Cloned (Comment) | Organism |
---|---|
expression of N-terminally His-tagged wild-type and mutant enzymes in Escherichia coli, recombinant wild-type enzyme shows a lower activity than the native one | Pedobacter heparinus |
Protein Variants | Comment | Organism |
---|---|---|
H105A | PCR overlap extension site-directed mutagenesis, very low expression level, no measurement of activity possible, reduced expression level | Pedobacter heparinus |
H110A | PCR overlap extension site-directed mutagenesis, reduced kcat, highly reduced Km compared to both recombinant and native wild-type enzymes, reduced expression level | Pedobacter heparinus |
H139A | PCR overlap extension site-directed mutagenesis, reduced kcat and increased Km compared to both recombinant and native wild-type enzymes, reduced expression level | Pedobacter heparinus |
H152A | PCR overlap extension site-directed mutagenesis, reduced Km and a kcat value between the recombinant and the native wild-type enzyme | Pedobacter heparinus |
H225A | PCR overlap extension site-directed mutagenesis, Km is the same as for the recombinant wild-type, reduced kcat | Pedobacter heparinus |
H234A | PCR overlap extension site-directed mutagenesis, Km is similar to the recombinant wild-type, reduced kcat | Pedobacter heparinus |
H241A | PCR overlap extension site-directed mutagenesis, highly reduced kcat, highly reduced Km compared to both recombinant and native wild-type enzymes | Pedobacter heparinus |
H295A | PCR overlap extension site-directed mutagenesis, inactive mutant | Pedobacter heparinus |
H36A | PCR overlap extension site-directed mutagenesis, reduced Km and a kcat value between the recombinant and the native wild-type enzyme | Pedobacter heparinus |
H424A | PCR overlap extension site-directed mutagenesis, reduced Km and kcat | Pedobacter heparinus |
H469A | PCR overlap extension site-directed mutagenesis, reduced Km and increased kcat compared to both recombinant and native wild-type enzymes | Pedobacter heparinus |
H510A | PCR overlap extension site-directed mutagenesis, inactive mutant | Pedobacter heparinus |
H539A | PCR overlap extension site-directed mutagenesis, Km between the recombinant and the native wild-type enzyme, kcat is increased compared to both wild-type enzymes | Pedobacter heparinus |
Inhibitors | Comment | Organism | Structure |
---|---|---|---|
diethyl dicarbonate | inactivation, 80% reversible by hydroxylamine within 6 h, mapping of modified histidine residues | Pedobacter heparinus |
KM Value [mM] | KM Value Maximum [mM] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
0.009 | - |
heparan sulfate | mutant H110A, pH 7.6, 35°C | Pedobacter heparinus | |
0.016 | - |
heparan sulfate | mutant H241A, pH 7.6, 35°C | Pedobacter heparinus | |
0.058 | - |
heparan sulfate | mutant H152A, pH 7.6, 35°C | Pedobacter heparinus | |
0.059 | - |
heparan sulfate | mutant H424A, pH 7.6, 35°C | Pedobacter heparinus | |
0.071 | - |
heparan sulfate | mutant H469A, pH 7.6, 35°C | Pedobacter heparinus | |
0.075 | - |
heparan sulfate | mutant H234A, pH 7.6, 35°C | Pedobacter heparinus | |
0.08 | - |
heparan sulfate | recombinant wild-type enzyme and mutant H225A, pH 7.6, 35°C | Pedobacter heparinus | |
0.092 | - |
heparan sulfate | mutant H539A, pH 7.6, 35°C | Pedobacter heparinus | |
0.098 | - |
heparan sulfate | mutant H36A, pH 7.6, 35°C | Pedobacter heparinus | |
0.143 | - |
heparan sulfate | native wild-type enzyme, pH 7.6, 35°C | Pedobacter heparinus | |
0.191 | - |
heparan sulfate | mutant H139A, pH 7.6, 35°C | Pedobacter heparinus |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Pedobacter heparinus | - |
- |
- |
Purification (Comment) | Organism |
---|---|
recombinant His-tagged wild-type and mutants from Escherichia coli | Pedobacter heparinus |
Reaction | Comment | Organism | Reaction ID |
---|---|---|---|
[GlcAbeta(1-4)GlcNbeta(1-4)]n = [GlcAbeta(1-4)GlcNbeta(1-4)]n-1 + 2-amino-2-deoxy-4-O-alpha-L-threo-hex-4-enopyranuronosyl-beta-D-glucopyranose | His295 and His510 are essential for catalytic activity | Pedobacter heparinus |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
heparan sulfate | - |
Pedobacter heparinus | ? | - |
? |
Synonyms | Comment | Organism |
---|---|---|
heparinase III | - |
Pedobacter heparinus |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
35 | - |
assay at | Pedobacter heparinus |
Turnover Number Minimum [1/s] | Turnover Number Maximum [1/s] | Substrate | Comment | Organism | Structure |
---|---|---|---|---|---|
5 | - |
heparan sulfate | mutant H241A, pH 7.6, 35°C | Pedobacter heparinus | |
22 | - |
heparan sulfate | mutant H225A, pH 7.6, 35°C | Pedobacter heparinus | |
23 | - |
heparan sulfate | mutant H234A, pH 7.6, 35°C | Pedobacter heparinus | |
24 | - |
heparan sulfate | mutant H424A, pH 7.6, 35°C | Pedobacter heparinus | |
37 | - |
heparan sulfate | mutant H110A, pH 7.6, 35°C | Pedobacter heparinus | |
68 | - |
heparan sulfate | mutant H139A, pH 7.6, 35°C | Pedobacter heparinus | |
78 | - |
heparan sulfate | recombinant wild-type enzyme, pH 7.6, 35°C | Pedobacter heparinus | |
83 | - |
heparan sulfate | mutant H152A, pH 7.6, 35°C | Pedobacter heparinus | |
86 | - |
heparan sulfate | mutant H36A, pH 7.6, 35°C | Pedobacter heparinus | |
94 | - |
heparan sulfate | native wild-type enzyme, pH 7.6, 35°C | Pedobacter heparinus | |
100 | - |
heparan sulfate | mutant H469A, pH 7.6, 35°C | Pedobacter heparinus | |
132 | - |
heparan sulfate | mutant H539A, pH 7.6, 35°C | Pedobacter heparinus |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.6 | - |
assay at | Pedobacter heparinus |
Ki Value [mM] | Ki Value maximum [mM] | Inhibitor | Comment | Organism | Structure |
---|---|---|---|---|---|
additional information | - |
additional information | inactivation kinetics | Pedobacter heparinus |