Binding of phosphatidylinositol 3,4,5-trisphosphate to the pleckstrin homology domain of protein kinase B induces a conformational change.
Protein kinase B (PKB/Akt) is a key regulator of cell growth, proliferation and metabolism. It possesses an N-terminal pleckstrin homology (PH) domain that interacts with equal affinity with the second messengers PtdIns(3,4,5)P3 and PtdIns(3,4)P2, generated through insulin and growth factor-mediated activation of phosphoinositide 3-kinase (PI3K). The binding of PKB to PtdIns(3,4,5)P3/PtdIns(3,4)P2 recruits PKB from the cytosol to the plasma membrane and is also thought to induce a conformational change that converts PKB into a substrate that can be activated by the phosphoinositide-dependent kinase 1 (PDK1). In this study we describe two high-resolution crystal structures of the PH domain of PKBalpha in a noncomplexed form and compare this to a new atomic resolution (0.98 A, where 1 A=0.1 nm) structure of the PH domain of PKBalpha complexed to Ins(1,3,4,5)P4, the head group of PtdIns(3,4,5)P3. Remarkably, in contrast to all other PH domains crystallized so far, our data suggest that binding of Ins(1,3,4,5)P4 to the PH domain of PKB, induces a large conformational change. This is characterized by marked changes in certain residues making up the phosphoinositide-binding site, formation of a short a-helix in variable loop 2, and a movement of variable loop 3 away from the lipid-binding site. Solution studies with CD also provided evidence of conformational changes taking place upon binding of Ins(1,3,4,5)P4 to the PH domain of PKB. Our data provides the first structural insight into the mechanism by which the interaction of PKB with PtdIns(3,4,5)P3/PtdIns(3,4)P2 induces conformational changes that could enable PKB to be activated by PDK1.
Milburn
Christine C
CC
Division of Biological Chemistry and Molecular Microbiology, School of Life Sciences, University of Dundee, Dundee DD1 5EH, UK.
Deak
Maria
M
Kelly
Sharon M
SM
Price
Nick C
NC
Alessi
Dario R
DR
Van Aalten
Daan M F
DM
D001665:Binding Sites; D001798:Blood Proteins / Q000378:metabolism; D002942:Circular Dichroism; D018360:Crystallography, X-Ray; D007295:Inositol Phosphates / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008968:Molecular Conformation; D018129:Phosphatidylinositol Phosphates / Q000737:chemistry* / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism; D011485:Protein Binding; D011487:Protein Conformation; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D013431:Sulfates / Q000737:chemistry
D001798:Blood Proteins; D007295:Inositol Phosphates; D018129:Phosphatidylinositol Phosphates; D010750:Phosphoproteins; D011518:Proto-Oncogene Proteins; D013431:Sulfates; C060974:phosphatidylinositol 3,4,5-triphosphate; C039077:platelet protein P47; C081655:inositol-1,3,4,5-tetrakisphosphate; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt