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Literature summary for 3.5.3.6 extracted from

  • Rodriguez, S.B.; Stitt, B.L.; Ash, D.E.
    Expression of peptidylarginine deiminase from Porphyromonas gingivalis in Escherichia coli: enzyme purification and characterization (2009), Arch. Biochem. Biophys., 488, 14-22.
    View publication on PubMedView publication on EuropePMC

Cloned(Commentary)

Cloned (Comment) Organism
two recombinant forms of the Porphyromonas gingivalis PAD, a truncated form with a 43-amino acid N-terminal deletion and the full-length form of PAD, are expressed in Escherichia coli BL21 Star (DE3)pLysS cells Porphyromonas gingivalis

Inhibitors

Inhibitors Comment Organism Structure
Cd2+ 53% inhibition at 1 mM Cd2+ Porphyromonas gingivalis
Zn2+ at 1 mM Zn2+, the enzyme activity is inhibited by 26% Porphyromonas gingivalis

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
0.01 0.02 N-alpha-benzoyl-L-arginine in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.021
-
bradykinin-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.033
-
bradykinin antagonist-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.07
-
L-Pro-L-Arg-L-Phe in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.089
-
L-Pro-L-Phe-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
1.2
-
L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
1.5
-
Nomega-hydroxy-L-arginine in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
1.9
-
D-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
4.7
-
homoarginine
-
Porphyromonas gingivalis
6
-
L-canavanine-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
6.9
-
L-Arginine hydroxamate in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
18
-
benzoyl-L-arginine ethyl ester in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
31
-
agmatine-L-Arg
-
Porphyromonas gingivalis

Metals/Ions

Metals/Ions Comment Organism Structure
additional information PAD is not a metalloenzyme Porphyromonas gingivalis

Molecular Weight [Da]

Molecular Weight [Da] Molecular Weight Maximum [Da] Comment Organism
51000
-
purified recombinant truncated PAD (missing 43 N-terminal amino acid residues but including Xpress and poly-His tags), SDS-PAGE Porphyromonas gingivalis
53000
-
active enzyme, gel filtration Porphyromonas gingivalis
60880
-
truncated PAD with its poly-His and Xpress epitope tags, calculated from amino acid sequence Porphyromonas gingivalis
60890
-
MALDI-TOF mass spectrometry Porphyromonas gingivalis

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
protein L-Arg + H2O Porphyromonas gingivalis
-
protein L-citrulline + NH3
-
?
protein L-Arg + H2O Porphyromonas gingivalis BAA-308-5 (W83)
-
protein L-citrulline + NH3
-
?

Organism

Organism UniProt Comment Textmining
Porphyromonas gingivalis
-
-
-
Porphyromonas gingivalis BAA-308-5 (W83)
-
-
-

Purification (Commentary)

Purification (Comment) Organism
Ni+-NTA resin column chromatography, ultrafiltration, and gel filtration Porphyromonas gingivalis

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
0.000345
-
crude extract, pH 9.5, at 37°C Porphyromonas gingivalis
0.175
-
after purification, pH 9.5, at 37°C Porphyromonas gingivalis

Storage Stability

Storage Stability Organism
25°C or -80°C, in the presence of 0.5 mM 2-mercaptoethanol or 1 mM dithiothreitol and 0.1% (w/v) sodium azide in 50 mM potassium phosphate buffer (pH 7.8), 2 weeks, no appreciable loss of activity Porphyromonas gingivalis
4°C or -20°C, in the presence of 0.5 mM 2-mercaptoethanol or 1 mM dithiothreitol and 0.1% (w/v) sodium azide in 50 mM potassium phosphate buffer (pH 7.8), 2 weeks, aggregation of the protein occurs Porphyromonas gingivalis

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
agmatine-L-Arg + H2O 3.2% activity compared to L-Arg Porphyromonas gingivalis N5-carbamoyl-N2-[4-(carbamoylamino)butyl]-L-ornithine + NH3
-
?
benzoyl-L-arginine ethyl ester + H2O 8.1% activity compared to L-Arg Porphyromonas gingivalis benzoyl-L-citrulline ethyl ester + NH3
-
?
benzoyl-L-arginine ethyl ester + H2O 8.1% activity compared to L-Arg Porphyromonas gingivalis BAA-308-5 (W83) benzoyl-L-citrulline ethyl ester + NH3
-
?
bradykinin-L-Arg + H2O 18.8% activity compared to L-Arg Porphyromonas gingivalis bradykinin-L-citrulline + NH3
-
?
D-Arg + H2O 26.4% activity compared to L-Arg Porphyromonas gingivalis ?
-
?
homoarginine + H2O 50% activity compared to L-Arg Porphyromonas gingivalis (2S)-2-amino-6-[(aminocarbonyl)amino]hexanoic acid + NH3
-
?
KRPPGFSPL + H2O 3.1% activity compared to L-Arg Porphyromonas gingivalis L-lysyl-N5-carbamoyl-L-ornithyl-L-prolyl-L-prolylglycyl-L-phenylalanyl-L-seryl-L-prolyl-L-leucine + NH3
-
?
L-Arg + H2O 100% activity Porphyromonas gingivalis citrulline + NH3
-
?
L-Arg + H2O 100% activity Porphyromonas gingivalis BAA-308-5 (W83) citrulline + NH3
-
?
L-arginine hydroxamate + H2O 68.2% activity compared to L-Arg Porphyromonas gingivalis ?
-
?
L-canavanine-L-Arg + H2O 2.5% activity compared to L-Arg Porphyromonas gingivalis O-ureido-L-homoserine + NH3
-
?
L-Pro-L-Arg-L-Phe + H2O 3.4% activity compared to L-Arg Porphyromonas gingivalis ?
-
?
L-Pro-L-Phe-L-Arg + H2O 78.2% activity compared to L-Arg Porphyromonas gingivalis ?
-
?
additional information no activity with Nomega,Nomega-dimethyl-L-arginine Porphyromonas gingivalis ?
-
?
additional information no activity with Nomega,Nomega-dimethyl-L-arginine Porphyromonas gingivalis BAA-308-5 (W83) ?
-
?
N-alpha-benzoyl-L-arginine + H2O 54.5% activity compared to L-Arg Porphyromonas gingivalis N-alpha-benzoyl-L-citrulline + NH3
-
?
N-alpha-benzoyl-L-arginine + H2O 54.5% activity compared to L-Arg Porphyromonas gingivalis BAA-308-5 (W83) N-alpha-benzoyl-L-citrulline + NH3
-
?
Nomega-hydroxy-L-arginine + H2O 2.1% activity compared to L-Arg Porphyromonas gingivalis Nomega-hydroxy-L-citrulline + NH3
-
?
protein L-Arg + H2O
-
Porphyromonas gingivalis protein L-citrulline + NH3
-
?
protein L-Arg + H2O
-
Porphyromonas gingivalis BAA-308-5 (W83) protein L-citrulline + NH3
-
?

Synonyms

Synonyms Comment Organism
PAD
-
Porphyromonas gingivalis
peptidylarginine deiminase
-
Porphyromonas gingivalis

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
55
-
-
Porphyromonas gingivalis

Temperature Stability [°C]

Temperature Stability Minimum [°C] Temperature Stability Maximum [°C] Comment Organism
4
-
PAD is cold sensitive, it aggregates within 30 min at 4°C, but aggregated PAD displays variable activity for up to one week after re-suspension with storage buffer Porphyromonas gingivalis
100
-
no product formation is observed in reaction mixtures containing the PAD sample that have been boiled Porphyromonas gingivalis

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.0046
-
Nomega-hydroxy-L-arginine in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.0052
-
L-canavanine-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.0068
-
bradykinin antagonist-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.007
-
agmatine-L-Arg
-
Porphyromonas gingivalis
0.0075
-
L-Pro-L-Arg-L-Phe in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.019
-
benzoyl-L-arginine ethyl ester in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.043
-
bradykinin-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.058
-
D-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.11
-
homoarginine
-
Porphyromonas gingivalis
0.12
-
N-alpha-benzoyl-L-arginine in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.15
-
L-Arginine hydroxamate in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.17
-
L-Pro-L-Phe-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.22
-
L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
9.5
-
-
Porphyromonas gingivalis

pH Range

pH Minimum pH Maximum Comment Organism
6 11 38.5% activity remaining at pH 6.0 and 3.4% activity remaining at pH 11.0 Porphyromonas gingivalis

pH Stability

pH Stability pH Stability Maximum Comment Organism
7 9.5 the enzyme at 13 mg/ml remains active at pH 7.0, 8.0 and 9.5 for several days at 25°C Porphyromonas gingivalis

Cofactor

Cofactor Comment Organism Structure
additional information PAD does not need a cofactor for catalysis and stability Porphyromonas gingivalis

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
0.00023
-
agmatine-L-Arg
-
Porphyromonas gingivalis
0.00093
-
L-canavanine-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.00099
-
benzoyl-L-arginine ethyl ester in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.003
-
Nomega-hydroxy-L-arginine in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.022
-
L-Arginine hydroxamate in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.024
-
homoarginine
-
Porphyromonas gingivalis
0.031
-
D-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.11
-
L-Pro-L-Arg-L-Phe in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.19
-
L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
0.21
-
bradykinin antagonist-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
1.9
-
L-Pro-L-Phe-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
2.1
-
bradykinin-L-Arg in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis
6 12 N-alpha-benzoyl-L-arginine in 50 mM CHES/HCl, pH 9.5 containing 10 mM dithiothreitol, at 37°C Porphyromonas gingivalis