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Literature summary for 3.2.1.55 extracted from

  • Maehara, T.; Fujimoto, Z.; Ichinose, H.; Michikawa, M.; Harazono, K.; Kaneko, S.
    Crystal structure and characterization of the glycoside hydrolase family 62 alpha-L-arabinofuranosidase from Streptomyces coelicolor (2014), J. Biol. Chem., 289, 7962-7972.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of wild-type and mutant enzymes in Streptomyces lividans strain 1326 and secretion to the culture medium Streptomyces coelicolor

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recomnbinant enzyme free or complexed with mercury derivative, L-arabinose, or xylooligosaccharides, sitting drop vapour diffusion method, mixing of 0.0012 ml of 8.3 mg/ml protein in 2 mM Tris-HCl buffer, pH 7.0, and 20 mM NaCl, with 0.001 ml of reservoir solution containing 20% w/v PEG 3350 and 0.2 M tripotassium citrate, pH 8.3, 20°C, X-ray diffraction structure determination and analysis at 1.4-2.1 A resolution, model building and molecular refinement Streptomyces coelicolor

Protein Variants

Protein Variants Comment Organism
D165E site-directed mutagenesis Streptomyces coelicolor
D165N site-directed mutagenesis Streptomyces coelicolor
D202N site-directed mutagenesis, inactive mutant Streptomyces coelicolor
E324Q site-directed mutagenesis Streptomyces coelicolor
E361Q site-directed mutagenesis, inactive mutant Streptomyces coelicolor
N425Q site-directed mutagenesis Streptomyces coelicolor
N462Q site-directed mutagenesis, the mutant shows 30% reduced activity compared to the wild-type enzyme Streptomyces coelicolor
W233A site-directed mutagenesis Streptomyces coelicolor
W233F site-directed mutagenesis Streptomyces coelicolor
W233Y site-directed mutagenesis Streptomyces coelicolor
W270A site-directed mutagenesis, the mutant shows 70% reduced activity compared to the wild-type enzyme Streptomyces coelicolor
W270F site-directed mutagenesis, the mutant shows 70% reduced activity compared to the wild-type enzyme Streptomyces coelicolor
W270Y site-directed mutagenesis, the mutant shows 50% reduced activity compared to the wild-type enzyme Streptomyces coelicolor
Y424A site-directed mutagenesis Streptomyces coelicolor
Y424F site-directed mutagenesis Streptomyces coelicolor
Y424W site-directed mutagenesis Streptomyces coelicolor
Y461A site-directed mutagenesis, the mutant shows 20% reduced activity compared to the wild-type enzyme Streptomyces coelicolor
Y461F site-directed mutagenesis, the mutant shows 80% reduced activity compared to the wild-type enzyme Streptomyces coelicolor
Y461W site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Streptomyces coelicolor

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information kinetics of mutant enzymes, overview Streptomyces coelicolor
1.9
-
4-nitrophenyl-alpha-L-arabinofuranoside ph 5.5, 37°C, recombinant wild-type enzyme Streptomyces coelicolor
7.3
-
wheat L-arabinoxylan ph 5.5, 37°C, recombinant wild-type enzyme Streptomyces coelicolor

Localization

Localization Comment Organism GeneOntology No. Textmining
extracellular
-
Streptomyces coelicolor
-
-

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
L-arabinoxylan + H2O Streptomyces coelicolor the enzyme efficiently removes the alpha-L-arabinosyl substituents from arabinoxylan ?
-
?
L-arabinoxylan + H2O Streptomyces coelicolor A3(2) the enzyme efficiently removes the alpha-L-arabinosyl substituents from arabinoxylan ?
-
?

Organism

Organism UniProt Comment Textmining
Streptomyces coelicolor O54161
-
-
Streptomyces coelicolor A3(2) O54161
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from Streptomyces lividans strain 1326 culture medium by affinity chromatography using lactosyl-Sepharose, dialysis and ultrafiltration Streptomyces coelicolor

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
4-nitrophenyl alpha-L-arabinofuranoside + H2O
-
Streptomyces coelicolor 4-nitrophenol + L-arabinofuranose
-
?
4-nitrophenyl alpha-L-arabinofuranoside + H2O
-
Streptomyces coelicolor A3(2) 4-nitrophenol + L-arabinofuranose
-
?
L-arabinoxylan + H2O the enzyme efficiently removes the alpha-L-arabinosyl substituents from arabinoxylan Streptomyces coelicolor ?
-
?
L-arabinoxylan + H2O the enzyme efficiently removes the alpha-L-arabinosyl substituents from arabinoxylan Streptomyces coelicolor A3(2) ?
-
?
additional information substrate specificity and substrate-binding mechanism, overview Streptomyces coelicolor ?
-
?
additional information substrate specificity and substrate-binding mechanism, overview Streptomyces coelicolor A3(2) ?
-
?
wheat L-arabinoxylan + H2O the enzyme efficiently removes the alpha-L-arabinosyl substituents from arabinoxylan Streptomyces coelicolor ?
-
?
wheat L-arabinoxylan + H2O the enzyme efficiently removes the alpha-L-arabinosyl substituents from arabinoxylan Streptomyces coelicolor A3(2) ?
-
?

Synonyms

Synonyms Comment Organism
exo-alpha-L-arabinofuranosidase UniProt Streptomyces coelicolor
ScAraf62A
-
Streptomyces coelicolor

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
30 45 assay at Streptomyces coelicolor

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.02
-
4-nitrophenyl-alpha-L-arabinofuranoside ph 5.5, 37°C, recombinant wild-type enzyme Streptomyces coelicolor
0.3
-
wheat L-arabinoxylan ph 5.5, 37°C, recombinant wild-type enzyme Streptomyces coelicolor

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
5.5 6 assay at Streptomyces coelicolor

General Information

General Information Comment Organism
evolution the enzyme belongs to the family 62 glycoside hydrolase, GH62. The crystal structure comprises a carbohydrate-binding module family 13 domain at its N-terminus and a catalytic domain at its C-terminus Streptomyces coelicolor
additional information the catalytic domain is a five-bladed beta-propeller consisting of five radially oriented anti-parallel beta-sheets, substrate binding at five subsites in the catalytic cleft and an L-arabinose-binding pocket at the bottom of the cleft. Asp202 and Glu361 are catalytic residues, and Trp270, Tyr461, and Asn462 are involved in the substrate-binding site for discriminating the substrate structures Streptomyces coelicolor

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.011
-
4-nitrophenyl-alpha-L-arabinofuranoside pH 5.5, 37°C, recombinant wild-type enzyme Streptomyces coelicolor
0.042
-
wheat L-arabinoxylan pH 5.5, 37°C, recombinant wild-type enzyme Streptomyces coelicolor