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Literature summary for 6.3.2.17 extracted from

  • Sheng, Y.; Khanam, N.; Tsaksis, Y.; Shi, X.M.; Lu, Q.S.; Bognar, A.L.
    Mutagenesis of folylpolyglutamate synthetase indicates that dihydropteroate and tetrahydrofolate bind to the same site (2008), Biochemistry, 47, 2388-2396.
    View publication on PubMed

Protein Variants

Protein Variants Comment Organism
A155H drastic increases in Km values Escherichia coli
A155W 13fold increase in Km value for dihydropteroate Escherichia coli
D151A mutation in residue specific for dihydropteroate binding, mutant is defective in using tetrahydrofolate as substrate Lacticaseibacillus casei
D154A mutation in residue specific for dihydropteroate binding, 200fold increase in Km value for substrate 5,6,7,8-tetrahydropteroyl-gamma-(L-Glu)2 Escherichia coli
F121A 50fold increase in Km for 5,10-methylene-5,6,7,8-tetrahydrofolic acid Lacticaseibacillus casei
F75A increase in Km value for 5,10-methylene-5,6,7,8-tetrahydrofolic acid, decreases in Km values for ATP and L-Glu Lacticaseibacillus casei
additional information construction of domain-swap chimera proteins between the Escherichia coli and the Lactobacillus casei enzymes. Chimera possess both folate or pteroate binding properties and enzymatic activities of their amino terminal portion, suggesting that the N-terminal domain determines the folate substrate specificity. Mutants with swapped omega loops, containing a folate binding site, retain the activities and folate or pteroate binding properties of the rest of the enzyme. Mutating Lactobacillus casei folypolyglutamate synthetase to contain an Escherichia coli folypolyglutamate synthetase dihydropteroate binding loop does not alter its substrate specificity to using dihydropteroate as a substrate Lacticaseibacillus casei
additional information expression of the amino-terminal domain with residues 1-287. The domain binds tetrahydrofolate and dihydropteroate with the same affinity as the intact enzyme. Construction of domain-swap chimera proteins between the Escherichia coli and the Lactobacillus casei enzymes. Chimera possess both folate or pteroate binding properties and enzymatic activities of their amino terminal portion, suggesting that the N-terminal domain determines the folate substrate specificity. Mutants with swapped omega loops, containing a folate binding site, retain the activities and folate or pteroate binding properties of the rest of the enzyme. Mutating Lactobacillus casei folypolyglutamate synthetase to contain an Escherichia coli folypolyglutamate synthetase dihydropteroate binding loop does not alter its substrate specificity to using dihydropteroate as a substrate Escherichia coli
R15E increases in Km values Lacticaseibacillus casei
S152A decrease in Vmax Lacticaseibacillus casei
S152W increases in Km values Lacticaseibacillus casei
T119W 100fold increase in Km for 5,10-methylene-5,6,7,8-tetrahydrofolic acid Lacticaseibacillus casei
T122H drastic increases in Km values Escherichia coli
T122W drastic increases in Km values Escherichia coli
Y414A decrease in Km values for ATP, increase in Km for L-Glu Lacticaseibacillus casei

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.0063
-
7,8-dihydropteroate wild-type, pH 9.8 Lacticaseibacillus casei
0.0063
-
7,8-dihydropteroate wild-type, pH 9.8 Escherichia coli
0.01
-
5,6,7,8-tetrahydrofolic acid mutant A155W, pH 9.8 Escherichia coli
0.01
-
5,10-methylene-5,6,7,8-tetrahydrofolyl-(L-Glu)2 mutant D151A, pH 9.8 Lacticaseibacillus casei
0.032
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid wild-type Lacticaseibacillus casei
0.034
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant S152A, pH 9.8 Lacticaseibacillus casei
0.037
-
ATP mutant D154A, pH 9.8 cosubstrate 7,8-dihydropteroate Escherichia coli
0.048
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant Y414A, pH 9.8 Lacticaseibacillus casei
0.05
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid wild-type Escherichia coli
0.053
-
L-Glu mutant F75A, pH 9.8 Lacticaseibacillus casei
0.054
-
ATP wild-type, pH 9.8 Escherichia coli
0.065
-
ATP mutant A155H, pH 9.8 cosubstrate 7,8-dihydropteroate Escherichia coli
0.071
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, pH 9.8 Lacticaseibacillus casei
0.071
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, pH 9.8 Escherichia coli
0.076
-
ATP mutant A155H, pH 9.8, cosubstrate 5,6,7,8-tetrahydrofolic acid Escherichia coli
0.08
-
7,8-dihydropteroate mutant A155W, pH 9.8 Escherichia coli
0.084
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant R15E, pH 9.8 Lacticaseibacillus casei
0.11
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant D151A, pH 9.8 Lacticaseibacillus casei
0.116
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant S152W, pH 9.8 Lacticaseibacillus casei
0.118
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid chimera containing N-terminal residues 1-71 of Lactobacillus casei enzyme, residues 82-93 of Escherichia coli enzyme, and residues 83-428 of Lactobacillus casei enzyme, pH 9.8 Lacticaseibacillus casei
0.118
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid chimera containing N-terminal residues 1-71 of Lactobacillus casei enzyme, residues 82-93 of Escherichia coli enzyme, and residues 83-428 of Lactobacillus casei enzyme, pH 9.8 Escherichia coli
0.12
-
L-Glu mutant A155H, pH 9.8, cosubstrate 5,6,7,8-tetrahydrofolic acid Escherichia coli
0.134
-
7,8-dihydropteroate chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, pH 9.8 Lacticaseibacillus casei
0.134
-
7,8-dihydropteroate chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, pH 9.8 Escherichia coli
0.14
-
7,8-dihydropteroate mutant D154A, pH 9.8 Escherichia coli
0.15
-
7,8-dihydropteroate mutant T122W, pH 9.8 Escherichia coli
0.17
-
L-Glu mutant A155H, pH 9.8, cosubstrate 7,8-dihydropteroate Escherichia coli
0.18
-
ATP mutant F75A, pH 9.8 Lacticaseibacillus casei
0.2
-
L-Glu mutant D154A, pH 9.8, cosubstrate 5,6,7,8-tetrahydrofolic acid Escherichia coli
0.204
-
5,10-methylene-5,6,7,8-tetrahydrofolyl-(L-Glu)2 mutant R15E, pH 9.8 Lacticaseibacillus casei
0.23
-
ATP mutant D154A, pH 9.8, cosubstrate 5,6,7,8-tetrahydrofolic acid Escherichia coli
0.23
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant F75A, pH 9.8 Lacticaseibacillus casei
0.23
-
ATP mutant Y414A, pH 9.8 Lacticaseibacillus casei
0.292
-
L-Glu chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, pH 9.8 Lacticaseibacillus casei
0.292
-
L-Glu chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, pH 9.8 Escherichia coli
0.3
-
5,6,7,8-tetrahydrofolyl-gamma-(L-Glu)2 mutant D154A, pH 9.8 Escherichia coli
0.3
-
L-Glu wild-type, pH 9.8 Escherichia coli
0.34
-
5,6,7,8-tetrahydrofolic acid mutant D154A, pH 9.8 Escherichia coli
0.39
-
5,6,7,8-tetrahydrofolic acid mutant T122W, pH 9.8 Escherichia coli
0.4
-
5,6,7,8-tetrahydrofolic acid mutant T122H, pH 9.8 Escherichia coli
0.448
-
5,6,7,8-tetrahydrofolic acid mutant A155H, pH 9.8 Escherichia coli
0.47
-
L-Glu wild-type, pH 9.8 Lacticaseibacillus casei
0.485
-
7,8-dihydropteroate mutant A155H, pH 9.8 Escherichia coli
0.555
-
ATP mutant A155W, pH 9.8 cosubstrate 7,8-dihydropteroate Escherichia coli
0.59
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, pH 9.8 Lacticaseibacillus casei
0.59
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, pH 9.8 Escherichia coli
0.98
-
7,8-dihydropteroate mutant T122H, pH 9.8 Escherichia coli
1.5
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant F121A, pH 9.8 Lacticaseibacillus casei
2.7
-
L-Glu mutant Y414A, pH 9.8 Lacticaseibacillus casei
3.3
-
L-Glu mutant D154A, pH 9.8, cosubstrate 5,6,7,8-tetrahydrofolic acid Escherichia coli
3.4
-
ATP mutant D151A, pH 9.8 Lacticaseibacillus casei
3.4
-
ATP wild-type, pH 9.8 Lacticaseibacillus casei
3.8
-
5,10-methylene-5,6,7,8-tetrahydrofolic acid mutant T119W, pH 9.8 Lacticaseibacillus casei
5.7
-
ATP chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, pH 9.8 Lacticaseibacillus casei
5.7
-
ATP chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, pH 9.8 Escherichia coli

Organism

Organism UniProt Comment Textmining
Escherichia coli P08192
-
-
Lacticaseibacillus casei P15925
-
-

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.138
-
chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Lacticaseibacillus casei
0.138
-
chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Escherichia coli
0.376
-
chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, substrate 7,8-dihydropteroate, pH 9.8 Lacticaseibacillus casei
0.376
-
chimera containing N-terminal residues 1-81 of Escherichia coli enzyme, residues 72-82 of Lactobacillus casei enzyme, and residues 93-422 of Escherichia coli enzyme, substrate 7,8-dihydropteroate, pH 9.8 Escherichia coli
0.912
-
chimera containing N-terminal residues 1-71 of Lactobacillus casei enzyme, residues 82-93 of Escherichia coli enzyme, and residues 83-428 of Lactobacillus casei enzyme, substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Lacticaseibacillus casei
0.912
-
chimera containing N-terminal residues 1-71 of Lactobacillus casei enzyme, residues 82-93 of Escherichia coli enzyme, and residues 83-428 of Lactobacillus casei enzyme, substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Escherichia coli
1.3
-
chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Lacticaseibacillus casei
1.3
-
chimera containing N-terminal residues 1-298 of Lactobacillus casei enzyme, residues 288-422 of Escherichia coli enzyme, substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Escherichia coli
4.1
-
substrate 7,8-dihydropteroate, pH 9.8 Escherichia coli
11.8
-
substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Lacticaseibacillus casei
13.2
-
substrate 5,10-methylene-5,6,7,8-tetrahydrofolic acid, pH 9.8 Escherichia coli

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ATP + 5,10-methylene-5,6,7,8-tetrahydrofolic acid + L-Glu
-
Escherichia coli ADP + phosphate + 5,10-methylene-5,6,7,8-tetrahydrofolyl-L-Glu
-
?
ATP + 5,10-methylene-5,6,7,8-tetrahydrofolic acid + L-glutamate
-
Lacticaseibacillus casei ADP + phosphate + 5,10-methylene-5,6,7,8-tetrahydrofolyl-Glu
-
?
ATP + 5,10-methylene-5,6,7,8-tetrahydrofolyl-(L-Glu)2 + L-glutamate
-
Lacticaseibacillus casei ADP + phosphate + 5,10-methylene-5,6,7,8-tetrahydrofolyl-(L-Glu)3
-
?
ATP + 5,6,7,8-tetrahydrofolic acid + L-Glu
-
Escherichia coli ADP + phosphate + 5,6,7,8-tetrahydrofolyl-Glu
-
?
ATP + 5,6,7,8-tetrahydrofolyl-gamma-(L-Glu)2 + L-Glu
-
Escherichia coli ADP + phosphate + 5,6,7,8-tetrahydrofolyl-gamma-(L-Glu)3
-
?
ATP + 7,8-dihydropteroate + L-Glu
-
Escherichia coli ADP + phosphate + 7,8-dihydropteroyl-L-Glu
-
?