Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary for 4.2.1.122 extracted from

  • Francis, D.; Winn, M.; Latham, J.; Greaney, M.; Micklefield, J.
    An engineered tryptophan synthase opens new enzymatic pathways to beta-methyltryptophan and derivatives (2017), ChemBioChem, 18, 382-386 .
    View publication on PubMed
Search for more literature for 4.2.1.122

Cloned(Commentary)

Cloned (Comment) Organism
recombinant expression of C-terminally His-tagged wild-type and mutant enzymes in Escherichia coli strain BL21(DE3) Salmonella enterica subsp. enterica serovar Typhimurium

Protein Variants

Protein Variants Comment Organism
L166A site-directed mutagenesis the beta-subunit, the mutant shows altered substrate specificity compared to the wild-type enzyme Salmonella enterica subsp. enterica serovar Typhimurium
L166V site-directed mutagenesis of the beta-subunit, the mutant shows altered substrate specificity compared to the wild-type enzyme, single-step enzymatic synthesis of beta-methyl-Trp derivatives, overview. Although 2-, 4-, 6- or 7-substituted indoles are accepted by StTrpS beta-L66V, along with L-Thr, 5-subsituted indoles prove to be very poor substrates for the enzyme. Variant beta-L166V can better accommodate L-Thr as a substrate Salmonella enterica subsp. enterica serovar Typhimurium
additional information an engineered variant of tryptophan synthase from Salmonella enterica (StTrpS) can catalyse the efficient condensation of L-threonine and various indoles to generate bmTrp and derivatives in a single step. Although L-serine is the natural substrate for TrpS, targeted mutagenesis of the StTrpS active site provides a variant (bL166V) that can better accommodate L-Thr as a substrate. The condensation of L-Thr and indole proceeds with retention of configuration at both alpha- and beta-positions to give (2S,3S)-beta-methyl-Trp Salmonella enterica subsp. enterica serovar Typhimurium

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
1.4
-
 L-serine recombinant mutant L166V, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
1.6
-
 L-serine recombinant wild-type enzyme, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
52
-
 L-serine recombinant mutant L166A, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
210
-
 L-threonine recombinant wild-type enzyme, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
310
-
 L-threonine recombinant mutant L166V, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
340
-
 L-threonine recombinant mutant L166A, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
L-serine + indole Salmonella enterica subsp. enterica serovar Typhimurium L-serine is the preferred substrate L-tryptophan + H2O
-
 ?

Organism

Organism UniProt Comment Textmining
Salmonella enterica subsp. enterica serovar Typhimurium
-
-
-
Salmonella enterica subsp. enterica serovar Typhimurium ATCC 37845
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant C-terminally His-tagged wild-type and mutant enzymes from Escherichia coli strain BL21-A1(DE3) by nickel affinity chromatography, ultrafiltration, and desalting gel filtration Salmonella enterica subsp. enterica serovar Typhimurium

Reaction

Reaction Comment Organism Reaction ID
L-serine + indole = L-tryptophan + H2O reaction mechanism, overview Salmonella enterica subsp. enterica serovar Typhimurium
a

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
L-serine + indole L-serine is the preferred substrate Salmonella enterica subsp. enterica serovar Typhimurium L-tryptophan + H2O
-
 ?
L-threonine + 2-methylindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-2-methyl-beta-methyltryptophan + H2O
-
 ?
L-threonine + 4-fluoroindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-4-fluoro-beta-methyltryptophan + H2O
-
 ?
L-threonine + 4-methylindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-4-methyl-beta-methyltryptophan + H2O
-
 ?
L-threonine + 5-bromoindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-5-bromo-beta-methyltryptophan + H2O
-
 ?
L-threonine + 5-chloroindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-5-chloro-beta-methyltryptophan + H2O
-
 ?
L-threonine + 5-phenylindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-5-phenyl-beta-methyltryptophan + H2O
-
 ?
L-threonine + 6-methylindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-6-methyl-beta-methyltryptophan + H2O
-
 ?
L-threonine + 7-chloroindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-7-chloro-beta-methyltryptophan + H2O
-
 ?
L-threonine + 7-fluoroindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-7-fluoro-beta-methyltryptophan + H2O
-
 ?
L-threonine + 7-methoxyindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-7-methoxy-beta-methyltryptophan + H2O
-
 ?
L-threonine + 7-methylindole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-7-methyl-beta-methyltryptophan + H2O
-
 ?
L-threonine + indole
-
 Salmonella enterica subsp. enterica serovar Typhimurium (2S,3S)-beta-methyltryptophan + H2O
-
 ?
additional information activity and isolated yields of L-beta-methyl-Trp are low compared to the wild-type reaction with l-Ser, which gives over 90% of L-Trp under the same conditions. Preperative scale synthesis (2S,3S)-L-beta-methyl tryptophan and derivatives Salmonella enterica subsp. enterica serovar Typhimurium ?
-
 ?

Synonyms

Synonyms Comment Organism
StTrpS (beta2)
-
 Salmonella enterica subsp. enterica serovar Typhimurium

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
 assay at Salmonella enterica subsp. enterica serovar Typhimurium

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.00068
-
 L-threonine recombinant wild-type enzyme, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.0035
-
 L-threonine recombinant mutant L166A, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.007
-
 L-threonine recombinant mutant L166V, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.082
-
 L-serine recombinant mutant L166V, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.18
-
 L-serine recombinant mutant L166A, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.35
-
 L-serine recombinant wild-type enzyme, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
7.8
-
 assay at Salmonella enterica subsp. enterica serovar Typhimurium

General Information

General Information Comment Organism
additional information leucine residue Leu166 (betaL166) is in close proximity to the beta-position of the alpha-aminoacrylate species Salmonella enterica subsp. enterica serovar Typhimurium

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.000003
-
 L-threonine recombinant wild-type enzyme, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.00001
-
 L-threonine recombinant mutant L166A, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.000023
-
 L-threonine recombinant mutant L166V, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.003
-
 L-serine recombinant mutant L166A, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.058
-
 L-serine recombinant mutant L166V, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium
0.22
-
 L-serine recombinant wild-type enzyme, pH 7.8, 37°C Salmonella enterica subsp. enterica serovar Typhimurium