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Literature summary for 3.2.1.96 extracted from

  • Fan, S.; Huang, W.; Wang, L.
    Remarkable transglycosylation activity of glycosynthase mutants of Endo-D, an endo-beta-N-acetylglucosaminidase from Streptococcus pneumoniae (2012), J. Biol. Chem., 287, 11272-11281.
    View publication on PubMedView publication on EuropePMC
Search for more literature for 3.2.1.96

Cloned(Commentary)

Cloned (Comment) Organism
expressed in Escherichia coli BL21(DE3) cells Streptococcus pneumoniae

Protein Variants

Protein Variants Comment Organism
H371W the mutant shows increased hydrolytic activity compared to the wild type Streptococcus pneumoniae
N322A the mutant demonstrates no product hydrolysis activity Streptococcus pneumoniae
N322A the mutant demonstrates remarkable transglycosylation activity with no product hydrolysis activity, having much higher catalytic efficiency for glycosylating the nonfucosylated GlcNAc acceptor Streptococcus pneumoniae
N322Q the mutant demonstrates remarkable transglycosylation activity with only marginal hydrolysis activity, having much higher catalytic efficiency for glycosylating the nonfucosylated GlcNAc acceptor Streptococcus pneumoniae
Y360F the mutant shows slightly reduced hydrolytic activity compared to the wild type Streptococcus pneumoniae

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.5
-
 Man3GlcNAc-oxazoline mutant enzyme N322Q, with Fmoc-Asn(Fucalpha1,6GlcNAc)-OH as acceptor, at pH 7.5 and 30°C Streptococcus pneumoniae
0.67
-
 Man3GlcNAc-oxazoline mutant enzyme N322A, with Fmoc-Asn(Fucalpha1,6GlcNAc)-OH as acceptor, at pH 7.5 and 30°C Streptococcus pneumoniae
0.73
-
 Fmoc-Asn(GlcNAc)-OH mutant enzyme N322Q, at pH 7.5 and 30°C Streptococcus pneumoniae
5.22
-
 Fmoc-Asn(GlcNAc)-OH mutant enzyme N322A, at pH 7.5 and 30°C Streptococcus pneumoniae
8.51
-
 Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH mutant enzyme N322A, at pH 7.5 and 30°C Streptococcus pneumoniae
18.51
-
 Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH mutant enzyme N322Q, at pH 7.5 and 30°C Streptococcus pneumoniae

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
100000
-
 x * 100000, SDS-PAGE Streptococcus pneumoniae
102000
-
 x * 102000, calculated from amino acid sequence Streptococcus pneumoniae

Organism

Organism UniProt Comment Textmining
Streptococcus pneumoniae Q93HW0
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni2+-immobilized HisTrap column chromatography Streptococcus pneumoniae

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
Man3GlcNAc-oxazoline + Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH
-
 Streptococcus pneumoniae ?
-
 ?
Man3GlcNAc-oxazoline + Fmoc-Asn(GlcNAc)-OH
-
 Streptococcus pneumoniae ?
-
 ?
additional information the enzyme also possesses transglycosylation activity with sugar oxazoline as the donor substrate, but the transglycosylation yield is low due to enzymatic hydrolysis of the donor substrate and the product. The enzyme favors nonfucosylated GlcNAc acceptor for transglycosylation Streptococcus pneumoniae ?
-
 ?
N-glycosylated immunglobuline G + H2O the enzyme prefers core-fucosylated N-glycan for hydrolysis Streptococcus pneumoniae ?
-
 ?

Subunits

Subunits Comment Organism
? x * 100000, SDS-PAGE Streptococcus pneumoniae
? x * 102000, calculated from amino acid sequence Streptococcus pneumoniae

Synonyms

Synonyms Comment Organism
endo-beta-N-acetylglucosaminidase
-
 Streptococcus pneumoniae
Endo-D
-
 Streptococcus pneumoniae

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.21
-
 Man3GlcNAc-oxazoline mutant enzyme N322A, with Fmoc-Asn(Fucalpha1,6GlcNAc)-OH as acceptor, at pH 7.5 and 30°C Streptococcus pneumoniae
0.24
-
 Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH mutant enzyme N322A, at pH 7.5 and 30°C Streptococcus pneumoniae
0.5
-
 Fmoc-Asn(GlcNAc)-OH mutant enzyme N322A, at pH 7.5 and 30°C Streptococcus pneumoniae
6.37
-
 Fmoc-Asn(GlcNAc)-OH mutant enzyme N322Q, at pH 7.5 and 30°C Streptococcus pneumoniae
10.17
-
 Man3GlcNAc-oxazoline mutant enzyme N322Q, with Fmoc-Asn(Fucalpha1,6GlcNAc)-OH as acceptor, at pH 7.5 and 30°C Streptococcus pneumoniae
14.32
-
 Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH mutant enzyme N322Q, at pH 7.5 and 30°C Streptococcus pneumoniae

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.0286
-
 Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH mutant enzyme N322A, at pH 7.5 and 30°C Streptococcus pneumoniae
0.096
-
 Fmoc-Asn(GlcNAc)-OH mutant enzyme N322A, at pH 7.5 and 30°C Streptococcus pneumoniae
0.31
-
 Man3GlcNAc-oxazoline mutant enzyme N322A, with Fmoc-Asn(Fucalpha1,6GlcNAc)-OH as acceptor, at pH 7.5 and 30°C Streptococcus pneumoniae
0.77
-
 Fmoc-Asn(Fucalpha(1->6)GlcNAc)-OH mutant enzyme N322Q, at pH 7.5 and 30°C Streptococcus pneumoniae
8.7
-
 Fmoc-Asn(GlcNAc)-OH mutant enzyme N322Q, at pH 7.5 and 30°C Streptococcus pneumoniae
20.4
-
 Man3GlcNAc-oxazoline mutant enzyme N322Q, with Fmoc-Asn(Fucalpha1,6GlcNAc)-OH as acceptor, at pH 7.5 and 30°C Streptococcus pneumoniae