Structural and Biophysical Characterization of the Interactions between the Death Domain of Fas Receptor and Calmodulin

Fernandez, Timothy F.; Samal, Alexandra B.; Bedwell, Gregory J.; Chen, Yabing; Saad, Jamil S.

doi:10.1074/jbc.m113.471821

Cited by 20 publications

(27 citation statements)

References 64 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…CaM samples were stored in a buffer containing 50 mM HEPES or Tris (pH 7.0), 150 mM NaCl, and 1 mM CaCl 2 . The CaM-N domain (residues 1-80) was expressed and purified as described (59). Because CaM-N has zero extinction coefficient, protein concentration was measured using the bicinchoninic acid assay (BCA; Thermo Scientific).…”

Section: Methodsmentioning

confidence: 99%

The Interplay between Calmodulin and Membrane Interactions with the Pleckstrin Homology Domain of Akt

Agamasu¹,

Ghanam²,

et al. 2017

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

The Akt protein, a serine/threonine kinase, plays important roles in cell survival, apoptosis, and oncogenes. Akt is translocated to the plasma membrane for activation. Akt-membrane binding is mediated by direct interactions between its pleckstrin homology domain (PHD) and phosphatidylinositol 3,4,5-trisphosphate (PI(3,4,5)P). It has been shown that Akt activation in breast cancer cells is modulated by calmodulin (CaM). However, the molecular mechanism of the interplay between CaM and membrane binding is not established. Here, we employed nuclear magnetic resonance (NMR) and biochemical and biophysical techniques to characterize how PI(3,4,5)P, CaM, and membrane mimetics (nanodisc) bind to Akt(PHD). We show that PI(3,4,5)P binding to Akt(PHD) displaces the C-terminal lobe of CaM but not the weakly binding N-terminal lobe. However, binding of a PI(3,4,5)P-embedded membrane nanodisc to Akt(PHD) with a 10-fold tighter affinity than PI(3,4,5)P is able to completely displace CaM. We also show that Akt(PHD) binds to both layers of the nanodisc, indicating proper incorporation of PI(3,4,5)P on the nanodisc surface. No detectable binding has been observed between Akt(PHD) and PI(3,4,5)P-free nanodiscs, demonstrating that PI(3,4,5)P is required for membrane binding, CaM displacement, and Akt activation. Using pancreatic cancer cells, we demonstrate that inhibition of Akt-CaM binding attenuated Akt activation. Our findings support a model by which CaM binds to Akt to facilitate its translocation to the membrane. Elucidation of the molecular details of the interplay between membrane and CaM binding to Akt may help in the development of potential targets to control the pathophysiological processes of cell survival.

show abstract

Section: Methodsmentioning

confidence: 99%

The Interplay between Calmodulin and Membrane Interactions with the Pleckstrin Homology Domain of Akt

Agamasu¹,

Ghanam²,

et al. 2017

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

show abstract

“…CaM samples were stored in a buffer containing 50 mM HEPES or Tris at pH 7.0, 150 mM NaCl, and 5 mM CaCl 2 . CaM-N (residues 1-80) and CaM-C (residues 76 -148) proteins were expressed and purified as described (36). CaM-N protein concentrations were measured using a bicinchoninic acid assay (BCA; Thermo Scientific).…”

Section: Methodsmentioning

confidence: 99%

Structural and Biophysical Characterization of the Interactions between Calmodulin and the Pleckstrin Homology Domain of Akt

Agamasu

Ghanam

Saad

2015

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

“…In addition, CCAs are frequently resistant to TRAIL-mediated apoptosis 307,308 . Interaction of Fas cell surface death receptor with calmodulin leads to the inhibition of Fas-induced apoptosis and might be involved in CCA chemoresistance 309 . Moreover, mutations in the pro-apoptotic tumour suppressor TP53 have been suggested as predictors of CCA outcome 209 .…”

Section: Mechanisms Of Chemoresistancementioning

confidence: 99%

Cholangiocarcinoma: current knowledge and future perspectives consensus statement from the European Network for the Study of Cholangiocarcinoma (ENS-CCA)

Bañales

Cardinale

Carpino

et al. 2016

Nat Rev Gastroenterol Hepatol

1,068

1,139

View full text Add to dashboard Cite

Structural and Biophysical Characterization of the Interactions between the Death Domain of Fas Receptor and Calmodulin

Cited by 20 publications

References 64 publications

The Interplay between Calmodulin and Membrane Interactions with the Pleckstrin Homology Domain of Akt

The Interplay between Calmodulin and Membrane Interactions with the Pleckstrin Homology Domain of Akt

Structural and Biophysical Characterization of the Interactions between Calmodulin and the Pleckstrin Homology Domain of Akt

Cholangiocarcinoma: current knowledge and future perspectives consensus statement from the European Network for the Study of Cholangiocarcinoma (ENS-CCA)

Contact Info

Product

Resources

About