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Literature summary for 3.2.1.86 extracted from

  • Yu, W.L.; Jiang, Y.L.; Pikis, A.; Cheng, W.; Bai, X.H.; Ren, Y.M.; Thompson, J.; Zhou, C.Z.; Chen, Y.
    Structural insights into the substrate specificity of a 6-phospho-beta-glucosidase BglA-2 from Streptococcus pneumoniae TIGR4 (2013), J. Biol. Chem., 288, 14949-14958.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 3.2.1.86

Cloned(Commentary)

Cloned (Comment) Organism
gene SP_0578 or bglA-2, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Streptococcus pneumoniae

Crystallization (Commentary)

Crystallization (Comment) Organism
purified recombinant enzyme in apoform or complexed with thiocellobiose 6-phosphate, sitting drop vapour diffusion method, mixing of 0.001 ml of 10 mg/ml protein in 20 mM Tris-Cl, pH 8.0, 100 mM NaCl with 0.001 ml of reservoir solution containing 15% PEG 5000MME, and 0.1 M sodium citrate, pH 5.6, 16°C, X-ray diffraction structure determination and analysis at 2.0-2.4 A resolution, molecular replacement method Streptococcus pneumoniae

Protein Variants

Protein Variants Comment Organism
E171A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
E171Q site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
E364A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
E364Q site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
K430A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
M423A site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Streptococcus pneumoniae
S424A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
W338A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
Y126A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
Y126F site-directed mutagenesis, the mutant shows reduced activity compared to the wild-type enzyme Streptococcus pneumoniae
Y303A site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
Y303F site-directed mutagenesis, inactive mutant Streptococcus pneumoniae
Y432F site-directed mutagenesis, inactive mutant Streptococcus pneumoniae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.478
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged wild-type enzyme Streptococcus pneumoniae
0.512
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant M423A Streptococcus pneumoniae
0.598
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant Y126F Streptococcus pneumoniae
1.135
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged wild-type enzyme Streptococcus pneumoniae
1.32
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant M423A Streptococcus pneumoniae
1.515
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant Y126F Streptococcus pneumoniae

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
cellobiose 6-phosphate + H2O Streptococcus pneumoniae
-
 beta-D-glucose + glucose 6-phosphate
-
 ?

Organism

Organism UniProt Comment Textmining
Streptococcus pneumoniae A0A0H2UP35 gene SP_0578 or bglA-2
-

Purification (Commentary)

Purification (Comment) Organism
recombinant tagged wild-type and mutant enzymes from Escherichia coli strain BL21(DE3) by nickel affinity chromatography, gel filtration, and ultrafiltration Streptococcus pneumoniae

Reaction

Reaction Comment Organism Reaction ID
6-phospho-beta-D-glucosyl-(1->4)-D-glucose + H2O = D-glucose + D-glucose 6-phosphate active site structure and double displacement mechanism, overview Streptococcus pneumoniae
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
4-nitrophenyl beta-D-glucopyranoside 6-phosphate + H2O
-
 Streptococcus pneumoniae 4-nitrophenol + beta-D-glucose 6-phosphate
-
 ?
cellobiose 6-phosphate + H2O
-
 Streptococcus pneumoniae beta-D-glucose + glucose 6-phosphate
-
 ?
additional information no activity with beta-(1,4)-thiocellobiose 6-phosphate Streptococcus pneumoniae ?
-
 ?

Synonyms

Synonyms Comment Organism
BglA-2
-
 Streptococcus pneumoniae

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Streptococcus pneumoniae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
138
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant Y126F Streptococcus pneumoniae
145
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant M423A Streptococcus pneumoniae
146
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant Y126F Streptococcus pneumoniae
168
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant M423A Streptococcus pneumoniae
170
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged wild-type enzyme Streptococcus pneumoniae
195
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged wild-type enzyme Streptococcus pneumoniae

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.5
-
 assay at Streptococcus pneumoniae

General Information

General Information Comment Organism
evolution the enzyme belongs to the glycosyl hydrolase family 1, GH1. Members of the GH-1 family share a common catalytic mechanism and exhibit similar structural folds, including a (beta/alpha)8 TIM-barrel Streptococcus pneumoniae
additional information the overall structure of enzyme BglA-2 adopts a typical (beta/alpha)8 TIM-barrel, with the active site located at the center of the convex surface of the beta-barrel. Residues Tyr126, Tyr303, and Trp338, at subsite +1 of BglA-2 determine substrate specificity with respect to 1,4-linked 6-phospho-beta-glucosides. Residues Ser424, Lys430, and Tyr432 of BglA-2 play important roles in the hydrolytic selectivity toward phosphorylated rather than non-phosphorylated compounds, comparative structural analysis. Tryptophan versus a methionine/alanine residue at subsite -1 may contribute to the catalytic and substrate selectivity with respect to structurally similar 6-phospho-beta-galactosidases and 6-phospho-beta-glucosidases assigned to the GH-1 family Streptococcus pneumoniae

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
90
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant Y126F Streptococcus pneumoniae
110
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant M423A Streptococcus pneumoniae
150
-
 cellobiose 6-phosphate pH 7.5, 37°C, recombinant His-tagged wild-type enzyme Streptococcus pneumoniae
240
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant Y126F Streptococcus pneumoniae
330
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged mutant M423A Streptococcus pneumoniae
410
-
 4-nitrophenyl-beta-D-glucopyranoside 6-phosphate pH 7.5, 37°C, recombinant His-tagged wild-type enzyme Streptococcus pneumoniae