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Literature summary extracted from

  • Zeng, J.; Gao, X.; Dai, Z.; Tang, B.; Tang, X.F.
    Effects of metal ions on stability and activity of hyperthermophilic pyrolysin and further stabilization of this enzyme by modification of a Ca2+-binding site (2014), Appl. Environ. Microbiol., 80, 2763-2772.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.4.21.B55 expression in Escherichia coli. At high temperatures, pyrolysin proform (Pls) is converted to mature pyrolysin (mPls) via autoprocessing of both the N- and C-terminal propeptides Pyrococcus furiosus

Protein Variants

EC Number Protein Variants Comment Organism
3.4.21.B55 D55A after teatment at 95°C for 12 h the mutant enzyme displays similar levels of residual activity to that of the wild-type enzyme. When incubated with 2 mM EDTA at 95°C for 10 min, the mutant enzyme is completely inactivated, whereas the wild-type enzyme displays a residual activity of 52.6% Pyrococcus furiosus
3.4.21.B55 D55A/D58A mutant enzyme is capable of maturation at 95°C and also exhibits thermal stability similar to wild-type enzyme in the absence of chelating agents Pyrococcus furiosus
3.4.21.B55 D55A/D58A/E59A the proform of the mutant enzyme is unable to mature at 95°C Pyrococcus furiosus
3.4.21.B55 D55A/E59A less stable than wild-type enzyme under non-chelating conditions Pyrococcus furiosus
3.4.21.B55 D55N/D58N/E59Q less stable than wild-type enzyme under non-chelating conditions Pyrococcus furiosus
3.4.21.B55 D58A after teatment at 95°C for 12 h the mutant enzyme displays similar levels of residual activity to that of the wild-type enzyme. When incubated with 2 mM EDTA at 95°C for 10 min, the mutant enzyme retains 9.6% of its original activity, whereas the wild-type enzyme displays a residual activity of 52.6% Pyrococcus furiosus
3.4.21.B55 D58A/E59A less stable than wild-type enzyme under non-chelating conditions Pyrococcus furiosus
3.4.21.B55 D818N/D820N half-life of the mutant enzyme at 95°C is 18 h, compared to 12 h measured for the wild-type enzyme. Inactivation of the enzymes at 100-115°C results in retention of a higher level of its original activity by the mutant enzyme compared to wild-type enzyme. The mutant enzyme displays higher specific activity towards azocasein than wild-type enzyme in the temperature range of 50-110°C. The mutant enzyme also shows higher initial velocity of hydrolysis than wild-type enzyme with azocasein or succinyl-Ala-Ala-Pro-Lys-4-nitroanilide as substrate Pyrococcus furiosus
3.4.21.B55 E59A after teatment at 95°C for 12 h the mutant enzyme exhibits a slightly lower residual activity than wild-type enzyme. When incubated with 2 mM EDTA at 95°C for 10 min, the mutant enzyme is completely inactivated, whereas the wild-type enzyme displays a residual activity of 52.6% Pyrococcus furiosus
3.4.21.B55 R276A after teatment at 95°C for 12 h the mutant enzyme displays similar levels of residual activity to that of the wild-type enzyme. When incubated with 2 mM EDTA at 95°C for 10 min, the mutant enzyme retains 9.6% of its original activity, whereas the wild-type enzyme displays a residual activity of 52.6% Pyrococcus furiosus

General Stability

EC Number General Stability Organism
3.4.21.B55 binding of Ca2+ increases the stability of the PPC domain and, in turn, stabilizes the global structure of the enzyme by modulating interdomain interactions Pyrococcus furiosus
3.4.21.B55 supplementation of Na+, Ca2+, or Mg2+ salts at concentrations similar to those in seawater destabilizes recombinant pyrolysin. Ca2+ and Mg2+ are stronger denaturants than Na+ for pyrolysin. Ca2+ also acts a stabilizer of pyrolysin by binding to the enzyme at specific sites. The destabilizing effect of metal ions on pyrolysin appears to be related to the disturbance of surface electrostatic interactions of the enzyme Pyrococcus furiosus

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining
3.4.21.B55 extracellular
-
 Pyrococcus furiosus
-
-

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.4.21.B55 Ca2+ supplementation of Na+, Ca2+, or Mg2+ salts at concentrations similar to those in seawater leads to an increase in enzyme activity. The activity level increases with the increase of the salt concentrations from 0.1 mM to 10 mM Pyrococcus furiosus
3.4.21.B55 Mg2+ supplementation of Na+, Ca2+, or Mg2+ salts at concentrations similar to those in seawater leads to an increase in enzyme activity. The activity level increases with the increase of the salt concentrations from 5 mM to 50 mM Pyrococcus furiosus
3.4.21.B55 Na+ supplementation of Na+, Ca2+, or Mg2+ salts at concentrations similar to those in seawater leads to an increase in enzyme activity. The activity level increases with the increase of the salt concentrations from 40 mM to 600 mM Pyrococcus furiosus

Organism

EC Number Organism UniProt Comment Textmining
3.4.21.B55 Pyrococcus furiosus
-
-
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.4.21.B55
-
 Pyrococcus furiosus

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.4.21.B55 azocasein + H2O
-
 Pyrococcus furiosus ?
-
 ?
3.4.21.B55 succinyl-Ala-Ala-Pro-Lys-4-nitroanilide + H2O
-
 Pyrococcus furiosus succinyl-Ala-Ala-Pro-Lys + 4-nitroaniline
-
 ?

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.4.21.B55 95
-
 assay at Pyrococcus furiosus
3.4.21.B55 110
-
 wild-type enzyme and mutant enzyme D818N/D820N Pyrococcus furiosus

Temperature Range [°C]

EC Number Temperature Minimum [°C] Temperature Maximum [°C] Comment Organism
3.4.21.B55 90 120 90°C: about 60% of maximal activity, 120°C: about 70% of maximal activity Pyrococcus furiosus

Temperature Stability [°C]

EC Number Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
3.4.21.B55 95
-
 half-life of wild-type enzyme: 12 h, half-life of mutant enzyme D818N/D820N is 18 h Pyrococcus furiosus
3.4.21.B55 105
-
 20 min, wild-type enzyme loses about 45% of its activity, mutant enzyme D818N/D820N loses about 30% of its activity Pyrococcus furiosus
3.4.21.B55 110
-
 5 min, wild-type enzyme loses about 30% of its activity, mutant enzyme D818N/D820N loses about 20% of its activity Pyrococcus furiosus
3.4.21.B55 115
-
 5 min, wild-type enzyme loses about 75% of its activity, mutant enzyme D818N/D820N loses about 70% of its activity Pyrococcus furiosus

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.4.21.B55 7.8
-
 assay at Pyrococcus furiosus