Structures of the PKC-ι kinase domain in its ATP-bound and apo forms reveal defined structures of residues 533–551 in the C-terminal tail and their roles in ATP binding

Takimura, Tetsuo; Kamata, Kenji; Fukasawa, Kazuhiro; Ohsawa, Hirokazu; Komatani, Hideya; Yoshizumi, Takashi; Takahashi, Ikuko; Kotani, Hidehito; Iwasawa, Yoshikazu

doi:10.1107/s0907444910005639

Cited by 38 publications

(41 citation statements)

References 18 publications

(20 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…7, C and D). These interactions are similar to those reported in previous studies of the crystal structure of ATPbound PKA (PDB code 1ATP) (23,24,29,30), PKC (PDB code 3A8W) (22), and PKC␤ (PDB code 3PFQ) (31), suggesting the accuracy of our PKC␦ homology models. ATP interacted with the same residues in WT and AS PKC␦, but the electrostatic interaction with the gatekeeper residue (Met-427) in WT PKC␦ was replaced by a weaker van der Waals interaction with Ala-427 in AS PKC␦.…”

Section: -(Benzyl)-atp Fits In the Nucleotide-binding Pocket Of As Pksupporting

confidence: 88%

See 1 more Smart Citation

Generation and Characterization of ATP Analog-specific Protein Kinase Cδ

Kumar

Weng

Geldenhuys

et al. 2015

Journal of Biological Chemistry

View full text Add to dashboard Cite

Background: A traceable form of PKC␦ is needed to investigate PKC␦ functions. Results: Analog-specific PKC␦ could utilize N 6 -(benzyl)-ATP to phosphorylate PKC␦ substrates and was specifically inhibited by PP1 analogs. Conclusion: Binding to the catalytic domain glycine-rich loop, Lys-378, Glu-428, Leu-430, and Phe-633 is the likely mechanism by which N 6 -(benzyl)-ATP and PP1 analogs interact with analog-specific PKC␦. Significance: This study provides tools to investigate PKC␦-mediated pathways.

show abstract

Section: -(Benzyl)-atp Fits In the Nucleotide-binding Pocket Of As Pksupporting

confidence: 88%

“…Because the x-ray crystal structure of PKC␦ is not available, PKC␦ homology models were created based on the crystal structure of human PKC (PDB code 3A8W) (22). The root mean square deviation for the overlaid structure of WT and AS PKC␦ is 0.279 Å (Fig.…”

Section: -(Benzyl)-atp Fits In the Nucleotide-binding Pocket Of As Pkmentioning

confidence: 99%

Generation and Characterization of ATP Analog-specific Protein Kinase Cδ

Kumar

Weng

Geldenhuys

et al. 2015

Journal of Biological Chemistry

View full text Add to dashboard Cite

show abstract

“…3A, graph). This protection from dephosphorylation was exaggerated in cells overexpressing a PKC ␤II construct containing a destabilizing mutation within the highly conserved NFD motif of the AGC protein kinase family, corresponding to F629A in PKC ␤II (33)(34)(35). This construct was more readily down-regulated by PDBu when compared with wild-type enzyme (Fig.…”

Section: Resultsmentioning

confidence: 94%

Active Site Inhibitors Protect Protein Kinase C from Dephosphorylation and Stabilize Its Mature Form

Gould

Antal

Reyes

et al. 2011

Journal of Biological Chemistry

View full text Add to dashboard Cite

Conformational changes acutely control protein kinase C (PKC). We have previously shown that the autoinhibitory pseudosubstrate must be removed from the active site in order for 1) PKC to be phosphorylated by its upstream kinase phosphoinositide-dependent kinase 1 (PDK-1), 2) the mature enzyme to bind and phosphorylate substrates, and 3) the mature enzyme to be dephosphorylated by phosphatases. Here we show an additional level of conformational control; binding of active site inhibitors locks PKC in a conformation in which the priming phosphorylation sites are resistant to dephosphorylation. Using homogeneously pure PKC, we show that the active site inhibitor Gö 6983 prevents the dephosphorylation by pure protein phosphatase 1 (PP1) or the hydrophobic motif phosphatase, pleckstrin homology domain leucine-rich repeat protein phosphatase (PHLPP). Consistent with results using pure proteins, treatment of cells with the competitive inhibitors Gö 6983 or bisindolylmaleimide I, but not the uncompetitive inhibitor bisindolylmaleimide IV, prevents the dephosphorylation and down-regulation of PKC induced by phorbol esters. Pulse-chase analyses reveal that active site inhibitors do not affect the net rate of priming phosphorylations of PKC; rather, they inhibit the dephosphorylation triggered by phorbol esters. These data provide a molecular explanation for the recent studies showing that active site inhibitors stabilize the phosphorylation state of protein kinases B/Akt and C.PKC isozymes comprise a family of multidomain proteins that are under exquisite conformational control. Two major mechanisms control the conformation of PKC family members: phosphorylation and second messenger-dependent membrane binding (1, 2). First, newly synthesized enzymes undergo a series of ordered phosphorylations that lock the enzyme into a stable, catalytically competent, and autoinhibited species (3, 4). This species is maintained in an autoinhibited conformation by a pseudosubstrate segment that blocks the substrate-binding cavity, a conformation that also protects the priming sites of PKC from dephosphorylation. The inactive species are localized throughout the cell, often tethered to scaffold proteins (5). This processing by phosphorylation is constitutive and required to protect PKC from degradation; unphosphorylated protein is rapidly degraded (1). Second, binding to lipid second messengers allosterically controls the enzyme by facilitating the release of the autoinhibitory pseudosubstrate segment from the substrate-binding cavity (6). Thus, this conformational transition is acutely controlled by activation of receptors that signal using diacylglycerol as the second messenger. The membranebound conformation has an increased sensitivity to phosphatases by 2 orders of magnitude (7), and prolonged activation, as occurs with phorbol esters (functional analogues of diacylglycerol), results in the dephosphorylation and subsequent degradation of PKC (8). Thus, phosphorylation converts newly synthesized PKC into a stable, degradation-resistan...

show abstract

“…Inspection of this structure revealed an interesting lattice contact between adjacent pairs of PKCi kinase domain molecules [ Fig. 1A: Protein Data Bank (PDB) code 3ZH8] not seen in previous PKCi crystal structures (23,38,39). Residues 469 to 485 and 510 to 514 from the C lobe of one kinase molecule (Fig.…”

Section: Structural Analysis Reveals An Invariant Dibasic Ripr Motif mentioning

confidence: 91%

A Cancer-Associated Mutation in Atypical Protein Kinase Cι Occurs in a Substrate-Specific Recruitment Motif

et al. 2013

View full text Add to dashboard Cite

Atypical protein kinase Cι (PKCι) has roles in cell growth, cellular polarity, and migration, and its abundance is frequently increased in cancer. We identified a protein interaction surface containing a dibasic motif (RIPR) that bound a distinct subset of PKCι substrates including lethal giant larvae 2 (LLGL2) and myosin X, but not other substrates such as Par3. Further characterization demonstrated that Arg471 in this motif was important for binding to LLGL2, whereas Arg474 was critical for interaction with myosin X, indicating that multiple complexes could be formed through this motif. A somatic mutation of the dibasic motif (R471C) was the most frequent mutation of PKCι in human cancer, and the intact dibasic motif was required for normal polarized epithelial morphogenesis in three-dimensional cysts. Thus, the R471C substitution is a change-of-function mutation acting at this substrate-specific recruitment site to selectively disrupt the polarizing activity of PKCι.

show abstract

Structures of the PKC-ι kinase domain in its ATP-bound and apo forms reveal defined structures of residues 533–551 in the C-terminal tail and their roles in ATP binding

Cited by 38 publications

References 18 publications

Generation and Characterization of ATP Analog-specific Protein Kinase Cδ

Generation and Characterization of ATP Analog-specific Protein Kinase Cδ

Active Site Inhibitors Protect Protein Kinase C from Dephosphorylation and Stabilize Its Mature Form

A Cancer-Associated Mutation in Atypical Protein Kinase Cι Occurs in a Substrate-Specific Recruitment Motif

Contact Info

Product

Resources

About