c-Jun Homodimer: Leucine Zipper Protein [PDB:1JUN]

The c-Jun protein is a transcriptional activator that binds DNA as either a homodimer or a heterodimer with the c-Fos protein [1, 2]. The three-dimensional structure of the leucine zipper domain of the c-Jun homodimer has been determined by nuclear magnetic resonance imaging [3, 4], and can be loaded by selecting Open from the File menu of the ProteinShader program and then using the file chooser box to select the 1JUN.pdb file.

Cartoon and space filling style displays

When the c-Jun leucine zipper domain is first loaded, it will be displayed as a pen-and-ink like drawing of ribbons (general loop regions) and tubes (alpha-helices) as shown in Figure 1A below. The domain is composed of two identical parallel chains, A and B, that are primarily alpha-helical in structure. To switch to a display style that uses spheres to represent atoms (Figure 1B), go to the Style menu above the canvas and use the Atom submenu to select Space Filling. The default size of each sphere is determined by the van der Waals radius for the atom type, which can be found by going to the ProteinShader API and clicking on AtomEnum. The default color is determined by the CPK (Corey, Pauling, and Koltun) color scheme for atoms, which can be found by going to the ProteinShader API and clicking on CPKColorEnum.

With colors based on atom type, it is very difficult to distinguish the two chains in a space filling style display, so in Figure 1B chain A is shown in red and chain B in green. To set these colors, go to the menu at the top right of the control panel and select Atom Color. Near the top left of the control panel, go to the Chain menu and select B. Choosing B should cause the menu right below the Selected radio button to show Chain as the current selection to apply modifications to. Clicking on the Chooser button will open a color chooser panel, which can now be used to select green for chain B. Leaving the color chooser panel open, go back to the Chain menu and select A so that red can be applied to chain A. Click on the OK button of the color chooser panel to close it.

To get a better idea of how the chains wrap around each other, either drag the mouse horizontally across the canvas, or select Motion from the menu at the top right of the control panel and hit the Start button. The text box or slider control below the Y-Axis label can be used to adjust the speed of rotation. The motion panel sets speed in degrees per second so that the rotation speed should be the same on any machine, but how smooth or jerky the motion is will depend on how powerful the graphics card is. After the image has been moved around, the original view can always be restored by going to the Orientation menu above the canvas and selecting Original.

A [1JUN.pdb pen-and-ink] B [1JUN.pdb space filling]

Figure 1. The leucine zipper domain of the c-Jun transcription factor is a coiled coil of parallel alpha-helices. (A) The two parallel chains are shown as ribbons and tubes using halftoning to produce a pen-and-ink style drawing. (B) The same structure is shown as a space filling style display with chain A in red and chain B in green. The images were captured as described in the Saving Images section of the tutorials main page.

Visualizing the leucine zipper

Like other members of the bZIP family of transcription factors, the c-Jun homodimer is stabilized by hydrophobic interactions between leucine residues that repeat at regular intervals along each chain [2, 3]. To visualize these leucine residues, first go to the Style menu above the canvas and select Tubes from the Cartoon submenu. The loops will be shown with a smaller diameter than the alpha-helix tubes. Now go to the menu at the top right of the control panel and select Cartoon Side Chains. Near the middle of the Cartoon Side Chains subpanel is a radio button labeled Global. Go to the menu right below Global and choose LEU. Now click the Space Filling button further below. The image should be that same as is in Figure 2A below, with the leucine side chains shown in red for chain A and green for chain B.

To get a better look at how the leucines pair up, rotate the image a few times by dragging the mouse horizontally across the canvas. It may also be helpful to temporarily set one of the chains to invisible as in Figure 2B, so go to the menu at the top right of the canvas and select Cartoon Visibility. Select either A or B from the Chain menu near the top left of the control panel. Make sure that the menu right below the Selected radio button is set to Chain, and then hit the Invisble button. With only a single chain visible as in Figure 2B, it should be easy to observe that the leucine residues are spaced every two turns of the alpha-helix, which corresponds to every seventh residue.

To better observe the pairing between the hydrophobic side chains of the leucine residues, the tubes used to represent the backbone of the alpha-helices can be set to be partially transparent as in Figure 2C. Go to the menu right below the Selected radio button to choose Model, and then click on the Translucent button near the bottom of the Cartoon Visibility subpanel. The translucent effect works a little better with a light background, so go to the Background menu above the canvas and select Light Gray. The slider near the bottom of the Cartoon Visibility panel can be used to fade the translucent tubes in and out. The side chains are not affected because they are an atom-style display, so the Atom Visibility subpanel would affect their translucency, not the Cartoon Visibility subpanel.

A [1JUN.pdb tubes & side chains] B [1JUN.pdb tubes & side chains] C [1JUN.pdb translucent tubes]

Figure 2. Illustration of the leucine residues of the leucine zipper domain of the c-Jun protein. (A) The two identical chains of the c-Jun homodimer are shown as tubes, while the leucine side chains are shown as a space filling display. The leucine residues for chain A are shown in red, while the leucine residues for chain B are shown in green. (B) Chain A has been set to invisible so that the leucine residues of chain B can be seen more clearly. Leucine residues occur every two turns of the alpha-helix (every 7 residues). (C) Same display as in (A), except that the tubes are shown using 75% translucency to make it easier to see the pairing of the hydrophobic leucine residues. The images were captured as described in the Saving Images section of the tutorials main page.

Destabilizing the leucine zipper

In addition to the leucines that form a zipper-like structure, a particularly interesting residue discussed by the authors of the c-Jun NMR paper is an asparagine at position 291 of each chain [3]. This polar residue is located at the dimer interface, and provides a destabilizing influence that is likely to facilitate the rapid exchange of zipper strands in vivo (c-Jun/c-Fos heterodimers have a higher DNA binding affinity than c-Jun homodimers, and control of dimer formation is part of how this family of transcription factors is regulated).

To visualize this residue, first go to the list of amino acids shown on the left side of the control panel and select ASN 291. Then use the menu at the top right of the control panel to go back to the Cartoon Side Chain subpanel. Make sure that Residues is showing in the menu below the Selected radio button, and then click on the Space Filling button. To change the color of the aspargine side chain, change the menu at the top right of the control panel to Atom color and click the Atom Type button. Setting the color by atom type will use gray for carbon, white for hydrogen, blue for nitrogen, and red for oxygen.

The manipulations described right above will affect only the ASN 291 of whichever chain (A or B) is currently showing in the Chain menu near the top left of the control panel, so the process will need to be repeated after flipping the Chain menu to the other chain. Using the Chain menu will cause the menu below the Selected radio button to automatically flip to Chain, so it will be necessary to change that menu back to Residues before clicking the Space Filling button on the Cartoon Side Chains subpanel or the Atom Type button on the Atom Color subpanel. Once ASN 291 has been set on both chains, the image should look the same as in Figure 3A below. To get a better look at the asparagine residues, the mouse motion controls can be used to zoom in on the image as shown in Figure 3B.

A [1JUN.pdb pen-and-ink tubes] B [1JUN.pdb space filling]

Figure 3. A polar asparagine residue has a destabilizing influence on the leucine zipper domain of the c-Jun homodimer. (A) The protein is shown as a tubes style display (75% transparency) with leucine side chains (red for chain A and green for chain B) as a space filling style display. The side chain for asparagine 291 of each chain is also shown, but colored by atom type (gray for carbon, white for hydrogen, blue for nitrogen, and red for oxygen). (B) The same display as in (A) after zooming in on the aspargine residues and rotating the image. The images were captured as described in the Saving Images section of the tutorials main page.

Multiple models

As a final point for this tutorial using c-Jun, there are actually seven slightly different models for the leucine zipper domain because the structure was determined using NMR (nuclear magnetic resonance imaging), which only provides a set of distance constraints for hydrogen atoms [5]. The ProteinShader program loads all of the models given in a Protein Data Bank file, and the Model menu at the top left of the control panel can be used to select any of the models. Currently, the program makes no attempt to synchronize changes to the models, but that option might be added at a later time. Right now, when a structure is first loaded, the Decorations subpanel is used to automatically apply Halftoning with texture maps (to get a pen-and-ink style effect) to only the first model. To apply the same effect to other models, the Halftoning button and its associated texture menus should be used. Any modifications to one model are remembered by only that one model.

References

1. Brandon C, Tooze J: Leucine zippers provide dimerization interactions for some eucaryotic transcription factors. In Introduction to Protein Structure. 2nd edition. New York: Garland Publishing; 1998: 191-193.

2. Angel PE, Herrlich PA (Eds): The FOS and JUN Families of Transcription Factors. Boca Raton: CRC Press; 1994.

3. Junius FK, O`Donoghue SI, Nilges M, Weiss AS, King GF: High resolution NMR solution structure of the leucine zipper domain of the c-Jun homodimer. J Biol Chem 1996, 271: 13663-13667.

4. c-Jun homodimer PDB entry 1JUN [http://www.rcsb.org/pdb/explore/explore.do?structureId=1JUN]

5. Brandon C, Tooze J: Determination of Protein Structure. In Introduction to Protein Structure. 2nd edition. New York: Garland Publishing; 1998: 373-392.