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Literature summary for 2.4.1.5 extracted from

  • Irague, R.; Tarquis, L.; Andre, I.; Moulis, C.; Morel, S.; Monsan, P.; Potocki-Veronese, G.; Remaud-Simeon, M.
    Combinatorial engineering of dextransucrase specificity (2013), PLoS ONE, 8, e77837.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
gene dsrS, sequence comparisons and phylogenetic analysis, recombinant expression of His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21 AI Leuconostoc mesenteroides

Protein Variants

Protein Variants Comment Organism
D306X random mutagenesis Leuconostoc mesenteroides
D460A/H463S/T464L site-directed mutagenesis Leuconostoc mesenteroides
D460M/H463Y/T464M/S512C site-directed mutagenesis Leuconostoc mesenteroides
D460X random mutagenesis Leuconostoc mesenteroides
F353T random mutagenesis Leuconostoc mesenteroides
F353W random mutagenesis Leuconostoc mesenteroides
F353X random mutagenesis Leuconostoc mesenteroides
H463R/T464D/S512T site-directed mutagenesis Leuconostoc mesenteroides
H463R/T464V/S512T site-directed mutagenesis Leuconostoc mesenteroides
H463X random mutagenesis Leuconostoc mesenteroides
additional information screening of diverse mutants of the eight conserved residues that are determined to be important for enzyme activity, overview. Construction of enzyme mutant DSR-S vardel DELTA4N Leuconostoc mesenteroides
N404X random mutagenesis Leuconostoc mesenteroides
S512C random mutagenesis Leuconostoc mesenteroides
S512X random mutagenesis Leuconostoc mesenteroides
T464X random mutagenesis Leuconostoc mesenteroides
W440X random mutagenesis Leuconostoc mesenteroides

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.5
-
sucrose pH 5.2, 25°C, recombinant mutant F353W Leuconostoc mesenteroides
3.9
-
sucrose pH 5.2, 25°C, recombinant mutant S512C Leuconostoc mesenteroides
5.3
-
sucrose pH 5.2, 25°C, recombinant mutant D460A/H463S/T464L Leuconostoc mesenteroides
6.9
-
sucrose pH 5.2, 25°C, recombinant mutant H463R/T464V/S512T Leuconostoc mesenteroides
15.5
-
sucrose pH 5.2, 25°C, recombinant mutant D460M/H463Y/T464M/S512C Leuconostoc mesenteroides
15.5
-
sucrose pH 5.2, 25°C, recombinant mutant H463R/T464D/S512T Leuconostoc mesenteroides
115.8
-
sucrose pH 5.2, 25°C, recombinant mutant F353T Leuconostoc mesenteroides

Metals/Ions

Metals/Ions Comment Organism Structure
Ca2+ required Leuconostoc mesenteroides

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
sucrose + [(1->6)-alpha-D-glucosyl]n Leuconostoc mesenteroides
-
D-fructose + [(1->6)-alpha-D-glucosyl]n+1
-
?
sucrose + [(1->6)-alpha-D-glucosyl]n Leuconostoc mesenteroides NRRL B-512F
-
D-fructose + [(1->6)-alpha-D-glucosyl]n+1
-
?

Organism

Organism UniProt Comment Textmining
Leuconostoc mesenteroides Q9L3Z9 gene dsrS
-
Leuconostoc mesenteroides NRRL B-512F Q9L3Z9 gene dsrS
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21 AI by nickel affinity chromatography Leuconostoc mesenteroides

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
sucrose + [(1->6)-alpha-D-glucosyl]n
-
Leuconostoc mesenteroides D-fructose + [(1->6)-alpha-D-glucosyl]n+1
-
?
sucrose + [(1->6)-alpha-D-glucosyl]n
-
Leuconostoc mesenteroides NRRL B-512F D-fructose + [(1->6)-alpha-D-glucosyl]n+1
-
?

Synonyms

Synonyms Comment Organism
Dsr S protein
-
Leuconostoc mesenteroides
DSR-S
-
Leuconostoc mesenteroides

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
25
-
assay at Leuconostoc mesenteroides

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
38.2
-
sucrose pH 5.2, 25°C, recombinant mutant D460M/H463Y/T464M/S512C Leuconostoc mesenteroides
43.9
-
sucrose pH 5.2, 25°C, recombinant mutant D460A/H463S/T464L Leuconostoc mesenteroides
86
-
sucrose pH 5.2, 25°C, recombinant mutant F353W Leuconostoc mesenteroides
172.1
-
sucrose pH 5.2, 25°C, recombinant mutant H463R/T464D/S512T Leuconostoc mesenteroides
207.1
-
sucrose pH 5.2, 25°C, recombinant mutant S512C Leuconostoc mesenteroides
338.8
-
sucrose pH 5.2, 25°C, recombinant mutant F353T Leuconostoc mesenteroides
610.2
-
sucrose pH 5.2, 25°C, recombinant mutant H463R/T464V/S512T Leuconostoc mesenteroides

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
5.2
-
assay at Leuconostoc mesenteroides

General Information

General Information Comment Organism
evolution the enzyme belongs to the glycoside-hydrolase family 70, sequence and structural analysis of GH70 enzymes, overview Leuconostoc mesenteroides
additional information three-dimensional enzyme molecular structure modeling, structure comparisons, overview. Role of peptide 460DYVHT464 in enzyme DSR-S linkage specificity, together with residue S512 it contributes to defining the +2 subsite topology, which may be critical for the enzyme regiospecificity. Molecular docking of sucrose in the active site of enzyme mutant DSR-S vardel DELTA4N Leuconostoc mesenteroides

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
2.5
-
sucrose pH 5.2, 25°C, recombinant mutant D460M/H463Y/T464M/S512C Leuconostoc mesenteroides
2.9
-
sucrose pH 5.2, 25°C, recombinant mutant F353T Leuconostoc mesenteroides
8.3
-
sucrose pH 5.2, 25°C, recombinant mutant D460A/H463S/T464L Leuconostoc mesenteroides
11.1
-
sucrose pH 5.2, 25°C, recombinant mutant H463R/T464D/S512T Leuconostoc mesenteroides
53.1
-
sucrose pH 5.2, 25°C, recombinant mutant S512C Leuconostoc mesenteroides
57.3
-
sucrose pH 5.2, 25°C, recombinant mutant F353W Leuconostoc mesenteroides
88.4
-
sucrose pH 5.2, 25°C, recombinant mutant H463R/T464V/S512T Leuconostoc mesenteroides