Information on EC 2.4.1.66 - procollagen glucosyltransferase

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The enzyme appears in viruses and cellular organisms

EC NUMBER
COMMENTARY hide
2.4.1.66
-
RECOMMENDED NAME
GeneOntology No.
procollagen glucosyltransferase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
UDP-glucose + (2S,5R)-5-O-(beta-D-galactosyl)-5-hydroxy-L-lysine-[procollagen] = UDP + (2S,5R)-5-O-[alpha-D-glucosyl-(1->2)-beta-D-galactosyl]-5-hydroxy-L-lysine-[procollagen]
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
hexosyl group transfer
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
Other types of O-glycan biosynthesis
-
-
SYSTEMATIC NAME
IUBMB Comments
UDP-glucose:(2S,5R)-5-O-(beta-D-galactosyl)-5-hydroxy-L-lysine-[procollagen] D-glucosyltransferase
Probably involved in the synthesis of carbohydrate units in complement (cf. EC 2.4.1.50 procollagen galactosyltransferase).
CAS REGISTRY NUMBER
COMMENTARY hide
9028-08-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
calf
-
-
Manually annotated by BRENDA team
tiger puffer
UniProt
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
malfunction
metabolism
physiological function
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
TDP-glucose + D-galactosylhydroxylysine
TDP + 1,2-D-glucosyl-D-galactosylhydroxylysine
show the reaction diagram
-
soluble enzyme, glucosylation at 60% of the rate of UDP-glucose
-
?
UDP-glucose + (2S,5R)-5-O-(beta-D-galactosyl)-5-hydroxy-L-lysine-[procollagen]
UDP + (2S,5R)-5-O-[alpha-D-glucosyl-(1->2)-beta-D-galactosyl]-5-hydroxy-L-lysine-[procollagen]
show the reaction diagram
-
-
-
-
?
UDP-glucose + 5-(D-galactosyloxy)-L-lysine-procollagen
UDP + 1,2-D-glucosyl-5-D-(galactosyloxy)-L-lysine-procollagen
show the reaction diagram
UDP-glucose + asialo alpha 1-glycoprotein
?
show the reaction diagram
UDP-glucose + calf skin gelatin
?
show the reaction diagram
-
-
-
-
?
UDP-glucose + D-galactosylhydroxylysine
UDP + 1,2-D-glucosyl-D-galactosylhydroxylysine
show the reaction diagram
UDP-glucose + D-galactosylsphingosine
UDP + 1,2-D-glucosyl-D-galactosylsphingosine
show the reaction diagram
UDP-glucose + fetuin minus sialic acid, galactose and N-acetylglucosamine
?
show the reaction diagram
UDP-glucose + galactosylhydroxylysyl residues in a calf skin gelatine
?
show the reaction diagram
-
-
-
-
?
UDP-glucose + ichthyocol
?
show the reaction diagram
-
fish collagen prepared from carp swim bladder, ichthyocol glycopeptides prepared by collagenase digestion are better substrates than native ichthyocol, better substrate than calf skin collagen
-
-
?
UDP-glucose + transferrin minus Fe3+ and sialic acid
?
show the reaction diagram
-
7% as effective as glucose-free bovine Achilles tendon collagen
-
-
?
UDP-glucose + type VII collagen
?
show the reaction diagram
-
-
-
-
?
additional information
?
-
NATURAL SUBSTRATES
NATURAL PRODUCTS
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
REVERSIBILITY
r=reversible
ir=irreversible
?=not specified
UDP-glucose + (2S,5R)-5-O-(beta-D-galactosyl)-5-hydroxy-L-lysine-[procollagen]
UDP + (2S,5R)-5-O-[alpha-D-glucosyl-(1->2)-beta-D-galactosyl]-5-hydroxy-L-lysine-[procollagen]
show the reaction diagram
-
-
-
-
?
UDP-glucose + 5-(D-galactosyloxy)-L-lysine-procollagen
UDP + 1,2-D-glucosyl-5-D-(galactosyloxy)-L-lysine-procollagen
show the reaction diagram
UDP-glucose + type VII collagen
?
show the reaction diagram
-
-
-
-
?
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
Ni2+
-
activates, soluble enzyme
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Acetylsalicylic acid
-
aspirin
Ca2+
-
5-10 mM: 10-15% inhibition of Mn2+-activated enzyme
cAMP
-
4.5 mM: 26% inhibition
Carminic acid
-
non-competitive, mechanism
Cd2+
-
inhibits Mn2+-activated enzyme
Co2+
-
inhibits at high concentrations, inhibits Mn2+-activated enzyme
Cu2+
-
inhibits Mn2+-activated enzyme
D-glucosamine
-
-
Fe2+
-
inhibits at high concentrations
Fe3+
-
inhibits Mn2+-activated enzyme, 0.03 mM: 27% inhibition of activity with 0.01 mM Mn2+ and 0.035 mM UDP-glucose
Mn2+
-
soluble enzyme, above 50 mM
Mn2+-UDP-complex
-
competitive inhibition to UDP-glucose, non-competitive to collagen
-
Ni2+
-
inhibits Mn2+-activated enzyme
p-chloromercuribenzoate
-
membrane-bound enzyme, 6 and 10 mM: 54% inhibition
p-hydroxymercuribenzoate
-
1 mM: complete inhibition
p-mercuribenzoate
-
0.0007 mM: 50% inhibition, substrates plus Mn2+ or Mn2+ alone partially protect
Sucrose
-
membrane-bound enzyme, 0.42 M: 88% inhibition, also inhibits soluble enzyme
UTP
-
membrane-bound enzyme, 6 mM: complete inhibition
Zn2+
-
kinetics
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
bovine serum albumin
-
50% activation of pure enzyme
-
dithiothreitol
-
activation, optimal concentration: 1 mM
additional information
-
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0021
calf skin collagen
-
-
-
0.167 - 0.63
Collagen
0.002 - 3.8
D-galactosylhydroxylysine
2.5
D-galactosylsphingosine
-
-
0.029
TDP-glucose
-
soluble enzyme
0.005 - 0.15
UDP-glucose
additional information
additional information
-
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
additional information
additional information
Gallus gallus
-
0.7 mol of glucosylgalactosylhydroxylysine residues formed per s per mol of enzyme
-
Ki VALUE [mM]
INHIBITOR
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.004
Mn2+-UDP-complex
-
-
-
0.003
UDP
-
-
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0.0019
-
purified soluble enzyme
0.0033
-
enzyme from embryo cartilage
0.0073
-
purified enzyme from plasma
0.031 - 0.041
-
enzyme from whole embryo
0.046
-
-
0.065 - 0.082
-
-
0.1
-
purified enzyme from blood platelets
0.33
-
GGT activity of LH3
additional information
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
5.8
-
skin enzyme
6 - 6.5
-
2 pH-optima: 6-6.5 and 7.5-8, regardless of size of acceptor
6.5 - 7.5
-
soluble enzyme
6.5 - 8
-
-
7 - 8
-
membrane-bound enzyme
7.5 - 8
-
2 pH-optima: 6-6.5 and 7.5-8, regardless of size of acceptor
8.3
-
2 pH-optima: 8.3 and 9.9
9.9
-
2 pH-optima: 8.3 and 9.9
additional information
-
pI: 4.2 and 8.1
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 11.5
-
about half-maximal activity at pH 4.0 and 11.5
5.2 - 6.5
-
about half-maximal activity at pH 5.2 and 6.5
TEMPERATURE OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
6.1
calculated from amino acid sequence
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
-
distribution, not: erythrocytes
Manually annotated by BRENDA team
-
embryo bone
Manually annotated by BRENDA team
-
fetal lung WI-38 and IMR-90 diploid fibroblasts, in cell suspension culture
Manually annotated by BRENDA team
-
-
Manually annotated by BRENDA team
-
peritoneal
Manually annotated by BRENDA team
highest expression
Manually annotated by BRENDA team
-
fetal lung WI-38 and IMR-90 diploid fibroblasts, in cell suspension culture
Manually annotated by BRENDA team
additional information
LOCALIZATION
ORGANISM
UNIPROT
COMMENTARY hide
GeneOntology No.
LITERATURE
SOURCE
-
5-10% of the total activity is surface-bound, the rest is of cytoplasmic origin
Manually annotated by BRENDA team
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
50000 - 55000
-
gel filtration
52000 - 54000
-
gel filtration
70000
-
about
72000
-
gel filtration
80000
-
value about, Western blot
85000
-
SDS-PAGE
180000
-
LH3 with GGT activity, gel filtration
additional information
-
amino acid composition
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
-
2 * 80000-85000, recombinant LH3 with GGT/GT activities, SDS-PAGE
monomer
-
1 * 72000-78000, value depends on the gel composition, SDS-PAGE
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
glycoprotein
-
-
additional information
-
structural domain fragment A of LH3 with GGT activity contains 2 N-glycosylation sites
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
20
-
10% loss of activity within 18 h, soluble enzyme, after DEAE-ion exchange chromatography
58
-
denaturation at
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
bovine serum albumin, final concentration of 0.1%, enhances stability
-
chromatography on Biogel A-1.5M decreases stability
-
concentrated enzyme, above 0.2 mg/ml, loses most of the activity
-
extremely stable to freezing, -40°C, and thawing cycles, soluble enzyme
-
loses activity during purification mostly due to inactivation
-
stable in crude tissue extracts
stable to repeated thawing and freezing, human liver or serum enzyme
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-20°C, enzyme concentration 0.1 mg/ml, several months, 33% loss of activity
-
-40°C, membrane-preparation, 1-2 months, stable
-
-40°C, soluble enzyme after Biogel A-1.5M chromatography, 2-4 weeks, stable
-
4°C, 0.1 M Tris-buffer, pH 7.2, 0.2 M NaCl, 1 week, 10% loss of activity
-
4°C, dilute enzyme solution, 1 week, more than 50% loss of activity
-
4°C, enzyme concentration 0.1 mg/ml, 1 week, 33% loss of activity
-
frozen, concentrated purified enzyme solution, little loss of activity
-
room temperature, soluble enzyme after DEAE-cellulose column, 18 h, 10% loss of activity
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
100fold; skin, partial, solubilized with Triton X-100
-
14600fold; affinity chromatography; solubilized with 0.5 M NaCl, 2 M urea increases solubilization rate; solubilized with Nonidet P-40; solubilized with Triton X-100
-
20fold
-
3110fold; affinity chromatography
-
39800fold from embryo
-
39800fold from embryo; affinity chromatography
-
5000fold; affinity chromatography; partial
-
50fold; partial
-
5400fold from plasma, 4400fold from blood platelets
-
80fold: soluble enzyme
-
His-tagged recombinant LH3/GGT/GT and N-terminal 30kDa fragment A of LH3 with GGT activity expressed in High Five insect cells
-
His-tagged recombinant LH3/GGT/GT protein expressed in Escherichia coli; partial
-
LH3-V5/His fusion protein is purified using nickel-nitrilotriacetic acid-agarose resin column chromatography
-
Ni-NTA-agarose
-
over 2000fold from whole embryo, 160fold from embryo cartilage; partial; solubilized with Triton X-100
-
partial
partial, His-tagged recombinant LH/GGT/GT protein expressed in Escherichia coli
-
partial; solubilized with CHAPS, Brij-35; solubilized with Nonidet P-40; solubilized with Triton X-100
-
partial; solubilized with Nonidet P-40
-
partial; solubilized with Triton X-100
-
skin, partial, solubilized with Triton X-100
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
a single gene codes for LH, GGT and GT activities; LH3 cDNA with GGT activity is cloned and expressed in Sf9 insect cells and in Escherichia coli XL1-Blue
-
cDNA sequence of GGT/LH3; LH3 cDNA with GGT activity is cloned and expressed in Sf9 insect cells and in Escherichia coli XL1-Blue
-
expressed in MDCK, COS-7 and HT-1080 cells
-
expressed in MEF cells
-
full-length LH3 and the N-terminal 30 kDa fragment A with GGT activity is cloned, sequenced and expressed in High Five insect cells
-
LH cDNA with GGT activity is cloned and expressed in Escherichia coli BL21 (DE3) pLysS, a single gene codes for LH, GGT and GT activities
-
LH cDNA with GGT activity is expressed in Escherichia coli XL1-Blue, cDNA sequence of GGT/LH
-
LH3-V5/His fusion protein is expressed in HEK-293 cells
-
overexpression in HT-1080 cells, expression of fragments in Escherichia coli
-
Plod3 gene encoding LH3/GGT is cloned and sequenced, gene structure and regulation, localized on chromosome 5
EXPRESSION
ORGANISM
UNIPROT
LITERATURE
67% expression of LH3 in humans with epidermolysis bullosa simplex
-
decrease in lysyl hydroxylase 3 in LH3 knock-out mice
-
enzyme levels are reduced in patients with recessive dystrophic epidermolysis bullosa
-
exogenous type VII collagen do not alter enzyme expression in cultured recessive dystrophic epidermolysis bullosa keratinocytes
-
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
A361G
-
mutant with reduced GGT activity
A380V
-
mutant with reduced GGT activity
A453I
-
mutant with a remarkably reduced GGT activity
C132I
-
mutant with dramatically reduced GGT activity
D331V
-
little effect on GGT activity
E371T
-
mutant with reduced GGT activity
E547K
-
mutant with increased GGT activity
F131L
-
mutant with markedly reduced GGT activity
F372Y
-
little effect on GGT activity
H165S
-
mutant with reduced GGT activity
K482A
-
mutant with increased GGT activity
K674V
-
no effect on GGT activity
L124V
-
mutant with 40% inhibited GGT activity
L125V
-
mutant with 70% inhibited GGT activity
L192K
-
mutant with reduced GGT activity
L196I
-
mutant with markedly reduced GGT activity
L42K
-
mutant with increased GGT activity
L491M
-
mutant with little increased GGT activity
L562I
-
mutant with increased GGT activity
L591I
-
mutant with increased GGT activity
M406L
-
mutant with reduced GGT activity
M642L
-
mutant with increased GGT activity
M724A
-
little effect on GGT activity
P591N
-
no effect on GGT activity
Q613E
-
mutant with little increased GGT activity
S204C
-
mutant with reduced GGT activity
V622L
-
no effect on GGT activity
A464I
-
no effect on GGT activity
C144I
-
mutant with dramatically reduced GGT activity
D187A/D189A/D190A/D191A
-
the mutant shows unreduced lysyl hydroxylase 3 activity
D392A
-
little effect on GGT activity
D669A
-
the mutant shows no lysyl hydroxylase 3 activity
F143L
-
mutant with reduced GGT activity
L136V
-
mutant with increased GGT activity
L137V
-
mutant with increased GGT activity
L208I
-
mutant with markedly reduced GGT activity
M650L
-
no effect on GGT activity
N23S
-
lysyl hydroxylase 3 activity is reduced by nearly 50%
R729A
-
the mutant shows no lysyl hydroxylase 3 activity
additional information