Cloned (Comment) | Organism |
---|---|
overproduction in Escherichia coli of an enzyme derivative with the mutation of the active-site serine residue to Cys (Pro-Tk-S359C), its derivatives lacking the N-propeptide (ProC-Tk-S359C) and both propeptides (Tk-S359C), and a His-tagged form of the C-propeptide (ProC*) | Thermococcus kodakarensis |
Protein Variants | Comment | Organism |
---|---|---|
additional information | construction of enzyme derivatives with the mutation of the active-site serine residue to Cys (Pro-Tk-S359C), Pro-Tk-S359C derivative lacking the N-propeptide (ProC-Tk-S359C) and both propeptides (Tk-S359C), and a His-tagged form of the isolated C-propeptide (ProC*). Comparison of the susceptibility of ProC* to proteolytic degradation in the presence and absence of Ca2+ suggests that the C-propeptide becomes highly resistant to proteolytic degradation in the presence of Ca2+ | Thermococcus kodakarensis |
Pro-Tk-S359C | construction of an enzyme derivative with the mutation of the active-site serine residue to Cys (Pro-Tk-S359C). Pro-Tk-S359C is purified mostly in an autoprocessed form in which the N-propeptide is autoprocessed but the isolated N-propeptide (ProN) forms a stable complex with ProC-Tk-S359C, indicating that the N-propeptide is autoprocessed first | Thermococcus kodakarensis |
ProC-Tk-S359C | construction of an enzyme derivative lacking the N-propeptide (ProC-Tk-S359C). The C-propeptide is autoprocessed and degraded when ProC-Tk-S359C is incubated at 80 °C in the absence of Ca2+. However, it is not autoprocessed in the presence of Ca2+. The enzymatic activity of ProC-Tk-S359C is higher than (but comparable to) that of Tk-S359C, an enzyme derivatives lacking both propeptides, suggesting that the C-propeptide is not important for activity. The Tm value of ProC-Tk-S359C is higher than that of Tk-S359C by 25.9°C in the absence of Ca2+ and 7.5 °C in the presence of Ca2+, indicating that the C-propeptide contributes to the stabilization of ProC-Tk-S359C | Thermococcus kodakarensis |
S359C | S359C is more stable than S359A. Tm value of is 58.0°C in the presence of 2.5 M GdnHCl and the absence of Ca2+ and 80.1°C in the presence of 6 m GdnHCl and 10 mm CaCl2 | Thermococcus kodakarensis |
Tk-S359C | construction of an enzyme derivative lacking both propeptides (Tk-S359C). The enzymatic activity of ProC-Tk-S359C, an enzyme derivatives lacking the N-propeptide is higher than (but comparable to) that of Tk-S359C, suggesting that the C-propeptide is not important for activity. The Tm value of ProC-Tk-S359C is higher than that of Tk-S359C by 25.9°C in the absence of Ca2+ and 7.5 °C in the presence of Ca2+, indicating that the C-propeptide contributes to the stabilization of ProC-Tk-S359C | Thermococcus kodakarensis |
Molecular Weight [Da] | Molecular Weight Maximum [Da] | Comment | Organism |
---|---|---|---|
63000 | - |
gel filtration, Pro-Tk-S359C (an enzyme derivative with the mutation of the active-site serine residue to Cys) | Thermococcus kodakarensis |
66000 | - |
x * 66000, Pro-Tk-S359C (an enzyme derivative with the mutation of the active-site serine residue to Cys) | Thermococcus kodakarensis |
Organism | UniProt | Comment | Textmining |
---|---|---|---|
Thermococcus kodakarensis | P58502 | sequence including singnal peptide (amino acid 1-24) and propeptide (amino acid 25-106) | - |
Posttranslational Modification | Comment | Organism |
---|---|---|
proteolytic modification | the N-propeptide is autoprocessed first in the maturation process of Pro-Tk-S359C (an enzyme derivative with the mutation of the active-site serine residue to Cys), although the C-propeptide is subsequently autoprocessed and degraded only in the absence of Ca2+. The C-propeptide is not autoprocessed in the presence of Ca2+, suggesting that Pro-Tk-SP derivative lacking N-propeptide (Val114-Gly640) (ProC-Tk-SP) is not an intermediate form but is the mature form of the enzyme. It is shown that the C-propeptide contributes to the stabilization of ProC-Tk-S359C | Thermococcus kodakarensis |
Purification (Comment) | Organism |
---|---|
- |
Thermococcus kodakarensis |
Substrates | Comment Substrates | Organism | Products | Comment (Products) | Rev. | Reac. |
---|---|---|---|---|---|---|
azocasein + H2O | - |
Thermococcus kodakarensis | ? | - |
? |
Subunits | Comment | Organism |
---|---|---|
? | x * 66000, Pro-Tk-S359C (an enzyme derivative with the mutation of the active-site serine residue to Cys) | Thermococcus kodakarensis |
Synonyms | Comment | Organism |
---|---|---|
Tk-SP | - |
Thermococcus kodakarensis |
Temperature Optimum [°C] | Temperature Optimum Maximum [°C] | Comment | Organism |
---|---|---|---|
80 | - |
assay at | Thermococcus kodakarensis |
Temperature Stability Minimum [°C] | Temperature Stability Maximum [°C] | Comment | Organism |
---|---|---|---|
58 | - |
Tm-value of Tk-S359C in absence of CaCl2. The Tm value of ProC-Tk-S359C is higher than that of Tk-S359C by 25.9°C in the absence of CaCl2 and 7.5°C in the presence of 10 mM CaCl2, indicating that the C-propeptide of ProC-Tk-S359C contributes to the stabilization of the protein by 25.9°C in Tm in the absence of Ca2+ and 7.5°C in Tm in the presence of Ca2+ | Thermococcus kodakarensis |
80.1 | - |
Tm-value of Tk-S359C in presence of 10 mM CaCl2. The Tm value of ProC-Tk-S359C is higher than that of Tk-S359C by 25.9°C in the absence of CaCl2 and 7.5°C in the presence of 10 mM CaCl2, indicating that the C-propeptide of ProC-Tk-S359C contributes to the stabilization of the protein by 25.9°C in Tm in the absence of Ca2+ and 7.5°C in Tm in the presence of Ca2+ | Thermococcus kodakarensis |
83.9 | - |
Tm-value of ProC-Tk-S359C in absence of CaCl2. The Tm value of ProC-Tk-S359C is higher than that of Tk-S359C by 25.9°C in the absence of CaCl2 and 7.5°C in the presence of 10 mM CaCl2, indicating that the C-propeptide of ProC-Tk-S359C contributes to the stabilization of the protein by 25.9°C in Tm in the absence of Ca2+ and 7.5°C in Tm in the presence of Ca2+ | Thermococcus kodakarensis |
87.6 | - |
Tm-value of ProC-Tk-S359C in presence of 10 mM CaCl2. The Tm value of ProC-Tk-S359C is higher than that of Tk-S359C by 25.9°C in the absence of CaCl2 and 7.5°C in the presence of 10 mM CaCl2, indicating that the C-propeptide of ProC-Tk-S359C contributes to the stabilization of the protein by 25.9°C in Tm in the absence of Ca2+ and 7.5°C in Tm in the presence of Ca2+ | Thermococcus kodakarensis |
pH Optimum Minimum | pH Optimum Maximum | Comment | Organism |
---|---|---|---|
7.5 | - |
assay at | Thermococcus kodakarensis |