Any feedback?
Please rate this page
(literature.php)
(0/150)

BRENDA support

Literature summary for 2.7.4.1 extracted from

  • Beassoni, P.R.; Gallarato, L.A.; Boetsch, C.; Garrido, M.N.; Lisa, A.T.
    Pseudomonas aeruginosa exopolyphosphatase is also a polyphosphate ADP phosphotransferase (2015), Enzyme Res., 2015, 404607 .
    View publication on PubMedView publication on EuropePMC

Activating Compound

Activating Compound Comment Organism Structure
Dimethylamine 9% activation compared to NH4+ Pseudomonas aeruginosa
methylamine 28% activation compared to NH4+ Pseudomonas aeruginosa
trimethylamine 5% activation compared to NH4+ Pseudomonas aeruginosa

Cloned(Commentary)

Cloned (Comment) Organism
gene ppx, recombinant expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strains XL10-Gold and BL21-CodonPlus Pseudomonas aeruginosa

Protein Variants

Protein Variants Comment Organism
additional information construction of a truncated enzyme mutants comprising amino acid residues 1-314 of 506 (N-paPpx(1-314)), or residues 1-303 (N-paPpx(1-303)), or residues 315-506 (C-paPpx(315-506)) of paPpx, amplified from Pseudomonas aeruginosa wild-type strain PAO1 chromosomal DNA through PCR. Only paPpx(1-506) and N-paPpx(1-314) are enzymatically active, while C-paPpx(315-506) lacks enzymatic activity Pseudomonas aeruginosa

KM Value [mM]

KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
additional information
-
additional information kinetic analysis of recombinant wild-type and truncated mutant enzymes, polyphosphatase activity (EC 3.6.1.11) and polyphosphate:ADP phosphotransferase activity, overview Pseudomonas aeruginosa
0.00249
-
polyphosphate65 pH 8.0, 37°C, full-length enzyme Pseudomonas aeruginosa
0.00311
-
polyphosphate25 pH 8.0, 37°C, full-length enzyme Pseudomonas aeruginosa
0.00311
-
polyphosphate65 pH 8.0, 37°C, truncated enzyme Pseudomonas aeruginosa
0.00336
-
polyphosphate25 pH 8.0, 37°C, truncated enzyme Pseudomonas aeruginosa

Metals/Ions

Metals/Ions Comment Organism Structure
Cs+ activates wild-type, full-length enzyme paPpx(1-506) Pseudomonas aeruginosa
K+ a nonessential activator of paPpx, presence of K+ does not affect the affinity of the enzyme for Mg2+, activates wild-type, full-length enzyme paPpx(1-506). The activity curve obtained with K+ is sigmoid and reaches its maximum activity at concentrations of 80 mM. Km(app)K+ is 42 mM Pseudomonas aeruginosa
Mg2+ required, values of Km(app)Mg2+ in paPpx(1-506) and NpaPpx(1-314) are 0.30 mM and 0.28 mM, respectively. The interaction between paPpx(1-506) and Mg2+ occurs in the N-terminal domain Pseudomonas aeruginosa
additional information behavior of the full-length paPpx(1-506) and N-paPpx(1-314) against different concentration of divalent ions such as Mg2+, Zn2+, Ca2+, and Mn2+ as effectors, in presence of a saturating concentration (0.008 mM) for the substrate polyphosphate65. The activation of both enzyme variants by Mg2+ is similar and shows no inhibition at high concentrations of this ion. The activation by Ca2+ and Mn2+ is negligible. Li+ and Na+ have no effects on enzyme activity, while NH4+, K+, Rb+, and Cs+ are activators of paPpx(1-506). Tetramethylammonium is not an activator of paPpx(1-506) Pseudomonas aeruginosa
NH4+ activates wild-type, full-length enzyme paPpx(1-506). The activity curve obtained with NH4+ is sigmoid and reaches its maximum activity at concentrations of 30 mM. Km(app)NH4+ is 10 mM Pseudomonas aeruginosa
Rb+ activates wild-type, full-length enzyme paPpx(1-506) Pseudomonas aeruginosa
Zn2+ Zn2+ is able to activate the enzyme only 20% compared to Mg2+ Pseudomonas aeruginosa

Natural Substrates/ Products (Substrates)

Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
ADP + (phosphate)n Pseudomonas aeruginosa
-
ATP + (phosphate)n-1
-
?

Organism

Organism UniProt Comment Textmining
Pseudomonas aeruginosa
-
-
-

Purification (Commentary)

Purification (Comment) Organism
recombinant His6-tagged wild-type and mutant enzymes from Escherichia coli strain BL21-CodonPlus by nickel affinity chromatography, dialysis, tag cleavage through thrombin, and again dialysis Pseudomonas aeruginosa

Specific Activity [micromol/min/mg]

Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
15
-
recombinant truncated enzyme, pH 8.0, 37°C, ATP formation, in presence of 80 mM K+ Pseudomonas aeruginosa
20
-
recombinant full-length enzyme, pH 8.0, 37°C, ATP formation, in presence of 80 mM K+ Pseudomonas aeruginosa
26.7
-
recombinant truncated enzyme, pH 8.0, 37°C, ATP formation, in presence of 25 mM NH4+ Pseudomonas aeruginosa
35
-
recombinant full-length enzyme, pH 8.0, 37°C, ATP formation, in presence of 25 mM NH4+ Pseudomonas aeruginosa
645
-
recombinant truncated enzyme, pH 8.0, 37°C, ATP formation, in presence of 5 mM Mg2+ and 80 mM K+ Pseudomonas aeruginosa
655
-
recombinant truncated enzyme, pH 8.0, 37°C, ATP formation, in presence of 5 mM Mg2+ Pseudomonas aeruginosa
715
-
recombinant truncated enzyme, pH 8.0, 37°C, ATP formation, in presence of 5 mM Mg2+ and 25 mM NH4+ Pseudomonas aeruginosa
3126.7
-
recombinant full-length enzyme, pH 8.0, 37°C, ATP formation, in presence of 5 mM Mg2+ and 80 mM K+ Pseudomonas aeruginosa
3150
-
recombinant full-length enzyme, pH 8.0, 37°C, ATP formation, in presence of 5 mM Mg2+ and 25 mM NH4+ Pseudomonas aeruginosa
3223.3
-
recombinant full-length enzyme, pH 8.0, 37°C, ATP formation, in presence of 5 mM Mg2+ Pseudomonas aeruginosa

Substrates and Products (Substrate)

Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
ADP + (phosphate)n
-
Pseudomonas aeruginosa ATP + (phosphate)n-1
-
?
ADP + polyphosphate25
-
Pseudomonas aeruginosa ATP + polyphosphate24
-
?
ADP + polyphosphate65
-
Pseudomonas aeruginosa ATP + polyphosphate64
-
?
additional information paPpx is an exopolyphosphatase (EC 3.6.1.11), and is also a polyphosphate:ADP phosphotransferase, and the active site is the same as that one involved in the hydrolase activity Pseudomonas aeruginosa ?
-
-

Subunits

Subunits Comment Organism
? x * 56419, sequence calculation, wild-type, full-length enzyme Pseudomonas aeruginosa

Synonyms

Synonyms Comment Organism
More cf. EC 3.6.1.11 Pseudomonas aeruginosa
paPpx
-
Pseudomonas aeruginosa
polyphosphate: ADP phosphotransferase
-
Pseudomonas aeruginosa
polyphosphate:ADP phosphotransferase
-
Pseudomonas aeruginosa

Temperature Optimum [°C]

Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
37
-
assay at Pseudomonas aeruginosa

Turnover Number [1/s]

Turnover Number Minimum [1/s] Turnover Number Maximum [1/s] Substrate Comment Organism Structure
0.29
-
polyphosphate65 pH 8.0, 37°C, truncated enzyme Pseudomonas aeruginosa
0.31
-
polyphosphate25 pH 8.0, 37°C, truncated enzyme Pseudomonas aeruginosa
3.93
-
polyphosphate65 pH 8.0, 37°C, full-length enzyme Pseudomonas aeruginosa
4.28
-
polyphosphate25 pH 8.0, 37°C, full-length enzyme Pseudomonas aeruginosa

pH Optimum

pH Optimum Minimum pH Optimum Maximum Comment Organism
8
-
assay at Pseudomonas aeruginosa

General Information

General Information Comment Organism
additional information homology structural model of full-length paPpx, in closed conformation, and of the N-terminal domain of paPpx in an open state, constructed by comparative modeling, molecular dynamic simulations, overview. Docking study with bound metals and/or ADP defining the N-paPpx(1-314) model in open conformation as receptor, docking with polyphosphate and ADP. Enzyme electrostatic potential calculations. A model of the paPpx N-terminal domain in complex with a polyP chain of 7 residues long and a molecule of ADP explains the phosphotransferase activity through docking techniques, overview Pseudomonas aeruginosa

kcat/KM [mM/s]

kcat/KM Value [1/mMs-1] kcat/KM Value Maximum [1/mMs-1] Substrate Comment Organism Structure
92.3
-
polyphosphate25 pH 8.0, 37°C, truncated enzyme Pseudomonas aeruginosa
93.3
-
polyphosphate65 pH 8.0, 37°C, truncated enzyme Pseudomonas aeruginosa
1376.2
-
polyphosphate25 pH 8.0, 37°C, full-length enzyme Pseudomonas aeruginosa
1578.3
-
polyphosphate65 pH 8.0, 37°C, full-length enzyme Pseudomonas aeruginosa