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iCn3D Tutorial: Protein Kinase B (AKT)

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    Protein Kinase B (AKT)

    A more complicated protein with multiple domains and at least two major conformations with different PDB structures: Protein Kinase B (aka AKT)

    PDB files:

    • 3CQW (active)
    • 3O96  (inactive)
    • 1UNQ  PH domain

    A.  Exploration of AKT – Domain Structure

    • PDB ID: 1UNQ
    • Description: Pleckstrin Homology Domain Of Protein Kinase B/Akt Bound To Ins(1,3,4,5)-Tetrakisphophate
    1. Open new iCn3D
    2. File, Retrieve by ID, MMDB
    3. Alt Click the ligand.  Style, Chemicals, Sphere
    4. Select, Select on 3D, check Chain, alt click the protein
    5. Color,  Secondary, Sheets in Yellow

    With the protein still selected:

    1. Analysis, Interactions
    2. For the second set choose chemicals
    3. Check just the noncovalent interactions shown below

      clipboard_e86b03ae938dbc70e2c3e0fc82f4120de.png
       
    4. Choose 3D Display Interactions.  This will display interacting side chains
    5. Selections, Save Selections, name it Interactions
    6. Style, Sidechains, Stick (so next step will just color side chains)
    7. Color, atom
    8. In defined sets choose Interactions.  Analysis, Label, Per Residue & No.
    9. Analysis, Label Scale, 3
    10. Style, Background, White
    11. File, Share Link, copy short link

    B.  Exploration of AKT – Self-defined aa, sequences, Render PH-domain, N-lobe, C-Lobe, Catalytic Loop, Activation Loop

    PDB ID: 3O96 (inactive)

    Description: 

    • Crystal Structure of Human AKT1 with an Allosteric Inhibitor
    • RAC-alpha serine/threonine-protein kinase; Chain A
    • Allosteric inhibitor IQO

    Cartoon structure

    clipboard_ed0412281c36fa1a7152b6d9d3529a4fa.png

    Instructions

    1. File, Retrieve by ID, MMDB
    2. Analysis, Seq. & Annotations, Uncheck Conserved Domains
    3. Analysis, Defined Sets.  Click 1st domain (PH = top 3O96_A_3d_domain)
    4. Style, Protein, C-Alpha Trace.
    5. Color , Unicolor, Yellow
    6. Defined Sets.  Choose 2st domain (N-Lobe).  Color, unicolor, Cyan
    7. Defined Sets.  Choose 3st domain (C-Lobe).  Color, unicolor, Grey, Light Gray
    8. Seq. & Annotations, Details  Tab, Uncheck Conserved Domains.
    9. Select by hand the catalytic  loop  (table below) by using the horizontal scroll to find and select 271V-287H.  Hover over each amino acids and click to select it or use your mouse to sweep out a rectangle over all the range of amino acids.
    10. Select, Save Selection, CatLoop
    11. Color, Unicolor, Red
    12. Repeat for Activation Loop and save the selection. Color Unicolor, Blue
    13. Style, Background, White
    14. File, Share Link, Lifelong Short Link

    Numbering or generic kinase and AKT

    SITE

    Generic
    Kinase

    Akt (+108)

    N lobe

    K72

    K180

    N lobe

    E91

    E199

    C lobe, cat loop

    R165

    R273

    C lobe, cat loop

    D166

    D274

    C lobe, act loop

    D184

    D292

    C lobe, act loop

     

    T197

    T305
    (NO SEQ)

    Approx Cat Loop

    163-179

    271V—287H

     

    Approx Act loop

    Start DFG (292-294) to APE

    184-200

    292-319

    308Tmiss toAPE end 319

     

    C.  Compare 3CQW (active) and 3O96  (inactive) – Superimpose two AKT structures

    PDB ID: 3CQW and 3O96

    Description

    3O96 (inactive): Crystal Structure of Human AKT1 with an Allosteric Inhibitor

    • RAC-alpha serine/threonine-protein kinase; Chain A
    • Allosteric inhibitor IQO

    3CQW (active):

    • Crystal Structure of Akt-1 complexed with substrate peptide (hence active form) and inhibitor
    • Peptide:  Glycogen synthase kinase-3 beta peptide

    Instructions

    1. File, Align, Structure to Structure
    2. Input 3CQW and 3O96
    3. Choose All Matching Molecules Superimposed
    4. In the top of the right hand alignment window, save the aligned state as presented (al_seq1).  Just the aligned sequences will be highlighted.
    5. View,  View Selection
    6. Style, Background, White
    7. File, Share Link, Lifelong Short Link
    8. Toggle between the two states by clicking the toggle menu button just underneath the File menu button

    iCn3D Tutorial: Protein Kinase B (AKT) is shared under a not declared license and was authored, remixed, and/or curated by LibreTexts.