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Literature summary extracted from

  • Chiang, P.; Sampaleanu, L.M.; Ayers, M.; Pahuta, M.; Howell, P.L.; Burrows, L.L.
    Functional role of conserved residues in the characteristic secretion NTPase motifs of the Pseudomonas aeruginosa type IV pilus motor proteins PilB, PilT and PilU (2008), Microbiology, 154, 114-126.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
3.6.1.15 genes pilB, pilT and pilU, DNA and amino acid sequence determination and anaylsis, expression of N-terminally His6-tagged wild-type and mutant enzymes in Escherichia coli strain Bl21 Pseudomonas aeruginosa
7.4.2.8 genes pilB, pilT and pilU from strain PAK, expression of wild-type and mutant enzymes in Escherichia coli strain Bl21(DE3). Expression of YFP-PilT, YFP-PilB, and YFP-PilU fusion enzymes, which retain their characteristic pattern of polar localization, with the exception of the Walker A mutant of YFP-PilT Pseudomonas aeruginosa

Protein Variants

EC Number Protein Variants Comment Organism
3.6.1.15 D160N site-directed mutagenesis, mutation of the Asp Box motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 E159Q site-directed mutagenesis, mutation of the Asp Box motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 E163Q site-directed mutagenesis, mutation of the Asp Box motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 E204Q site-directed mutagenesis, mutation of the Walker B motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 G135S site-directed mutagenesis, mutation of the Walker A motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 H222A site-directed mutagenesis, mutation of the His Box motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 H229A site-directed mutagenesis, mutation of the His Box motif residue in PilT Pseudomonas aeruginosa
3.6.1.15 additional information mutation of conserved Walker A or Walker B residues in any of the three ATPases abrogates twitching motility, and for the Walker A mutant of PilT causes loss of polar localization, construction of diverse mutants of the type IV pilus motor proteins, overview Pseudomonas aeruginosa
7.4.2.8 D160N mutation in the Walker A motif of PilT, and in PilU Pseudomonas aeruginosa
7.4.2.8 D355N mutation of PilB Pseudomonas aeruginosa
7.4.2.8 E159Q mutation in the Walker A motif of PilT, and in PilU Pseudomonas aeruginosa
7.4.2.8 E163Q mutation in the Walker A motif of PilT, and in PilU Pseudomonas aeruginosa
7.4.2.8 E204 mutation in the Walker B motif of PilT, and in PilU, inactive PilT mutant Pseudomonas aeruginosa
7.4.2.8 E354Q mutation of PilB Pseudomonas aeruginosa
7.4.2.8 E358Q mutation of PilB Pseudomonas aeruginosa
7.4.2.8 E396Q mutation of PilB Pseudomonas aeruginosa
7.4.2.8 G135S mutation in the Asp Box of PilT, and in PilU, inactive PilT mutant Pseudomonas aeruginosa
7.4.2.8 G331S mutation of PilB Pseudomonas aeruginosa
7.4.2.8 H222A mutation in the His Box of PilT, and in PilU Pseudomonas aeruginosa
7.4.2.8 H229A mutation in the His Box of PilT, and in PilU Pseudomonas aeruginosa
7.4.2.8 H414A mutation of PilB Pseudomonas aeruginosa
7.4.2.8 H421A mutation of PilB Pseudomonas aeruginosa
7.4.2.8 additional information a constructed YFP-PilB construct does not complement a pilB mutant, mutation of conserved Walker A or Walker B residues in any of PilB, PilT and PilU ATPases abrogates twitching motility, and for the Walker A mutant of PilT causes loss of polar localization, overview Pseudomonas aeruginosa

Localization

EC Number Localization Comment Organism GeneOntology No. Textmining

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
3.6.1.15 Mg2+
-
 Pseudomonas aeruginosa
7.4.2.8 Mg2+ required Pseudomonas aeruginosa

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
7.4.2.8 ATP + H2O Pseudomonas aeruginosa
-
 ADP + phosphate
-
 ?
7.4.2.8 ATP + H2O Pseudomonas aeruginosa PAK
-
 ADP + phosphate
-
 ?
7.4.2.8 additional information Pseudomonas aeruginosa PilB and PilT are bipolar proteins belonging to the secretion NTPase superfamily, and power pilus extension and retraction, respectively, while the unipolar PilT paralogue PilU supports pilus retraction in an unknown manner. In all three proteins, the third acidic residue in the Asp Box and the second His of the His Box are crucial for function, mutation of these residues causes loss of PilT ATPase activity in vitro ?
-
 ?
7.4.2.8 additional information Pseudomonas aeruginosa PAK PilB and PilT are bipolar proteins belonging to the secretion NTPase superfamily, and power pilus extension and retraction, respectively, while the unipolar PilT paralogue PilU supports pilus retraction in an unknown manner. In all three proteins, the third acidic residue in the Asp Box and the second His of the His Box are crucial for function, mutation of these residues causes loss of PilT ATPase activity in vitro ?
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
3.6.1.15 Pseudomonas aeruginosa
-
 genes pilB, pilT and pilU
-
3.6.1.15 Pseudomonas aeruginosa PAK
-
 genes pilB, pilT and pilU
-
7.4.2.8 Pseudomonas aeruginosa
-
 genes pilB, pilT and pilU
-
7.4.2.8 Pseudomonas aeruginosa PAK
-
 genes pilB, pilT and pilU
-

Purification (Commentary)

EC Number Purification (Comment) Organism
3.6.1.15 recombinant His6-tagged wild-type and mutant enzymes from Escherichia coli strain Bl21 by nickel affinity chromatography Pseudomonas aeruginosa

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
3.6.1.15 additional information
-
 activity of mutant enzymes, overview Pseudomonas aeruginosa
3.6.1.15 0.00008
-
 PilU Pseudomonas aeruginosa
3.6.1.15 0.00012
-
 PilT Pseudomonas aeruginosa
3.6.1.15 0.00018
-
 PilB Pseudomonas aeruginosa
7.4.2.8 0.00108
-
 recombinant PilT mutant H229A Pseudomonas aeruginosa
7.4.2.8 0.00275
-
 recombinant PilT mutant E163Q Pseudomonas aeruginosa
7.4.2.8 0.005
-
 recombinant wild-type PilU Pseudomonas aeruginosa
7.4.2.8 0.0074
-
 recombinant wild-type PilT Pseudomonas aeruginosa
7.4.2.8 0.0108
-
 recombinant wild-type PilB Pseudomonas aeruginosa

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
3.6.1.15 ATP + H2O recombinant His-tagged PilB, PilT, and PilU show ATPase activity in vitro with requirement for three invariant acidic residues in the Asp Box motif, and for two invariant His residues in the His Box motif to varying extents, overview Pseudomonas aeruginosa ADP + phosphate
-
 ?
3.6.1.15 ATP + H2O recombinant His-tagged PilB, PilT, and PilU show ATPase activity in vitro with requirement for three invariant acidic residues in the Asp Box motif, and for two invariant His residues in the His Box motif to varying extents, overview Pseudomonas aeruginosa PAK ADP + phosphate
-
 ?
3.6.1.15 NTP + H2O
-
 Pseudomonas aeruginosa NDP + phosphate
-
 ?
3.6.1.15 NTP + H2O
-
 Pseudomonas aeruginosa PAK NDP + phosphate
-
 ?
7.4.2.8 ATP + H2O
-
 Pseudomonas aeruginosa ADP + phosphate
-
 ?
7.4.2.8 ATP + H2O the enzyme shows requirement for three invariant acidic residues in the Walker Asp Box motif, and for two invariant His residues in the His Box motif, structure-function analysis and modelling, overview. The Walker A motif is involved in ATP binding, while the Walker B motif is involved in the hydrolysis of ATP Pseudomonas aeruginosa ADP + phosphate
-
 ?
7.4.2.8 ATP + H2O
-
 Pseudomonas aeruginosa PAK ADP + phosphate
-
 ?
7.4.2.8 ATP + H2O the enzyme shows requirement for three invariant acidic residues in the Walker Asp Box motif, and for two invariant His residues in the His Box motif, structure-function analysis and modelling, overview. The Walker A motif is involved in ATP binding, while the Walker B motif is involved in the hydrolysis of ATP Pseudomonas aeruginosa PAK ADP + phosphate
-
 ?
7.4.2.8 additional information PilB and PilT are bipolar proteins belonging to the secretion NTPase superfamily, and power pilus extension and retraction, respectively, while the unipolar PilT paralogue PilU supports pilus retraction in an unknown manner. In all three proteins, the third acidic residue in the Asp Box and the second His of the His Box are crucial for function, mutation of these residues causes loss of PilT ATPase activity in vitro Pseudomonas aeruginosa ?
-
 ?
7.4.2.8 additional information PilB and PilT are bipolar proteins belonging to the secretion NTPase superfamily, and power pilus extension and retraction, respectively, while the unipolar PilT paralogue PilU supports pilus retraction in an unknown manner. In all three proteins, the third acidic residue in the Asp Box and the second His of the His Box are crucial for function, mutation of these residues causes loss of PilT ATPase activity in vitro Pseudomonas aeruginosa PAK ?
-
 ?

Subunits

EC Number Subunits Comment Organism
3.6.1.15 More the type IV pilus motor proteins contain conserved residues in the Walker A, Walker B, and Asp Box and His Box motifs characteristic of secretion NTPases, overview Pseudomonas aeruginosa
7.4.2.8 homohexamer secretion NTPases of the type II secretion and type IV pili systems, including PilB, PilT and PilU, function as toroidal homohexamers and, in addition toWalker A and Walker B motifs, contain unique Asp Box and His Box motifs Pseudomonas aeruginosa

Synonyms

EC Number Synonyms Comment Organism
3.6.1.15 More PilB and PilT are bipolar proteins belonging to the secretion NTPase superfamily Pseudomonas aeruginosa
3.6.1.15 PilB
-
 Pseudomonas aeruginosa
3.6.1.15 PilT
-
 Pseudomonas aeruginosa
3.6.1.15 PilU
-
 Pseudomonas aeruginosa
3.6.1.15 T4P motor protein
-
 Pseudomonas aeruginosa
3.6.1.15 type IV pilus motor protein
-
 Pseudomonas aeruginosa
7.4.2.8 More PilB and PilT are bipolar proteins belonging to the secretion NTPase superfamily Pseudomonas aeruginosa
7.4.2.8 PilB
-
 Pseudomonas aeruginosa
7.4.2.8 PilT
-
 Pseudomonas aeruginosa

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
3.6.1.15 24
-
 assay at Pseudomonas aeruginosa
7.4.2.8 24
-
 assay at Pseudomonas aeruginosa

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
3.6.1.15 7.5
-
 assay at Pseudomonas aeruginosa
7.4.2.8 7.5
-
 assay at Pseudomonas aeruginosa