marcus, S.L.; Polakowski, R.; Seto, N.O.L.; Leinala, E.; Borisova, S.; Blancher, A.; Roubinet, F.; Evans, S.V.; Palcic, M.M.
A single point mutation reverses the donor specificity of human blood group B-synthesizing galactosyltransferase
J. Biol. Chem.
278
12403-12405
2003
Homo sapiens (P16442)
Patenaude, S.I.; Seto, N.O.L.; Borisova, S.N.; Szpacenko, A.; Marcus, S.L.; Palcic, M.M.; Evans, S.V.
The structural basis for specificity in human ABO(H) blood group biosynthesis
Nat. Struct. Biol.
9
685-690
2002
Homo sapiens (P16442)
Seto, N.O.L.; Compston, C.A.; Szpacenko, A.; Palcic, M.M.
Enzymatic synthesis of blood group A and B trisaccharide analogues
Carbohydr. Res.
324
161-169
2000
Homo sapiens
Mukherjee, A.; Palcic, M.M.; Hindsgaul, O.
Synthesis and enzymatic evaluation of modified acceptors of recombinant blood group A and B glycosyltransferases
Carbohydr. Res.
326
1-21
2000
Homo sapiens
Seto, N.O.; Compston, C.A.; Evans, S.V.; Bundle, D.R.; Narang, S.A.; Palcic, M.M.
Donor substrate specificity of recombinant human blood group A, B and hybrid A/B glycosyltransferases expressed in Escherichia coli
Eur. J. Biochem.
259
770-775
1999
Homo sapiens
Seto, N.O.L.; Palcic, M.M.; Compston, C.A.; Li, H.; Bundle, D.R.; Narang, S.A.
Sequential interchange of four amino acids from blood group B to blood group A glycosyltransferase boosts catalytic activity and progressively modifies substrate recognition in human recombinant enzymes
J. Biol. Chem.
272
14133-14138
1997
Homo sapiens
Lowary, T.L.; Hindsgaul, O.
Recognition of synthetic O-methyl, epimeric, and amino analogues of the acceptor alpha-L-Fuc p-(1-2)-beta-D-Gal p-OR by the blood-group A and B gene-specified glycosyltransferases
Carbohydr. Res.
251
33-67
1994
Homo sapiens
Nakajima, T.; Furukawa, K.; Takenaka, O.
Blood group A and B glycosyltransferase in nonhuman primate plasma
Exp. Clin. Immunogenet.
10
21-30
1993
Macaca fascicularis, Homo sapiens, Pan troglodytes
Yazawa, S.; Nakajima, T.; Kameyama, N.; Saga, K.I.; Tachikawa, T.
An enzyme-linked immunosorbent assay for blood-group A and B enzymes
Carbohydr. Res.
239
329-335
1993
Homo sapiens
David, L.; Leitao, D.; Sobrinho-Simoes, M.; Bennett, E.P.; White, T.; Mandel, U.; Dabelsteen, E.; Clausen, H.
Biosynthetic basis of incompatible histo-blood group A antigen expression: anti-A transferase antibodies reactive with gastric cancer tissue of type O individuals
Cancer Res.
53
5494-5500
1993
Homo sapiens
Navaratnam, N.; Findlay, J.B.C.; Keen, J.N.; Watkins, W.M.
Purification, properties and partial amino acid sequence of the blood-group-A-gene-associated alpha-3-N-acetylgalactosaminyltransferase from human gut mucosal tissue
Biochem. J.
271
93-98
1990
Homo sapiens
Takeya, A.; Hosomi, O.; Ishiura, M.
Complete purification and characterization of alpha-3-N-acetylgalactosaminyltransferase encoded by the human blood group A gene
J. Biochem.
107
360-368
1990
Homo sapiens
Clausen, H.; White, T.; Takio, K.; Titani, K.; Stroud, M.; Holmes, E.; Karkov, J.; Thim, L.; Hakomori, S.
Isolation to homogeneity and partial characterization of a histo-blood group A defined Fuc alpha 1-2Gal alpha 1-3-N-acetylgalactosaminyltransferase from human lung tissue
J. Biol. Chem.
265
1139-1145
1990
Homo sapiens
Nagai, M.; Dave, V.; Kaplan, B.E.; Yoshida, A.
Human blood group glycosyltransferases. I. Purification of N-acetylgalactosaminyltransferase
J. Biol. Chem.
253
377-379
1987
Homo sapiens
Kobata, A.; Grollman, E.F.; Ginsburg, V.
An enzymic basis for blood type A in humans
Arch. Biochem. Biophys.
124
609-612
1968
Homo sapiens
Nguyen, H.P.; Seto, N.O.L.; Cai, Y.; Leinala, E.K.; Borisova, S.N.; Palcic, M.M.; Evans, S.V.
The influence of an intramolecular hydrogen bond in differential recognition of inhibitory acceptor analogs by human ABO(H) blood group A and B glycosyltransferases
J. Biol. Chem.
278
49191-49195
2003
Homo sapiens (P16442)
Letts, J.A.; Rose, N.L.; Fang, Y.R.; Barry, C.H.; Borisova, S.N.; Seto, N.O.; Palcic, M.M.; Evans, S.V.
Differential recognition of the type I and II H antigen acceptors by the human ABO(H) blood group A and B glycosyltransferases
J. Biol. Chem.
281
3625-3632
2006
Homo sapiens (P16442), Homo sapiens
Alfaro, J.A.; Zheng, R.B.; Persson, M.; Letts, J.A.; Polakowski, R.; Bai, Y.; Borisova, S.N.; Seto, N.O.; Lowary, T.L.; Palcic, M.M.; Evans, S.V.
ABO(H) blood group A and B glycosyltransferases recognize substrate via specific conformational changes
J. Biol. Chem.
283
10097-10108
2008
Homo sapiens (P16442)
Yamamoto, F.; Yamamoto, M.; Blancher, A.
Generation of histo-blood group B transferase by replacing the N-acetyl-D-galactosamine recognition domain of human A transferase with the galactose-recognition domain of evolutionarily related murine alpha1,3-galactosyltransferase
Transfusion
50
622-630
2009
Homo sapiens
Johal, A.; Schuman, B.; Alfaro, J.; Borisova, S.; Seto, N.; Evans, S.
Sequence-dependent effects of cryoprotectants on the active sites of the human ABO(H) blood group A and B glycosyltransferases
Acta Crystallogr. Sect. D
68
268-276
2012
Homo sapiens
Schuman, B.; Fisher, S.; Kovalevsky, A.; Borisova, S.; Palcic, M.; Coates, L.; Langan, P.; Evans, S.
Preliminary joint neutron time-of-flight and X-ray crystallographic study of human ABO(H) blood group A glycosyltransferase
Acta Crystallogr. Sect. F
67
258-262
2011
Homo sapiens (P16442), Homo sapiens
Grimm, L.; Weissbach, S.; Fluegge, F.; Begemann, N.; Palcic, M.; Peters, T.
Protein NMR studies of substrate binding to human blood group A and B glycosyltransferases
ChemBioChem
18
1260-1269
2017
Homo sapiens (P16442)
Fluegge, F.; Peters, T.
Insights into allosteric control of human blood group A and B glycosyltransferases from dynamic NMR
ChemistryOpen
8
760-769
2019
Homo sapiens (P16442)
Blackler, R.J.; Gagnon, S.M.; Polakowski, R.; Rose, N.L.; Zheng, R.B.; Letts, J.A.; Johal, A.R.; Schuman, B.; Borisova, S.N.; Palcic, M.M.; Evans, S.V.
Glycosyltransfer in mutants of putative catalytic residue Glu303 of the human ABO(H) A and B blood group glycosyltransferases GTA and GTB proceeds through a labile active site
Glycobiology
27
370-380
2017
Homo sapiens (P16442)
Gagnon, S.M.L.; Legg, M.S.G.; Sindhuwinata, N.; Letts, J.A.; Johal, A.R.; Schuman, B.; Borisova, S.N.; Palcic, M.M.; Peters, T.; Evans, S.V.
High-resolution crystal structures and STD NMR mapping of human ABO(H) blood group glycosyltransferases in complex with trisaccharide reaction products suggest a molecular basis for product release
Glycobiology
27
966-977
2017
Homo sapiens (P16442)
Gagnon, S.; Legg, M.; Polakowski, R.; Letts, J.; Persson, M.; Lin, S.; Zheng, R.; Rempel, B.; Schuman, B.; Haji-Ghassemi, O.; Borisova, S.; Palcic, M.; Evans, S.
Conserved residues Arg188 and Asp302 are critical for active site organization and catalysis in human ABO(H) blood group A and B glycosyltransferases
Glycobiology
28
624-636
2018
Homo sapiens (P16422)
Kano, T.; Kondo, K.; Hamako, J.; Matsushita, F.; Sakai, K.; Matsui, T.
Effects of plasma glycosyltransferase on the ABO(H) blood group antigens of human von Willebrand factor
Int. J. Hematol.
108
139-144
2018
Homo sapiens (P16442)
Fluegge, F.; Peters, T.
Complete assignment of Ala, Ile, Leu, Met and Val methyl groups of human blood group A and B glycosyltransferases using lanthanide-induced pseudocontact shifts and methyl-methyl NOESY
J. Biomol. NMR
70
245-259
2018
Homo sapiens (P16442)
Cid , E.; Yamamoto, M.; Yamamoto, F.
Amino acid substitutions at sugar-recognizing codons confer ABO blood group system-related alpha1,3 Gal(NAc) transferases with differential enzymatic activity
Sci. Rep.
9
846
2019
Homo sapiens (P16442)
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