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Literature summary for 3.2.1.97 extracted from

  • Willis, L.M.; Zhang, R.; Reid, A.; Withers, S.G.; Wakarchuk, W.W.
    Mechanistic investigation of the endo-alpha-N-acetylgalactosaminidase from Streptococcus pneumoniae R6 (2009), Biochemistry, 48, 10334-10341.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
expression of wild-type and mutant enzymes in Escherichia coli strain AD202 Streptococcus pneumoniae R6

Protein Variants

Protein Variants Comment Organism
D658A site-directed mutagenesis, the substrate specificity and activity is altered compared to the wild-type enzyme Streptococcus pneumoniae R6
D764A site-directed mutagenesis, the substrate specificity and activity is altered compared to the wild-type enzyme Streptococcus pneumoniae R6
D764F site-directed mutagenesis, the substrate specificity and activity is altered compared to the wild-type enzyme Streptococcus pneumoniae R6
E796A site-directed mutagenesis, the substrate specificity and activity is altered compared to the wild-type enzyme Streptococcus pneumoniae R6
E796Q site-directed mutagenesis, the substrate specificity and activity is altered compared to the wild-type enzyme Streptococcus pneumoniae R6
additional information construction of a DELTA1-39 deletion and DELTA1-39,DELTA1568-1767 deletion mutants Streptococcus pneumoniae R6

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information kinetic analysis, overview Streptococcus pneumoniae R6
0.0012
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796Q Streptococcus pneumoniae R6
0.0013
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
0.021
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
0.033
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant D764A Streptococcus pneumoniae R6
0.034
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
0.048
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39,DELTA1568-1767 deletion mutant Streptococcus pneumoniae R6
0.062
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
0.212
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796Q Streptococcus pneumoniae R6
2.1
-
2,4-dinitrophenyl-GalNAc-alpha-R pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
2.4
-
2,4-dinitrophenyl-GalNAc-alpha-R pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6

Metals/Ions

Metals/Ions Comment Organism Structure
NaCl effects on kinetics, overview Streptococcus pneumoniae R6
NaN3 effects on kinetics, overview Streptococcus pneumoniae R6
Sodium formate effects on kinetics, overview Streptococcus pneumoniae R6

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
additional information Streptococcus pneumoniae R6 SpGH101 specifically removes an O-linked disaccharide Gal-beta-1,3-GalNAc-alpha from glycoproteins ?
-
?

Organism

Organism UniProt Comment Textmining
Streptococcus pneumoniae R6 Q8DR60
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant wild-type and mutant enzymes from Escherichia coli strain AD202 by anion exchange chromatography, ultrafiltration, gel filtration, and hydrophobic interaction chromatography Streptococcus pneumoniae R6

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc + H2O
-
Streptococcus pneumoniae R6 2,4-dinitrophenol + beta-D-Gal-(1->3)-alpha-D-GalNAc
-
?
2,4-dinitrophenyl-GalNAc-alpha-R + H2O
-
Streptococcus pneumoniae R6 2,4-dinitrophenol + GalNAc-alpha-R
-
?
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc + H2O
-
Streptococcus pneumoniae R6 4-nitrophenol + beta-D-Gal-(1->3)-alpha-D-GalNAc
-
?
additional information SpGH101 specifically removes an O-linked disaccharide Gal-beta-1,3-GalNAc-alpha from glycoproteins Streptococcus pneumoniae R6 ?
-
?
additional information the enzyme contains several putative carbohydrate binding modules, structure-function relationship, D764 and E796 are the nucleophile and general acid-base residues, respectively, overview Streptococcus pneumoniae R6 ?
-
?

Subunits

Subunits Comment Organism
More analysis of the three-dimensional structure of the recombinant protein, the enzyme contains several putative carbohydrate binding modules, structure-function relationship, overview Streptococcus pneumoniae R6

Synonyms

Synonyms Comment Organism
endo-alpha-N-acetylgalactosaminidase
-
Streptococcus pneumoniae R6
More the enzyme belongs to the glycoside hydrolase family 101, GH101 Streptococcus pneumoniae R6
SpGH101
-
Streptococcus pneumoniae R6

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Streptococcus pneumoniae R6

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.76
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant D764A Streptococcus pneumoniae R6
1
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
1.6
-
2,4-dinitrophenyl-GalNAc-alpha-R pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
2.1
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796Q Streptococcus pneumoniae R6
5
-
2,4-dinitrophenyl-GalNAc-alpha-R pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
14
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796Q Streptococcus pneumoniae R6
18
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
541
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
579
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
748
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39,DELTA1568-1767 deletion mutant Streptococcus pneumoniae R6

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
additional information
-
bell-shaped dependence of kcat/Km upon pH Streptococcus pneumoniae R6
6.5
-
assay at Streptococcus pneumoniae R6

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.0006
-
2,4-dinitrophenyl-GalNAc-alpha-R pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
0.002
-
2,4-dinitrophenyl-GalNAc-alpha-R pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
0.0099
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796Q Streptococcus pneumoniae R6
0.023
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant D764A Streptococcus pneumoniae R6
0.048
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
0.987
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant D658A Streptococcus pneumoniae R6
9.3
-
4-nitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6
11.7
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796Q Streptococcus pneumoniae R6
13.8
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, mutant E796A Streptococcus pneumoniae R6
15.6
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39,DELTA1568-1767 deletion mutant Streptococcus pneumoniae R6
15.9
-
2,4-dinitrophenyl beta-D-Gal-(1->3)-alpha-D-GalNAc pH 6.5, 37°C, DELTA1-39 deletion mutant Streptococcus pneumoniae R6