Information on EC 4.2.2.11 - guluronate-specific alginate lyase

Word Map on EC 4.2.2.11
Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
Specify your search results
Select one or more organisms in this record:
Show additional data
Do not include text mining results
Include (text mining) results (more...)
Include results (AMENDA + additional results, but less precise; more...)


The expected taxonomic range for this enzyme is: Bacteria, Eukaryota

EC NUMBER
COMMENTARY hide
4.2.2.11
-
RECOMMENDED NAME
GeneOntology No.
guluronate-specific alginate lyase
REACTION
REACTION DIAGRAM
COMMENTARY hide
ORGANISM
UNIPROT
LITERATURE
Eliminative cleavage of alginate to give oligosaccharides with 4-deoxy-alpha-L-erythro-hex-4-enuronosyl groups at their non-reducing ends and alpha-L-guluronate at their reducing end
show the reaction diagram
REACTION TYPE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
C-O bond cleavage
-
-
elimination
-
-
elimination of an alcohol from a polysaccharide
-
-
-
-
PATHWAY
BRENDA Link
KEGG Link
MetaCyc Link
alginate degradation
-
-
Fructose and mannose metabolism
-
-
SYSTEMATIC NAME
IUBMB Comments
alginate alpha-L-guluronate-uronate lyase
The enzyme catalyses the degradation of alginate by a beta-elimination reaction. It cleaves the (1->4) bond between alpha-L-guluronate and either alpha-L-guluronate or beta-D-mannuronate, generating oligosaccharides with 4-deoxy-alpha-L-erythro-hex-4-enuronosyl groups at their non-reducing ends and alpha-L-guluronate at the reducing end. Depending on the composition of the substrate, the enzyme produces oligosaccharides ranging from two to six residues, with preference for shorter products. cf. EC 4.2.2.3, mannuronate-specific alginate lyase.
CAS REGISTRY NUMBER
COMMENTARY hide
64177-88-4
-
ORGANISM
COMMENTARY hide
LITERATURE
UNIPROT
SEQUENCE DB
SOURCE
-
-
-
Manually annotated by BRENDA team
polysaccharide lyase family 15
UniProt
Manually annotated by BRENDA team
strain No. 272
-
-
Manually annotated by BRENDA team
strain No. 272
-
-
Manually annotated by BRENDA team
strain H-4
SwissProt
Manually annotated by BRENDA team
isolated from brown seaweed
-
-
Manually annotated by BRENDA team
Bacillus sp. ATB-1015
ATB-1015 strain
-
-
Manually annotated by BRENDA team
ALY-1 strain
-
-
Manually annotated by BRENDA team
type 25, intracellular and extracellular enzymes
-
-
Manually annotated by BRENDA team
strain M-1,intracellular and extracellular enzyme
-
-
Manually annotated by BRENDA team
; gene alg2A
UniProt
Manually annotated by BRENDA team
isolated from rotten seaweed
AB489222
GenBank
Manually annotated by BRENDA team
bifunctional enzyme, activities against poly(beta-D-mannuronate), EC 4.2.2.3, and poly(alpha-L-guluronate), EC 4.2.2.11
-
-
Manually annotated by BRENDA team
strain H-4
SwissProt
Manually annotated by BRENDA team
bifunctional enzyme, activities against poly(beta-D-mannuronate), EC 4.2.2.3, and poly(alpha-L-guluronate), EC 4.2.2.11
-
-
Manually annotated by BRENDA team
strain F6
-
-
Manually annotated by BRENDA team
gene algMytC
UniProt
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
isolated from gut microflora of abalone
UniProt
Manually annotated by BRENDA team
AL-128
-
-
Manually annotated by BRENDA team
AL-128
-
-
Manually annotated by BRENDA team
-
-
-
Manually annotated by BRENDA team
GENERAL INFORMATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
evolution
physiological function
additional information
SUBSTRATE
PRODUCT                       
REACTION DIAGRAM
ORGANISM
UNIPROT
COMMENTARY
(Substrate) hide
LITERATURE
(Substrate)
COMMENTARY
(Product) hide
LITERATURE
(Product)
Reversibility
r=reversible
ir=irreversible
?=not specified
acetylated alginate
?
show the reaction diagram
-
-
-
-
?
acetylated poly-(beta-(1->4)-D-mannuronan)
?
show the reaction diagram
-
-
-
-
?
alginate
?
show the reaction diagram
alpha-L-guluronosyl linkage in alginate
?
show the reaction diagram
poly(alpha-(1->4)-L-guluronic acid)
?
show the reaction diagram
poly(alpha-L-guluronate)
?
show the reaction diagram
poly(beta-(1->4)-D-mannuronic acid)
?
show the reaction diagram
preferred over poly(alpha-(1->4)-L-guluronic acid)
-
-
?
poly(beta-(1->4)-D-mannuronic acid/alpha-(1->4)-L-guluronic acid)
?
show the reaction diagram
alternating structure of alpha-L-guluronic acid and beta-D-mannuronic acid. 120% of the activity with alginate
-
-
?
poly(beta-D-mannuronic acid/alpha-(1->4)-L-guluronic acid)
?
show the reaction diagram
poly-(alpha-L-guluronate)
?
show the reaction diagram
-
-
-
?
poly-(beta-(1->4)-D-mannuronan)
?
show the reaction diagram
poly-alpha-L-guluronic acid
4-deoxy-erythro-hex-4-ene pyranosyluronate
show the reaction diagram
-
degradation of alginate
-
-
?
poly-alpha-L-guluronic acid
?
show the reaction diagram
poly-alpha-L-guluronic acid
unsaturated di-, tri- and tetrasaccharides
show the reaction diagram
poly-beta1,4-D-mannuronan
?
show the reaction diagram
saturated deca((1-4)-alpha-L-guluronan)
?
show the reaction diagram
saturated hepta((1-4)-alpha-L-guluronan)
?
show the reaction diagram
saturated hexa((1-4)-alpha-L-guluronan)
unsaturated tetramer and a saturated dimer
show the reaction diagram
saturated penta((1-4)-alpha-L-guluronan)
?
show the reaction diagram
saturated tetra((1-4)-alpha-L-guluronan)
?
show the reaction diagram
sodium alginate
?
show the reaction diagram
unsaturated hepta((1-4)-alpha-L-guluronan)
?
show the reaction diagram
unsaturated hexa((1-4)-alpha-L-guluronan)
unsaturated triguluronic acid + unsaturated tetraguluronic acid
show the reaction diagram
unsaturated pentaguluronan
?
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
alginate
?
show the reaction diagram
poly-alpha-L-guluronic acid
4-deoxy-erythro-hex-4-ene pyranosyluronate
show the reaction diagram
-
degradation of alginate
-
-
?
poly-alpha-L-guluronic acid
unsaturated di-, tri- and tetrasaccharides
show the reaction diagram
sodium alginate
?
show the reaction diagram
additional information
?
-
METALS and IONS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
(NH4)2SO4
-
relative activity: 409%
Ag+
1 mM, 131% of initial activity
Ba2+
-
1 mM, 143% of initial activity
CH3COO- NH4+
-
relative activity: 298%
CH3COONa
-
relative activity: 233%
Co2+
-
1 mM, 123% of initial activity
CoCl2
-
relative activity: 102%
Cs+
0.2 M, 2500% of initial activity
Cu2+
-
2 mM, 66% inhibition
CuSO4
-
relative activity: 93%
EDTA
-
relative activity: 14%
EGTA
-
relative activity: 14%
HgCl2
-
relative activity: 29%
K2HPO4
-
relative activity: 439%
KCl
-
relative activity: 521%
Li+
0.2 M, 1880% of initial activity
NaBr
-
relative activity: 712%
NaF
-
relative activity: 419%
NH4+
0.2 M, 2270% of initial activity
NH4Cl
-
relative activity: 549%
NiCl2
-
relative activity: 106%
PdCl2
-
relative activity: 5%
Rb+
0.2 M, 2500% of initial activity
Sr2+
-
1 mM, 124% of initial activity
SrCl2
-
relative activity: 54%
ZnCl2
-
relative activity: 111%
additional information
INHIBITORS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
Al3+
-
1 mM, 47% residual activity
AlCl3
-
53% inhibition at 1 mM
Ba(OH)2
-
slightly inhibitory, percentage of inhibition is dependent on substrate
Ba2+
-
50 mM, isoform A 89%, isoform B 44%, isoform C 42% of initial activity
BaCl2
-
28% inhibition at 1 mM
Cd2+
-
2 mM, 25% residual activity
CuCl2
ethylene glycol tetraacetic acid
-
2 mM, 23% residual activity
Fe3+
-
1 mM, 8% residual activity
FeCl2
-
1 mM, 29% remaining activity
FeCl3
-
92% inhibition at 1 mM
glutathione
-
2 mM, 23% residual activity
Guanidinium chloride
HgCl2
-
1 mM, 23% remaining activity
N-bromosuccinimide
-
-
Ni2+
-
2 mM, 65% residual activity
NiCl2
-
33% inhibition at 1 mM
NiSO4
-
slightly inhibitory, percentage of inhibition is dependent on substrate
p-chloromercuribenzoate
-
-
SDS
-
10 mM SDS causes complete loss of activity whereas the ordered beta-structure is created
Sr2+
-
2 mM, 70% residual activity
trinitrobenzene sulfonic acid
-
-
Triton X-100
slightly activating at 0.001%, slightly inhibitory at 0.01%
ZnCl2
additional information
-
ACTIVATING COMPOUND
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
alginates with low guluronic content
-
cultures in the presence of alginates with low guluronic content give reduced biomass but favor enzyme production
Triton X-100
slightly activating at 0.001%, slightly inhibitory at 0.01%
Tween 20
slightly activating at 0.001-0.01%
Tween 40
slightly activating at 0.001-0.01%
Tween 80
slightly activating at 0.001-0.01%
KM VALUE [mM]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
0.0026
acetylated alginate
-
pH 7.1, 25C
-
0.005
acetylated poly-(beta-(1->4)-D-mannuronan)
-
pH 7.1, 25C
-
0.011
alginate
-
pH 7.1, 25C
0.013
poly-(beta-(1->4)-D-mannuronan)
-
pH 7.1, 25C
0.077
saturated deca((1-4)-alpha-L-guluronan)
-
pH 7.0, 50 mM phosphate buffer
-
0.099 - 0.192
saturated hepta((1-4)-alpha-L-guluronan)
-
0.11 - 0.226
saturated hexa((1-4)-alpha-L-guluronan)
-
0.232 - 0.3
saturated penta((1-4)-alpha-L-guluronan)
-
4.25
saturated tetra((1-4)-alpha-L-guluronan)
-
pH 8.5, 30 C, 100 mM glycine in 0.1 N NaCl and 0.1 N NaOH
-
0.0101 - 6.18
sodium alginate
additional information
alginate
TURNOVER NUMBER [1/s]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
1.2
acetylated alginate
Pseudomonas aeruginosa
-
pH 7.1, 25C
-
1.5
acetylated poly-(beta-(1->4)-D-mannuronan)
Pseudomonas aeruginosa
-
pH 7.1, 25C
-
32 - 999
alginate
32
poly-(beta-(1->4)-D-mannuronan)
Pseudomonas aeruginosa
-
pH 7.1, 25C
0.0149 - 872.8
sodium alginate
kcat/KM VALUE [1/mMs-1]
SUBSTRATE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
IMAGE
500
acetylated alginate
Pseudomonas aeruginosa
-
pH 7.1, 25C
197766
300
acetylated poly-(beta-(1->4)-D-mannuronan)
Pseudomonas aeruginosa
-
pH 7.1, 25C
197765
3100
alginate
Pseudomonas aeruginosa
-
pH 7.1, 25C
2943
2500
poly-(beta-(1->4)-D-mannuronan)
Pseudomonas aeruginosa
-
pH 7.1, 25C
197764
0.58 - 2570
sodium alginate
120502
additional information
alginate
Flavobacterium sp.
G9CHX6
kcat/Km value 26 ml/mg/s, pH 8.5, 30C
2943
SPECIFIC ACTIVITY [µmol/min/mg]
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
1.14
wild-type, pH 7.5, 30C
1.54
-
pH 5.0, 40C
1.98
-
wild-type enzyme, pH 7.0, 50C, substrate alginate
41.4
-
purified recombinant His-tagged enzyme, pH 7.0, 50C, substrate alginate
46
-
purified recombinant His-tagged enzyme, substrate poly(alpha-L-guluronate), pH 7.0, 50C
67
-
pH 6.5, 35C
233
-
isoform C, substrate sodium alginate, pH 7.0, 30C
265.4
purified recombinant His-tagged enzyme, pH 8.5, 45C
330
-
isoform B, substrate poly(alpha-L-guluronate), pH 7.0, 30C
390
-
isoform A, substrate poly(alpha-L-guluronate), pH 7.0, 30C
467
-
isoform B, substrate sodium alginate, pH 7.0, 30C
653
-
isoform A, substrate sodium alginate, pH 7.0, 30C
765
-
substrate poly(alpha-L-guluronic acid), 22C, pH 7.5
1401
pH 7.5, 30C
2152
-
pH 7.0, 38C; purified enzyme, pH 7.0, 38C
2490
-
30C, pH 8.0
4803
-
pH 8.5, 50C
8409
AB489222
pH 7.0, 30C; purified native enzyme, pH 7.0, 50C
pH OPTIMUM
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
pH RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 8.5
no activity outside this range
5 - 10
more than 60% of maximum activity within
5 - 11
-
activity range, profile overview
5.5 - 9.5
AB489222
activity range, profile overview
6 - 7.5
-
intracellular enzyme
6
highest activity at pH 6.0, which sharply decreases when the pH is higher or lower than pH 6.0
7 - 8
-
about 70% activity retained at pH 9.0
7 - 10
90% of maximal activity at pH 8.5 and pH 10.0, 30% at pH 7.5, inactive below, profile overview
7.5 - 8
-
maximal activity, slightly dependent on buffer composition
additional information
TEMPERATURE RANGE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
15 - 45
30% of maximal activity at 15C and 45C
20 - 70
40% of maximal activity at 20C and 70C, profile overview
20
50% of maximum activity
20 - 70
25 - 40
-
more than 80% of maximum activity
25 - 60
AB489222
activity range, profile overview
25 - 40
-
isoforms A, B, more than 60% of maximum activity
25
-
isoform C, 54% of maximum activity
30.55
more than 70% of maximum activity within
50
30% of maximum activity
pI VALUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4
-
isoform A, isoelectric focusing
4.3 - 4.4
-
-
4.4
-
isoform B, isoelectric focusing
4.6
-
isoform C, isoelectric focusing
6.6
sequence calculation
7.3 - 7.4
8.5
recombinant enzyme without signal peptide, calculated
8.6
recombinant enzyme with signal peptide, calculated
SOURCE TISSUE
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
SOURCE
PDB
SCOP
CATH
ORGANISM
UNIPROT
Zobellia galactanivorans (strain DSM 12802 / CCUG 47099 / CIP 106680 / NCIMB 13871 / Dsij)
Zobellia galactanivorans (strain DSM 12802 / CCUG 47099 / CIP 106680 / NCIMB 13871 / Dsij)
MOLECULAR WEIGHT
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
25009
x * 25009, calculated
25270
-
MALDI-TOF mass spectrometry
27000
-
gel filtration
28400
-
amino acid analysis
29643
1 * 29643, calculated, 1 * 28000, SDS-PAGE
31600
-
1 * 31600, intracellular, SDS-PAGE
32700
-
x * 32700, recombinant His-tagged enzyme without, SDS-PAGE
33900
-
x * 33900, SDS-PAGE
37000
-
x * 37000, SDS-PAGE
37573
x * 37573, sequence calculation, x * 43000, recombinant His-tagged enzyme, SDS-PAGE
38300
monomeric recombinant His-tagged enzyme, gel filtration
39000
-
Sephacryl S-200 HR column chromatography
40000 - 42000
-
-
42000
x * 42000, recombinant His-tagged enzyme, SDS-PAGE
42400
x * 45500, recombinant enzyme with signal peptide, x * 42400, recombinant enzyme without signal peptide, calulated
45000
-
x * 45000 plus x * 50000, SDS-PAGE
45500
x * 45500, recombinant enzyme with signal peptide, x * 42400, recombinant enzyme without signal peptide, calulated
47800
x * 47800, recombinant His-tagged enzyme, SDS-PAGE
50000
-
x * 45000 plus x * 50000, SDS-PAGE
54426
x * 54426, calculated
60250
-
x * 60250, isoform A, x * 36000, isoform B, x * 23000, isoform C, SDS-PAGE
69500
dimeric recombinant His-tagged enzyme, gel filtration
77500
x * 77500, SDS-PAGE of recombinant protein used to maltose binding protein
78000
x * 79900, including signal peptide, x * 78000, without signal peptide, calculated. x * 80000, SDS-PAGE
79100
x * 81600, about, sequence calculation, x * 79100, recombinant enzyme, SDS-PAGE
79900
x * 79900, including signal peptide, x * 78000, without signal peptide, calculated. x * 80000, SDS-PAGE
80000
x * 79900, including signal peptide, x * 78000, without signal peptide, calculated. x * 80000, SDS-PAGE
81600
x * 81600, about, sequence calculation, x * 79100, recombinant enzyme, SDS-PAGE
SUBUNITS
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
homodimer
monomer
monomer or dimer
additional information
POSTTRANSLATIONAL MODIFICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
proteolytic modification
Crystallization/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
crystal structures of native protein and its inactive mutant H531A in complex with alginate trisaccharide, at 2.10 and 2.99 A resolutions with final R-factors of 18.3 and 19.9%, respectively. The enzyme is comprised of an alpha/alpha-barrel plus anti-parallel beta-sheet as a basic scaffold. His311 and Tyr365 are the catalytic base and acid, respectively. A short alpha-helix in the central alpha/alpha-barrel domain and a conformational change at the interface between the central and C-terminal domains are essential for the exolytic mode of action
hanging-drop vapour-diffusion, structure at 1.2 A resolution
-
20C, drop solution comprising 1.4 M NaCl, 0.1 M potassium sodium phosphate and 0.1 M 2-morpholinoethanesulfonate-sodium hydroxide pH 6.5, vapor-diffusion method, monoclinic cristals
-
homology modeling of structure. Residues Asn198, His199, Arg246, and Tyr253 are conserved for the catalytic active site
purified recombinant soluble His-tagged wild-type and selenomethionine-labeled enzyme, and two mutant enzymes H202L and Y258A, and Y258A DELTAMMG variant, free or in complex with an alginate trisaccharide, hanging drop vapour diffusion method, mixing of 15 mg/ml protein in in 20 mM HEPES, pH 7.5, 100 mM KCl, with 0.1 M Tris, pH 8.0, 5% 2-methyl-2,4-pentanediol, 10% PEG 6000, 3 days, 16C, X-ray diffraction structure determination and analysis at 1.85 A, 1.7 A, 2.45 A, and 1.9 A resolution, respectively
hanging-drop vapour-diffusion, A1-II structure at 2.2 A resolution, A1-II' structure at 1.0 A resolution
-
purified recombinant catalytic domain of AlyA1PL7, hanging drop vapor diffusion method, 0.002 ml of 11.3 mg/ml protein solution with 0.1 mg/ml oligoguluronate, are mixed with 0.001 ml of reservoir solution containing 0.2 M KSCN and 28% PEG-MME 2000, and equilibration against 0.5 ml reservoir solution, 21C, screening and method optimization, X-ray diffraction structure determination and analysis at 1.43 A resolution; purified recombinant full-length dimeric AlyA5, hanging drop vapor diffusion method, 0.002 ml of 7.7 mg/ml protein solution with 0.1 mg/ml oligoglucuronate, are mixed with 0.001 ml of reservoir solution containing PEG 3350 and 0.2 M sodium/potassium tartrate, and equilibration against 02 ml reservoir solution, 21C, screening and method optimization, X-ray diffraction structure determination and analysis at 1.75 A resolution
pH STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
4 - 10
4
24 h, 20% residual activity
730135
4 - 10
purified recombinant enzyme, 25C, 24 h, over 70% activity remaining
730184
5 - 11
-
stable
648533
5
-
relatively stable in alkaline pH and unstable in a range outside acidic pH 5.0
34064
5 - 7
-
isoform C
713799
5 - 9
-
isoforms A and B
713799
5.5 - 8.5
AB489222
12 h, more than 60% residual activity; purified enzyme, 12 h, over 60% activity within this pH range remains pH 5.5 to 8.5, mostly stable at pH 7.5, half inactivation at pH 9.5
729531
6 - 9
-
A1m is quite stable in incubation at 30C for 1 h between pH 6 and 9
693771
6 - 7.5
-
sharp drop of activity below pH 6.0 and above pH 7.5
648523
6 - 10
-
stable
648525
6 - 9
-
; purified native enzyme, 40C, stable within this pH range
729470
6.5 - 8.5
-
stable
648534
8
-
most stable at pH 8.0
716092
8 - 9
the purified recombinant His-tagged enzyme exhibits stable and high activity at pH between 8.0 and 9.0
729058
8.5 - 10
purified recombinant enzyme, 25C, 24 h, over 90% activity remaining
730184
TEMPERATURE STABILITY
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
0
-
intracellular enzyme shows a weak residual activity of 9%
25 - 35
purified recombinant enzyme, pH 9.0, completely stable at 25C for 24 h and at 35C for 1 h, but loss of 60% activity after 24 h at 35C
30 - 50
-
enzyme is rapidly inactivated upon incubation at 30-50C for 15 min
30
-
stable at temperatures up to 30 C
35
-
30 min, isoform B 59% residual activity, isoform C 55% residual activity
45
purified recombinant His-tagged enzyme, pH 8.5, 1 h, stable up to; stable below
55
-
intracellular enzyme shows a weak residual activity of 9%
63
-
half-life 20 min
90
-
10 min, 40% residual activity; purified native enzyme, 10 min, pH 7.0, 40% activity remains
100
-
incubation for 15 min causes complete loss of activity, activity diminishes with increasing incubation temperatures below 100 C, addition of hen egg-white lysozyme decreased its thermal activity dramatically
GENERAL STABILITY
ORGANISM
UNIPROT
LITERATURE
activity of AlgMytC is stable in the presence of various detergents
more labile to heat treatment in Tris-HCl buffer, pH 7.0 than in phosphate buffer with similar pH
-
stable to freeze-drying, retains activity for several months in this form
-
unstable to heating, shows low activity when mantained in 50 mM Tris-HCl buffer
-
STORAGE STABILITY
ORGANISM
UNIPROT
LITERATURE
-80C, presence of 10% glycerol, stable for several months
-
4C, stable for 2 weeks
-
cell-free extract at -20 C
-
Purification/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
ammonium sulfate precipitation, cation exchange chromatography and gel filtration
-
ammonium sulfate precipitation, gel filtration and anion-exchange chromatography
-
anion exchange chromatography, and Sephadex G-100 chromatography
-
cation exchange chromatography, chromatofocusing and gel filtration
-
native enzyme 1.75fold by ammonium sulfate fractionation and anion exchange chromatography
AB489222
native extracellular enzyme 23.6fold from cell culture supernatant; purification from culture liquid
-
partial, ammonium sulfate precipitation and ion-exchange chromatography
-
purification leads to a mixture of three monomeric alginate lyases with molecular masses of 60.25, 36, and 23 kDa
-
recombinant Alg17C from Escherichia coli strain BL21(DE3)
recombinant His-tagged enzyme 5.74fold from Escherichia coli strain BL21(DE3) by nickel affinity chromatography
recombinant His-tagged enzyme by nickel affinity chromatography from Escherichia coli strain BL21(DE3)
recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration to homogeneity; recombinant His-tagged enzyme from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and gel filtration to homogeneity
recombinant His-tagged enzyme lacking the signal peptide from Escherichia coli strain BL21(DE3) by nickel affinity chromatography and ultrafiltration
-
recombinant protein
recombinant soluble and renatured His-tagged enzyme from Escherichia coli strain BL21(DE3)
recombinant soluble His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) Rosetta culture supernatant by nickel affinity chromatography and gel filtration
using a SuperQ Toyopearl column and Butyl Toyopearl column
-
Cloned/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
baculoviral expression system
-
DNA and amino acid sequence determination and analysis, expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
DNA and amino acid sequence determination and analysis, phylogenetic analysis
AB489222
expressed in Escherichia coli
-
expression in Escherichia coli
expression in Escherichia coli; gene alg2A, DNA and amino acid sequence determination and analysis, sequence comparisons and phylogenetic tree, expression of His-tagged enzyme in Escherichia coli strain BL21(DE3)
gene alg17C, expression in Escherichia coli strain BL21(DE3)
gene alg7D, overexpression of soluble His-tagged enzyme lacking the signal peptide in Escherichia coli strain BL21(DE3)
-
gene algMytC, DNA and amino acid sequence determination and analysis, sequence comparisons, recombinant expression of N-terminally His-tagged enzyme in Escherichia coli strain BL21(DE3) partly in inclusion bodies
phylogenetic analysis, recombinant expression of His-tagged AlyA1PL7 in Escherichia coli strain BL21(DE3); phylogenetic analysis, recombinant expression of His-tagged AlyA5 in Escherichia coli strain BL21(DE3)
subcloning of His-tagged wild-type and mutant enzymes in Escherichia coli strain DH5-alpha, expression in Escherichia coli strain BL21(DE3) Rosetta, the enzymes are secreted from periplasmic space
ENGINEERING
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
H311A
0.3% of wild-type activity
R199A
4.3% of wild-type activity
W467A
0.45% of wild-type activity
Y365F
0.011% of wild-type activity
A78S
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 820 U/mg, against poly(alpha-L-guluronic acid) 938 U/mg. Ratio of activities 1.1
A78S/T89I
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 228 U/mg, against poly(alpha-L-guluronic acid) 34.3 U/mg. Ratio of activities 0.2
A78S/T89I/A217E
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 187 U/mg, against poly(alpha-L-guluronic acid) 29.8 U/mg. Ratio of activities 0.2
G26E
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 692 U/mg, against poly(alpha-L-guluronic acid) 787 U/mg. Ratio of activities 1.1
G26E/P39H
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 246 U/mg, against poly(alpha-L-guluronic acid) 19.5 U/mg. Ratio of activities 0.1. In the absence of Ca2+, no detectable activity against G-M linkages
G304V
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 828 U/mg, against poly(alpha-L-guluronic acid) 878 U/mg. Ratio of activities 1.1
I51M
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 731 U/mg, against poly(alpha-L-guluronic acid) 786 U/mg. Ratio of activities 1.1
I51M/T89I
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 227 U/mg, against poly(alpha-L-guluronic acid) 18.8 U/mg. Ratio of activities 0.1
I51M/T89I/G304V
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 153 U/mg, against poly(alpha-L-guluronic acid) 13.3 U/mg. Ratio of activities 0.1
P39H
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 356 U/mg, against poly(alpha-L-guluronic acid) 37 U/mg. Ratio of activities 0.1
P39T
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 216 U/mg, against poly(alpha-L-guluronic acid) 71 U/mg. Ratio of activities 0.3
S35R
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 443 U/mg, against poly(alpha-L-guluronic acid) 448 U/mg. Ratio of activities 1.0
S35R/P39T
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 32.6 U/mg, against poly(alpha-L-guluronic acid) 6.9 U/mg. Ratio of activities 0.2
S35R/P39T/A224V
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 43 U/mg, against poly(alpha-L-guluronic acid) 3.4 U/mg. Ratio of activities 0.1
S37I
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 53.8 U/mg, against poly(alpha-L-guluronic acid) 4.3 U/mg. Ratio of activities 0.1
S86L
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 31 U/mg, against poly(alpha-L-guluronic acid) 9.1 U/mg. Ratio of activities 0.3
T85A
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) 24.8 is U/mg, against poly(alpha-L-guluronic acid) 4.4 U/mg. Ratio of activities 0.32
T89I
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 218 U/mg, against poly(alpha-L-guluronic acid) 31 U/mg. Ratio of activities 0.1
V6I
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is858 U/mg, against poly(alpha-L-guluronic acid) 851 U/mg. Ratio of activities 1.0
V6I/T85A
-
activity against poly(beta-D-mannuronic acid/alpha-L-guluronic acid) is 23.6 U/mg, against poly(alpha-L-guluronic acid) 2.2 U/mg. Ratio of activities 0.1
H202L
site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme
H415A
site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
N201A
site-directed mutagenesis, inactive mutant
Q149A
site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
R260A
site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
R438A
site-directed mutagenesis, the mutant shows highly reduced activity compared to the wild-type enzyme
Y258A
site-directed mutagenesis, inactive mutant
Y450A
site-directed mutagenesis, inactive mutant
additional information
-
engineering of different lyases, each of which cleaves only one of the four possible linkages in alginates: G-G, G-M, M-G, and M-M. The substitutions conferring altered specificity to the mutant enzymes are located in conserved regions in the polysaccharide lyase family 7 alginate lyases. Structure-function analyses suggests that the improved G-G specificity might be caused by increased affinity for nonproductive binding of the alternating G-M structure
Renatured/COMMENTARY
ORGANISM
UNIPROT
LITERATURE
6M urea collapses the circular dicroic spectral bands of the native enzyme caused by aromatic amino acids side chains and causes dissapearance of the enzyme activity, removal of the denaturant by dialysis restored the native-like spectrum and the activity, but some parts of the enzyme conformation are defective in the renaturation conditions
-
solubilization of enzyme from inclusion bodies by 8 M urea in Tris-HCl, pH 8.0
APPLICATION
ORGANISM
UNIPROT
COMMENTARY hide
LITERATURE
agriculture
-
oligosaccharides produced by alginase may act as osmoprotective agents during the plant germination process. The relative root length of Brassica campestris L. increases with the addition of oligosaccharides with reduced degrees of polymerization. The oligosaccharides with lower degree of polymerization-values are effective in reducing the effect of salt stress on the activity of the superoxide dismutase and guaiacol peroxidase, and oligosaccharides with moderate degree of polymerization-values can reduce the increase in lipid peroxidation activities induced by salt stress
biofuel production
Alg17C can be used as the key enzyme to produce alginate monomers in the process of utilizing alginate for biofuels and chemicals production
food industry
industry
synthesis
additional information
-
use of recombinant abalone alginate lyase and beta-1,4-endoglucanase for protoplast isolation from Laminaria japonica. In Laminaria japonica blades pretreated with proteinase K and incubated in artificial seawater containing alginate lyase and beta-1,4-endoglucanase, the protoplast number is increased up to 5000000 protoplasts/g fresh weight. These cells are mostly derived from the epidermal layer rather than the cortical layer. Results suggest that at least three enzymes, alginate lyase, cellulase, and protease, are essential for effective protoplast isolation from Laminaria japonica