First, we will look at the Angiotensin II Type-1 membrane receptor as compared to Angiotensin II Type-2 membrane receptor.

Opening An Account
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Go to the Biology Workbench homepage in Workbench panel. Click on the link that says, "Registration". If you already have an account, go ahead and log into the program. If not, you need to open an account first.  It is free and it is available immediately. Once your account is created, you will be taken to another website Click on the link that reads Biology Workbench to go to the main page.

Starting a New Session
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There are a few different sets of tools supplied by the Workbench. They are Session Tools, Protein Tools, Nucleic Tools, Alignment Tools and Structure Tools (Alpha). To maintain some organization in your research, you should create a new session for every new topic you research.
1. From the window highlight the line that reads Start New Session and click Run, You will be taken to another window
2. Type the name of the new session, for example, Angiotensin, in the space provided and then click the Start New Session button You will be taken to another window.



The page will reload and the name of your new session will appear below the Default Session. You can click back and forth between the default and Superkingdoms sessions. However, for the remainder of this tutorial, make sure that the Superkingdoms session is selected.

3. Because Angiotensin II Type-1 membrane receptor and Angiotensin II Type-2 membrane receptor are proteins we will use the Protein Tools. We will use the Ndjinn Multiple Database search.

4. Use the term "angiotensin II receptor" for your search and select the Swissprot Database. For Hits per Page choose ALL.

5. Select "Type 1 angiotensin II receptor" and "Type 2 angiotensin II receptor" for humans and Import Sequences.

6. Select both sequences by clicking on the boxes to the left of the names. In the window select CLUSTALW. - (perform selected operation) When new page appears go to the bottom and (submit). This aligns the sequences. Remember you are looking at amino acid sequences. Do you want to know which letter stands for which amino acid?

Question 1 What do the different colors of amino acids mean?

7. To understand the coding better you need to think about amino acid structure. The green coding refers to strong group. If you looked at the molecular formula for the 2 amino acids in common, you would see they look very much alike. The strong groups mean they have a side chain that has the same charge or are the same size and therefore act very similiar in their chemical reactions. The dark blue coding refers to a weak group. To help you Pay special attention to the amino acids in black. They represent where changes have occurred...EVOLUTION. Differences in the proteins have accumulated due to mutations of the DNA sequence. Can you find Steve's DRY motif that was so important to him?

Question 2 What does the color coding tell you about the DRY group?

8. Now back up a few pages to your search results. Choose the Type II receptor for rat, mouse and sheep. and Import Sequences.

9. Select the Type II sequences for all 4 species by clicking on the boxes to the left of the names. In the window select CLUSTALW. - (perform selected operation) When new page appears go to the bottom and (submit).

Question 3 Complete the table below based on your alignment.
(HINT: Each complete row consists of 60 amino acids) Look at each animal's protein vs the human ATII. Count the number of differences in their amino acids. (For the sheep only compare the regions that they both have.) Record that in column B. Subtract that from the total and you will have the number of amino acids conserved.

Organism		A. Number of Amino Acid Positions conserved (they were the same as the human)	B. Number of Different Amino Acid Positions Non-Conserved areas	Percentage of Conserved Amino Acids =(Column A/ total A+B amino acids *100%)
Human vs.	Rat
	Mouse
	Sheep

Question 4 Which animal should Steve use in for his research on Type II angiotensin? Why?

Question 5 Do you find the DRY motif in any or all of these animals? Which ones?

10. Scroll down to the Clustal W Dendogram. This shows how closely relately organisms are. Using the right click button, copy and paste this into your word document.

Question 6 Explain the evolutionary relationships of the 4 animals based on this single protein.