3.4.21.B57: pernisine
This is an abbreviated version!
For detailed information about pernisine, go to the full flat file.
Word Map on EC 3.4.21.B57
-
3.4.21.B57
-
hyperthermophilic
-
archaeon
-
subtilisin-like
-
pernix
-
aeropyrum
-
subtilisins
-
thermococcus
-
prion
-
medicine
-
kodakaraensis
-
proregion
-
mesophilic
-
autoprocessed
-
detergents
-
edta
-
codon-optimised
-
far-uv
-
tk-subtilisin
-
high-temperature
-
cacl2
-
n-propeptide
-
roll
-
hyperthermostable
- 3.4.21.B57
-
hyperthermophilic
- archaeon
-
subtilisin-like
- pernix
-
aeropyrum
- subtilisins
-
thermococcus
- prion
- medicine
- kodakaraensis
-
proregion
-
mesophilic
-
autoprocessed
- detergents
- edta
-
codon-optimised
-
far-uv
- tk-subtilisin
-
high-temperature
- cacl2
- n-propeptide
-
roll
-
hyperthermostable
Reaction
the enzyme can digest the pathological prion protein isoform (PrPSc) from different species, e.g. human, bovine, deer and mouse =
Synonyms
pernisine, subtilase, Tk-SP, Tk-subtilisin, TKS
ECTree
3.4.21.B57 pernisineAdvanced search results
results (
results (Engineering
Engineering on EC 3.4.21.B57 - pernisine
Please wait a moment until all data is loaded. This message will disappear when all data is loaded.
top
PROTEIN VARIANTS 
ORGANISM
UNIPROT
COMMENTARY 
LITERATURE
S355A
site-directed mutagenesis, catalytically inactive active site mutant
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
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
S324A
S324C
site-directed mutagenesis, structure comparison of the mutant pro-enzyme with the wild-type pro-enzyme
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
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
additional information
S324A
the crystal structure of the active site mutant of Tk-subtilisin (S324A-subtilisin), which is refolded in the presence of Ca2+ and absence of Tk-propeptide, is determined at 2.16 A resolution. This structure is the same as that of Tk-subtilisin matured from Pro-Tk-subtilisin. The counting of amino acids refers to the enzyme protein without the signal peptide (amino acid 1-24) and the propeptide (amino acid 25-106)
S324A
site-directed mutagenesis, structure comparison of the mutant pro-enzyme with the wild-type pro-enzyme
additional information
construction of a series of active-site mutants of with (Tk-S359A/C) and without (Tk-S359A/CDeltaJ) beta-jelly roll domain. Both Tk-S359C and Tk-S359CDeltaJ exhibit protease activities, indicating that the beta-jelly roll domain is not required for folding or activity. The Tm value of Tk-S359ADeltaJ determined by far-UV CD spectroscopy in the presence of 10-mM CaCl2 is lower than that of Tk-S359A by 29.4°C. The Tm value of Tk-S359A is decreased by 29.5 °C by the treatment with 10 mM ethylenediaminetetraacetic acid, indicating that the beta-jelly roll domain contributes to the stabilization of Tk-S359A only in a Ca2+-bound form
additional information
-
construction of a series of active-site mutants of with (Tk-S359A/C) and without (Tk-S359A/CDeltaJ) beta-jelly roll domain. Both Tk-S359C and Tk-S359CDeltaJ exhibit protease activities, indicating that the beta-jelly roll domain is not required for folding or activity. The Tm value of Tk-S359ADeltaJ determined by far-UV CD spectroscopy in the presence of 10-mM CaCl2 is lower than that of Tk-S359A by 29.4°C. The Tm value of Tk-S359A is decreased by 29.5 °C by the treatment with 10 mM ethylenediaminetetraacetic acid, indicating that the beta-jelly roll domain contributes to the stabilization of Tk-S359A only in a Ca2+-bound form
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+
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+
additional information
Pro-Tk-subtilisin variants with complete amino acid substitutions at Gly56 are constructed. Pro-G56W, Pro-G56E and Pro-G56S are overproduced, purified, and characterized. Their maturation rates increase in the order wild-type enzyme or = G56W-propeptide > G56S-propeptide > G56E-propeptide, indicating that they are inversely correlated with the maturation rates of Pro7-Tk-subtilisin and its derivatives
additional information
the Leu69Pro mutation in the propeptide accelerates the maturation of Pro-Tk-subtilisin by reducing the binding ability of Tk-propeptide to Tk-subtilisin
additional information
the Pro-Tk-subtilisin derivative with the F17His mutation (Pro-F17H), Tk-propeptide derivative with the same mutation (F17H-propeptide), and two active-site mutants of Pro-F17H (Pro-F17H/S324A and Pro-F17H/S324C) are constructed
additional information
to analyze the role of the Ca2+-binding loop, three mutant proteins, Deltaloop-Tk-subtilisin (Ca2+-binding loop is removed), DeltaCa2-Pro-S324A (Ca2+-binding site Ca2 is removed), and DeltaCa3-Pro-S324A (Ca2+-binding site Ca3 is removed), are constructed. The structures of DeltaCa2-Pro-S324A (Ca2+-binding site Ca 2 is removed) and DeltaCa3-Pro-S324A (Ca2+-binding site Ca3 is removed) are identical to that of Pro-S324A, except that they lack the Ca2 and Ca3 sites, respectively, and the structure of the Ca2+-binding loop is destabilized. These proteins are slightly more stable than Pro-S324A
additional information
generation of IS1-deletion mutants of S324A and S324C enzyme variants
