Prediction of Secondary Structure

1. Now that we have seen how the wildtype and mutant prion sequences differ from one another, let's look to see how these differences affect folding of the resulting proteins. Scroll back up to the top of the screen and click "Return".

2. This will return you to the Alignment Tools homepage. Click on the "Protein Tools" button at the top to bring you to the Protein Tools homepage.



3. 1. Check the boxes next to both sequences. Then, scroll down the menu and highlight the tool that says "GOR4 ¡V Predict Secondary Structure of PS". This tool predicts the amount of alpha helix and beta sheet (in other words, secondary structure) present in a protein sequence. Click "Run".



4. We do not need to concern ourselves with the next screen. Simply click "Submit". The new screen that appears shows the results. You can see right away that the wildtype sequence (the one on the top) has both alpha helices (shown in red letters) and beta sheets (blue letters).



5. The mutant sequence (the one on the bottom) has ONLY beta sheets (shown in blue letters). The mutant sequence contains NO alpha helices.



But, let's be careful here. Remember that the mutant sequence is much shorter than the wildtype sequence so we must compare only the segments where the mutant sequence and the wildtype sequence overlap. Let's look at the part of the wildtype sequence that overlaps the mutant sequence. In the figure below, there is a highlighted region that shows the region where the mutant sequence overlaps the wildtype sequence. This is so we can see how many alpha helices (red) and beta sheets (blue) there are in the region where the wildtype sequence overlaps with the mutant sequence. Mark off with a pencil where the wildtype sequence overlaps the mutant sequence by comparing the following two figures:



This is the region in the wildtype sequence that overlaps the mutant sequence. We need to figure out how many alpha helices and beta sheets are in this highlighted region so we can compare the number to the mutant sequence. Since we can't see the colors because the region is highlighted, compare it to the figure below.



Now mark with your pencil on this wildtype sequence where it overlaps with the mutant sequence by referring to the highlighted section in the figure above.

After careful comparison, you will see that in the region of overlap the wildtype sequence contains 5 beta sheets and 1 alpha helix. On the other hand, the mutant sequence contains 9 beta sheets and NO alpha helices. This constitutes a huge difference in secondary structure which would undoubtedly affect protein folding, which is presumably responsible for the change from a functional prion protein to a disease-causing prion protein.