All ETDs from UAB

Advisory Committee Chair

Jamil S Saad

Advisory Committee Members

Terje Dokland

Aaron Lucius

Chad Petit

Peter E Prevelige Jr

Document Type


Date of Award


Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine


Growth and survival signals initiated at the cell surface by receptor-ligand interactions often involve a downstream activation of Akt, a serine/threonine kinase critical in regulating cell survival, apoptosis and oncogenesis. Hyper-activation of Akt is a common tumorigenic event, thereby making the Akt activation pathway a prime target for cancer therapy. The translocation of Akt to the plasma membrane (PM) for phosphorylation is a critical step in the Akt activation pathway. It has been established that membrane binding of Akt is mediated by direct interactions between its pleckstrin homology domain (PHD) and membrane lipid phosphatidylinositol 3,4,5-trisphosphate [PI(3,4,5)P3]. There is now emerging evidence that in addition to binding to PI(3,4,5)P3, the PH domain also binds to calmodulin (CaM) to promote Akt activation in many breast cancer cells. Upon epidermal growth factor (EGF) stimulation of these cells, CaM co-localizes with Akt at the PM to mediate Akt membrane anchoring and activation. However, the molecular mechanism of the interplay between lipid and CaM binding to Akt(PHD) is not known. In this dissertation, we employed nuclear magnetic resonance (NMR), biochemical and biophysical techniques to characterize how CaM and membrane lipids bind to Akt(PHD). We show that CaM binds tightly to Akt(PHD). The CaM binding interface in Akt(PHD) was mapped to two loops adjacent to the PI(3,4,5)P3 binding site, which represents a novel CaM binding motif. Next, using a membrane mimetic (nanodisc) incorporating native PI(3,4,5)P3, we demonstrate that Akt(PHD) binds much tighter to membrane than to CaM. We also show that the ternary complex formation between Akt(PHD), membrane and CaM is not cooperative and membrane lipids compete with CaM to displace it from Akt(PHD). Collectively, we provide the first mechanistic details in support of a model by which CaM binds to Akt to facilitate its translocation to the membrane. Our findings have potential to further the search for drugs to regulate the aberrant Akt activation pathway.



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