# Visualize alphafold2 predicted structure with PYMOL

- In the previous slide we plotted our MSA alignment, the pLDDT scores, and the predicted alignment error. However, it is also useful to visualize the actual predicted protein structure and compare it to the known structure if there is one. Here we use a software called PyMOL to do just that:

![](img/pymol-overview.png)

- Here we see that PyMOL takes either the PDB ID or a PDB file and creates a visualization for us to examine. If you have not done so already please [download PyMOL](https://access.tufts.edu/pymol) and open the app. You should see a window like the following:

![](img/pymol-session.png)

- Here we have a:

  - **History Window** with log of previous commands
  - **Command Interface** to enter PyMOL commands
  - **List of Objects Loaded** which list of objects/proteins that have been loaded into PyMOL
  - **Visualization Window** to visualize protiens loaded into PyMOL

- Let's try on our data!

## Download AlphaFold Output

- First we will need to download our predicted structure pdb file for the mutant, Cas12a mut2-CWF. To this, go to `Files > Home Directory`:

![](img/home-dir.png)

- Now, click on `cas12a_af2_sp24` and download the following file, then click `5XUS_mut2cwf_modified_prerun`, and download `ranked_0.pdb`:

![](img/ondemand-screenshot-ranked0.png)

## Importing Structures

- To visualize this protein structure in PyMOL, open PyMOL on your computer
- Go to File > Open - then choose your pdb file, `ranked_0.pdb`
- The file we have loaded is the Cas12a-CWF mutant, let's fetch the structure for the wild type Cas12a protein:

```bash
fetch 5xus
```

- We will now align this structure with the Cas12a-CWF mutant. So in the PyMOL command prompt enter:

```bash
align ranked_0, 5xus
```

## Viewing Structures

- To view one structure at a time, you can use the `disable` command to hide one of the structures:

```bash
disable 5xus
```

- To see this structure again we can simply use the `enable` command:

```bash
enable 5xus
```

## Visualizing Variants

- Now that we have aligned/colored our structures, let's select residues on the RuvC Domain on the Cas12a mut2-CWF and the Cas12a wild-type:

```bash
select resi 863+952+965+1214 and name CA
```

> [!NOTE]\
> Note we are only selecting the alpha carbons so that when we label these residues we only have one label per residue

- To label these residues we can use the following:

```bash
label sele, " %s%s" % (resn,resi)
```

- With these residues selected we can color them to visualize them easier:

```bash
color red, sele
```

- Let's now zoom into this region:

```bash
zoom sele
```

- Use your mouse to drag and rotate the structure to take a look at different angles of it. Locate V863, can you see the difference between wild type protein structure `5XUS` and our predicted structure `ranked_0`?
  ![](img/pymol-v863.png)

- To capture this image we can go to `File > Export Image As > PNG... > Save PNG image as ...` and enter a name for your image!

- Alternatively, you can take a screen shot and save.

- Reference Paper:
  [Ma et al. 2022](https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9825149/)

> [!NOTE]
> If you encounter any issues or this doesn't work as expected, please feel free to reach out to Shirley Li, xue.li37@tufts.edu