HINT 2.30 Manual: Chapter 5I

LESSON 4: Calculating the Hydropathic Structure of a "Key"

This lesson continues with the HIV1 molecule as used in Lesson 3. Delete the grid and contour objects created in Lesson 3 by using the Delete Object command. If you are entering the Hint Tutorial at this point, follow the instructions in Steps 1 and 2 of Lesson 1 and Steps 1, 2 and 3 of Lesson 3.

  1. Set Complement HintMap parameters for the active site of HIV-1

    For this map you want to define the region of the active site as the grid dimensions. The easiest way to do this now is to read in the known inhibitor, and use it as a template for the Molecule Region. Go to the Molecule pulldown and select the Get command. Choose PDB as the Get File Type and turn Heteroatoms on. Enter ./a74704.pdb as the File Name; turn Reference Object on and select HIV1 as the Object Name; press Execute. We will return to this molecule in Step 3, but for now, pulldown Setup (on the Hint menu bar) to the Grid command. Select Grid Center as Molecule Region and choose A74704 as the Molecule_Region. The box defined by this molecule should coincide to the region of the active site of HIV-1.

    From the Molecule pulldown, select the Display command and turn the Display of A74704 off for now.

    From the Setup pulldown, select Grid Complement. Most of this is identical to the Grid Molecule command discussed in Lessons 2 and 3. We will set most parameters identical to the choices of Lesson 3. From Grid_Types select Hydrophobic/Polar; enter a value of 6.0 for the CutOff Radius; set the Volume_Averaging parameter to off; and set the Van der Waals Limit to 1.00.. Press Execute to set the Grid Complement parameters.

    Next, set the Hint_DistFunct parameters. For a HintGrid Complement calculation select exp(-nr) for the Hydropathic Term and set the Steric_Term to off. In this case we will add directionality to the polar portion of the Hint Complement map by setting Direction Vectors to Hybridizd/Lone Pair and using a Vector Focus of 0.5. This will emphasize the higher electron density and polarity of lone pairs and pi orbitals (i.e., directionality) to give what we believe is a more accurate view of the hydrogen bonding and interaction ability of these atoms. Press Execute to set these parameters.

  2. Calculate the HintGrid Complementary maps

    Go to the HintGrid pulldown and select the Complementary command. Parameters in this block initiate the Complementary HintGrid calculation. First make sure that HIV1 is the parameter in the Molecule parameter box. Next, choose Background as the Execute_Mode parameter. The Job Name and Grid Filename parameters will be chosen for you automatically if you touch these boxes and press return. Enter a textual Comment of your choice. Select Auto_Get_Grid to be on. Finally, press Execute to begin the map calculation. This map calculation should take 15-30 minutes.

    Repeat the map calculation to create a parallel Acid/Base Complement map for HIV1. Return to Grid_Complement Setup to select Acid/Base from Grid_Types. Choose Lewis as the Acid/Base Definition to use the Lewis model for acids and bases. Make sure all other parameters are the same as before. Press Execute. Return to Complement HintGrid and repeat the calculation; new default values for Job Name and Grid Filename will be chosen.

    When this second map calculation is complete (about 15-30 minutes) you may contour the HintGrid maps for display. Go to the Grid pulldown on the Hint menu bar and select the Contour command. First select the Grid Object box and pick HIV1_CO_HP from the associated Value Aid. Next select the Contour Name Root box and enter HIVCH. This will be the root portion of all Contour Objects created from HIV_CO_HP. For the Contour Select parameter pick the Single Value option. Contour the Hydrophobic portion of this map. Enter a value of 150.0 for the Contour Level and pick a green for the Contour Color parameter. Now select the Grid Object box and pick HIV1_CO_AB from the associated Value Aid. Next select the Contour Name Root box and enter HIVCP. Contour this map with -50.0 (red) and 50.0 (blue). You probably will have to use the "Slab" to decrease the display depth-of-field to an interpretable level. This display shows regions of the active site that would best be fit with a hydrophobic "key" (in green) and regions that would best be matched with Lewis acid (red) or Lewis base (blue) polar groups in the inhibitor molecule.

  3. Examine hydropathic structure of an inhibitor

    One known inhibitor for HIV protease is the A74704 molecule designed, synthesized and analyzed by the Erickson group at Abbott and at the National Cancer Institute. As you recall, we read in this molecule in step 1 of this Lesson in order to use its coordinates to define the active site. Now you will see how well it fits the predictions made by Hint regarding substrate design. Go to the Molecule pulldown and select the Display command to redisplay A74704. Color the molecule by atom. You should note how the two leucine-like group of A74704 are enveloped by the hydrophobic green contours of the Complement map, and how many of the polar features of the inhibitor are spatially near analogous regions in the "key" map for HIV. In particular note that the inhibitor hydroxyl group is in an acid-like (red) contour.

    Inhibitor A74704 and HINT complement map of HIV-1 Protease