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Literature summary extracted from

  • Sweetman, D.; Stavridou, I.; Johnson, S.; Green, P.; Caddick, S.E.; Brearley, C.A.
    Arabidopsis thaliana inositol 1,3,4-trisphosphate 5/6-kinase 4 (AtITPK4) is an outlier to a family of ATP-grasp fold proteins from Arabidopsis (2007), FEBS Lett., 581, 4165-4171.
    View publication on PubMed

Cloned(Commentary)

EC Number Cloned (Comment) Organism
2.7.1.159 expression in Escherichia coli Arabidopsis thaliana
2.7.1.159 in Escherichia coli Rosetta, transformed with pET28a and pET28c containing 3 kinase cDNAs. Vector-encoded hexa-histidine (His) tags are added to all proteins. Arabidopsis thaliana

KM Value [mM]

EC Number KM Value [mM] KM Value Maximum [mM] Substrate Comment Organism Structure
2.7.1.159 0.042
-
 1D-myo-inositol 1,3,4,6-tetrakisphosphate
-
 Arabidopsis thaliana

Metals/Ions

EC Number Metals/Ions Comment Organism Structure
2.7.1.159 LiCl 10 mM in activity assay Arabidopsis thaliana
2.7.1.159 MgCl2 6 mM in activity assay Arabidopsis thaliana

Natural Substrates/ Products (Substrates)

EC Number Natural Substrates Organism Comment (Nat. Sub.) Natural Products Comment (Nat. Pro.) Rev. Reac.
2.7.1.159 1D-myo-inositol-1,3,4,5-tetrakisphosphate + ATP Arabidopsis thaliana enzyme displays isomerase activity 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ATP
-
 ?
2.7.1.159 1D-myo-inositol-1,3,4-trisphosphate + ATP Arabidopsis thaliana
-
 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ADP
-
 ?
2.7.1.159 1D-myo-inositol-1,4,6-trisphosphate + 1D-myo-inositol-3,4,6-trisphosphate + 2 ATP Arabidopsis thaliana racemic mixture 2 1D-myo-inositol-1,3,4,6-tetrakisphosphate + 2 ADP
-
 ?
2.7.1.159 1D-myo-inositol-3,4,5,6-tetrakisphosphate + ATP Arabidopsis thaliana enzyme displays isomerase activity 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ATP
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,3,4-trisphosphate Arabidopsis thaliana
-
 ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,3,4-trisphosphate Arabidopsis thaliana
-
 ADP + 1D-myo-inositol 1,3,4,6-tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,3,4-trisphosphate Arabidopsis thaliana The substrate is converted to InsP4 but not to InsP5 ADP + 1D-myo-inositol 1,3,4,6-tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,4,6-trisphosphate + 1D-myo-inositol 3,4,6-trisphosphate Arabidopsis thaliana racemic mix of phosphates ADP + inositol tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 3,4,5,6-tetrakisphosphate Arabidopsis thaliana
-
 ADP + 1D-myo-inositol 1,3,4,5,6-pentakisphosphate
-
 ?
2.7.1.159 additional information Arabidopsis thaliana AtITPK4 differs from other family members in that it not display inositol 3,4,5,6-tetrakisphosphate 1-kinase activity. It displays inositol 1,4,5,6-tetrakisphosphate and inositol-1,3,4,5-tetrakisphosphate isomerase activity. With inositol tetrakisphosphate substrates, ITPK4 displays an isomerase activity (in the absence of inositol pentakisphosphate product) interpreting as phosphatase and kinase activity. AtITPK4 shows isomerase activity against other substrates. Removal of the 1- or 3-phosphate of inositol-1,3,4,6-tetrakisphosphate and subsequent rephosphorylation at the same position generates inositol-1,3,4,6-tetrakisphosphate. ?
-
 ?
2.7.1.159 additional information Arabidopsis thaliana enzyme displays both kinase and phosphatase activity. ?
-
 ?

Organism

EC Number Organism UniProt Comment Textmining
2.7.1.159 Arabidopsis thaliana O80568
-
-
2.7.1.159 Arabidopsis thaliana O80568 AtITPK4 protein is distinguished from the other proteins by an extended N-terminal sequence (39 residues longer than that of AtITPK2). A blast search with amino acids 1-80 of AtITPK4 as query fails to identify significant homology to any proteins of other known functional annotation. In alignment, AtITPK4 has an extra 121 amino acids N-terminal to the start of the Entamoeba enzyme. AtITPK4 shows 14.9% identity and 25.7% similarity to the Entamoeba enzyme in a pairwise alignment.
-
2.7.1.159 Arabidopsis thaliana Q9SBA5 enzyme belongs to a larger family of ATP-grasp fold proteins
-
2.7.1.159 Arabidopsis thaliana O81893 enzyme belongs to a larger family of ATP-grasp fold proteins
-
2.7.1.159 Arabidopsis thaliana Q9SUG3 enzyme belongs to a larger family of ATP-grasp fold proteins
-

Purification (Commentary)

EC Number Purification (Comment) Organism
2.7.1.159 Cells are pelleted and resuspended in buffer, sonicated, centrifuged and the supernatant of the lysate is retained. Ni2+-nitrilotriacetate resin prewashed in NiSO4 is added to the lysate and incubated. The resin is washed twice with 10 ml of 50 mM NaH2PO4, 300 mM NaCl and 20 mM imidazole (pH 8) to elute non-specifically bound proteins, and washed with of NaH2PO4, 300 mM NaCl and 250 mM imidazole to elute AtIPK1 protein. Bacterial lysates and protein fractions from the purification are analysed by SDS-PAGE. Protein concentration is determined by the Bradford assay. Arabidopsis thaliana
2.7.1.159 Cells are pelleted and resuspended in buffer, sonicated, centrifuged and the supernatant of the lysate is retained. Ni2+-nitrilotriacetate resin prewashed in NiSO4 is added to the lysate and incubated. The resin is washed twice with 10 ml of 50 mM NaH2PO4, 300 mM NaCl and 20 mM imidazole (pH 8) to elute non-specifically bound proteins, and washed with of NaH2PO4, 300 mM NaCl and 250 mM imidazole to elute AtIPK1 protein. Bacterial lysates and protein fractions from the purification are analysed by SDS-PAGE. Protein concentration is determined by the Bradford method. Arabidopsis thaliana
2.7.1.159 Cells are pelleted and resuspended in buffer, sonicated, centrifuged and the supernatant of the lysate is retained. Ni2+-nitrilotriacetate resin prewashed in NiSO4 is added to the lysate and incubated. The resin is washed twice with 10 ml of 50 mM NaH2PO4, 300 mM NaCl and 20 mM imidazole (pH 8) to elute non-specifically bound proteins, and washed with of NaH2PO4, 300 mM NaCl and 250 mM imidazole to elute AtIPK1 protein. Bacterial lysates and protein fractions from the purification are analysed by SDS/PAGE. Protein concentration is determined by Bradford assay. Arabidopsis thaliana
2.7.1.159 Cells are pelleted and resuspended in buffer, sonicated, centrifuged and the supernatant of the lysate is retained. Ni2+-nitrilotriacetate resin prewashed in NiSO4 is added to the lysate and incubated. The resin is washed twice with 10 ml of 50 mM NaH2PO4, 300 mM NaCl and 20 mM imidazole (pH 8) to elute non-specifically bound proteins, and washed with of NaH2PO4, 300 mM NaCl and 250 mM imidazole to elute AtIPK1 protein. Bacterial lysates and protein fractions from the purification are analysed by SDS/PAGE. Protein concentration is determined by the Bradford method. Arabidopsis thaliana

Source Tissue

EC Number Source Tissue Comment Organism Textmining
2.7.1.159 anther
-
 Arabidopsis thaliana
-
2.7.1.159 carpel
-
 Arabidopsis thaliana
-
2.7.1.159 cauline leaf
-
 Arabidopsis thaliana
-
2.7.1.159 cauline leaf weak expression Arabidopsis thaliana
-
2.7.1.159 flower expressed in male and female organs of young and mature flowers Arabidopsis thaliana
-
2.7.1.159 flower
-
 Arabidopsis thaliana
-
2.7.1.159 flower bud young, AtITPK4 transcript is expressed throughout early flower development which culminates in the opening of the flower bud Arabidopsis thaliana
-
2.7.1.159 leaf
-
 Arabidopsis thaliana
-
2.7.1.159 leaf weak expression, vasculature Arabidopsis thaliana
-
2.7.1.159 petal
-
 Arabidopsis thaliana
-
2.7.1.159 pollen
-
 Arabidopsis thaliana
-
2.7.1.159 root
-
 Arabidopsis thaliana
-
2.7.1.159 sepal
-
 Arabidopsis thaliana
-
2.7.1.159 silique
-
 Arabidopsis thaliana
-
2.7.1.159 silique weak expression, revealed expression in the seed and pod wall Arabidopsis thaliana
-
2.7.1.159 stamen
-
 Arabidopsis thaliana
-
2.7.1.159 stem
-
 Arabidopsis thaliana
-

Specific Activity [micromol/min/mg]

EC Number Specific Activity Minimum [µmol/min/mg] Specific Activity Maximum [µmol/min/mg] Comment Organism
2.7.1.159 additional information
-
 not detected in mature flowers, but expression of trancript is confirmed by mRNA in situ analysis Arabidopsis thaliana
2.7.1.159 0.0044
-
-
 Arabidopsis thaliana

Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
2.7.1.159 1D-myo-inositol-1,3,4,5-tetrakisphosphate + ATP enzyme displays isomerase activity Arabidopsis thaliana 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ATP
-
 ?
2.7.1.159 1D-myo-inositol-1,3,4-trisphosphate + ATP
-
 Arabidopsis thaliana 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ADP
-
 ?
2.7.1.159 1D-myo-inositol-1,4,5,6-tetrakisphosphate + ATP AtITPK4 is more active against 1D-myo-inositol-1,4,5,6-tetrakisphosphate than against 1D-myo-inositol-3,4,5,6-tetrakisphosphate. Arabidopsis thaliana 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ATP
-
 ?
2.7.1.159 1D-myo-inositol-1,4,6-trisphosphate + 1D-myo-inositol-3,4,6-trisphosphate + 2 ATP racemic mixture Arabidopsis thaliana 2 1D-myo-inositol-1,3,4,6-tetrakisphosphate + 2 ADP
-
 ?
2.7.1.159 1D-myo-inositol-3,4,5,6-tetrakisphosphate + ATP enzyme displays isomerase activity Arabidopsis thaliana 1D-myo-inositol-1,3,4,6-tetrakisphosphate + ATP
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,3,4-trisphosphate
-
 Arabidopsis thaliana ADP + 1D-myo-inositol 1,3,4,5-tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,3,4-trisphosphate
-
 Arabidopsis thaliana ADP + 1D-myo-inositol 1,3,4,6-tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,3,4-trisphosphate The substrate is converted to InsP4 but not to InsP5 Arabidopsis thaliana ADP + 1D-myo-inositol 1,3,4,6-tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 1,4,6-trisphosphate + 1D-myo-inositol 3,4,6-trisphosphate racemic mix of phosphates Arabidopsis thaliana ADP + inositol tetrakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol 3,4,5,6-tetrakisphosphate
-
 Arabidopsis thaliana ADP + 1D-myo-inositol 1,3,4,5,6-pentakisphosphate
-
 ?
2.7.1.159 ATP + 1D-myo-inositol-1,3,4-trisphosphate
-
 Arabidopsis thaliana ?
-
 ?
2.7.1.159 additional information the enzyme displays inositol 1,4,5,6-tetrakisphosphate and inositol 1,3,4,5-tetrakisphosphate isomerase activity. It lacks the Ins(3,4,5,6)P4 1-kinase Arabidopsis thaliana ?
-
 ?
2.7.1.159 additional information AtITPK4 differs from other family members in that it not display inositol 3,4,5,6-tetrakisphosphate 1-kinase activity. It displays inositol 1,4,5,6-tetrakisphosphate and inositol-1,3,4,5-tetrakisphosphate isomerase activity. With inositol tetrakisphosphate substrates, ITPK4 displays an isomerase activity (in the absence of inositol pentakisphosphate product) interpreting as phosphatase and kinase activity. AtITPK4 shows isomerase activity against other substrates. Removal of the 1- or 3-phosphate of inositol-1,3,4,6-tetrakisphosphate and subsequent rephosphorylation at the same position generates inositol-1,3,4,6-tetrakisphosphate. Arabidopsis thaliana ?
-
 ?
2.7.1.159 additional information enzyme displays both kinase and phosphatase activity. Arabidopsis thaliana ?
-
 ?

Synonyms

EC Number Synonyms Comment Organism
2.7.1.159 At2G43980
-
 Arabidopsis thaliana
2.7.1.159 AtITPK1
-
 Arabidopsis thaliana
2.7.1.159 AtITPK2
-
 Arabidopsis thaliana
2.7.1.159 AtITPK3
-
 Arabidopsis thaliana
2.7.1.159 AtITPK4
-
 Arabidopsis thaliana
2.7.1.159 inositol 1,3,4-trisphosphate 5/6-kinase
-
 Arabidopsis thaliana
2.7.1.159 inositol 1,3,4-trisphosphate 5/6-kinase 4
-
 Arabidopsis thaliana

Temperature Optimum [°C]

EC Number Temperature Optimum [°C] Temperature Optimum Maximum [°C] Comment Organism
2.7.1.159 30
-
 assay at Arabidopsis thaliana

pH Optimum

EC Number pH Optimum Minimum pH Optimum Maximum Comment Organism
2.7.1.159 7.5
-
 assay at Arabidopsis thaliana

Cofactor

EC Number Cofactor Comment Organism Structure
2.7.1.159 ATP
-
 Arabidopsis thaliana
2.7.1.159 ATP commentary Arabidopsis thaliana