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Literature summary extracted from

  • Bobrov, K.; Borisova, A.; Eneyskaya, E.; Ivanen, D.; Shabalin, K.; Kulminskaya, A.; Rychkov, G.
    Improvement of the efficiency of transglycosylation catalyzed by alpha-galactosidase from Thermotoga maritima by protein engineering (2013), Biochemistry, 78, 1112-1123 .
    View publication on PubMed

Protein Variants

EC Number Protein Variants Comment Organism
3.2.1.22 F328A transglycosylation is markedly increased compared to wild-type enzyme. kcat/Km value is more than twofold lower than for the wild type enzyme. 16-fold higher yield of products with the (alpha1,2)-bond as compared to products produced by the wild type enzyme Thermotoga maritima
3.2.1.22 G385L transglycosylation is markedly increased compared to wild-type enzyme. Mutation G385L results in a twofold decrease in the specific activity of the enzyme during its purification. The kinetic parameters are not determined for the mutant G385L due to its instability Thermotoga maritima
3.2.1.22 L195C kcat/Km value is more than twofold lower than for the wild type enzyme Thermotoga maritima
3.2.1.22 P402D transglycosylation is markedly increased compared to wild-type enzyme. kcat/Km value is more than twofold lower than for the wild type enzyme. 4-fold higher yield of products with the (alpha1,2)-bond as compared to products produced by the wild type enzyme Thermotoga maritima
3.2.1.22 P402S no significant changes in the kinetic parameters of hydrolysis Thermotoga maritima
3.2.1.22 W85Y no significant changes in the kinetic parameters of hydrolysis Thermotoga maritima

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
3.2.1.22 0.1
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme 328A Thermotoga maritima
3.2.1.22 0.1
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme F194K Thermotoga maritima
3.2.1.22 0.11
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, wild-type enzyme Thermotoga maritima
3.2.1.22 0.11
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme G385L Thermotoga maritima
3.2.1.22 0.11
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme P402S Thermotoga maritima
3.2.1.22 0.32
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme L195C Thermotoga maritima
3.2.1.22 0.43
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme P402D Thermotoga maritima

Organism

EC Number Organism UniProt Comment Textmining
3.2.1.22 Thermotoga maritima G4FEF4
-
-
3.2.1.22 Thermotoga maritima DSM 3109 G4FEF4
-
-

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.2.1.22 4-nitrophenyl alpha-D-galactopyranoside + H2O hydrolysis is accompanied by transglycosylation resulting in production of a mixture of (alpha1,2)-, (alpha1,3)-, and (alpha1,6)-4-nitrophenyl-digalactosides Thermotoga maritima 4-nitrophenol + D-galactopyranose
-
 ?
3.2.1.22 4-nitrophenyl alpha-D-galactopyranoside + H2O hydrolysis is accompanied by transglycosylation resulting in production of a mixture of (alpha1,2)-, (alpha1,3)-, and (alpha1,6)-4-nitrophenyl-digalactosides Thermotoga maritima DSM 3109 4-nitrophenol + D-galactopyranose
-
 ?

Subunits

EC Number Subunits Comment Organism
3.2.1.22 ? x * 60000, wild type enzyme and its mutants W85Y, F194K, L195C, F328A, P402D, P402S, and G385L, SDS-PAGE Thermotoga maritima

Turnover Number [1/s]

EC Number Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
3.2.1.22 1.1
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme 328A Thermotoga maritima
3.2.1.22 8
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, wild-type enzyme Thermotoga maritima
3.2.1.22 8.15
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme G385L Thermotoga maritima
3.2.1.22 9.8
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme P402S Thermotoga maritima
3.2.1.22 10.73
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme P402D Thermotoga maritima
3.2.1.22 12.2
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme L195C Thermotoga maritima
3.2.1.22 20.4
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme F194K Thermotoga maritima

kcat/KM [mM/s]

EC Number kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
3.2.1.22 11.57
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme 328A Thermotoga maritima
3.2.1.22 24.95
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme P402D Thermotoga maritima
3.2.1.22 38.76
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme L195C Thermotoga maritima
3.2.1.22 72.07
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, wild-type enzyme Thermotoga maritima
3.2.1.22 74.09
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme G385L Thermotoga maritima
3.2.1.22 88.28
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme P402S Thermotoga maritima
3.2.1.22 198.06
-
 4-nitrophenyl alpha-D-galactopyranoside 37°C, pH 5.0, mutant enzyme F194K Thermotoga maritima