Spring 2018 Protein Structure Assignment

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  Chem 153A: Fall 2017 Protein structure assignment. Turn in to Gradescope by April 18, 2018; 11:59pm Name: Sunjin Kim UID: 404605409 Lec: 2 Part A (10 pts): The Protein Databank: Go to the PDB website www.rcsb.org  Look at the Molecule of the Month article, for April 2018 . Choose one of the pdb structures linked in the article  and answer the following questions: 1. (1 pt) What is the PDB code for the structure? 5m2g 2. (1 pt) How many unique polypeptide chains are in this structure? 1 3. (1 pt) What is the amino acid length of each unique polypeptide? 464 4. (1 pt) What is the name of each polypeptide? Tetrachloroethene reductive dehalogenase catalytically active subunit 5. (1 pt) How many total polypeptide chains are in this structure? 2 6. (1 pt) What are the chains labeled? A,B 7. (1 pt) What ligands or other molecules were in this structure? Iron/sulfur cluster, norpseudo-B12, 2,4,6-Tribromophenol, glycerol, benzamidine 8. (1 pt) What technique was used to determine this structure? X-ray diffraction Search the protein databank for the protein featured in the Molecule of the Month article and answer the following questions: Enter your EXACT  search term here: : Dehalogenases 9. (1 pt) How many structures of this protein have been published? __________341_______________ 10. (1 pt) How many papers have been published containing structures of this protein (# of citations)?  _________________105 citations_________________  Part B (15 pts): General protein structural features: Using pymol open the pdb file for Alcohol Dehydrogenase, 1D1S and answer the following questions about that structure: 1. (1 pt) How many polypeptide chains are in this structure?__________4________________________ To visualize the features easier follow these instructions: Hide everything for the structure Type “select chain A ” into the command line  (make sure the A is capitalized) For the selection show cartoon, you may also want to color differently to see the features better. ANSWER THE FOLLOWING QUESTIONS FOR CHAIN A ONLY , BASED ON WHAT YOU SEE IN PYMOL . 2. (1 pt) What is the amino acid at the C-terminus of Chain A? Phenylalanine 3. (1 pt) How many α -helices are in Chain A? 12 4. (2 pts) How many amino acids make up the longest α -helix? 19 5. (1 pt) How many β -strands are in this structure?_18 6. (1 pts) Find the largest β -sheet in the structure. What type of beta sheet is it? Parallel beta sheet 7. (1 pt) This beta sheet is part of a Rossman fold (an NAD binding domain) that includes amino acids 163-338. Visualize the NAD in the structure. Which end of the Beta sheet does the NAD bind to? The C-terminus end 8. (2 pts) The Rossman fold is characterized by a beta sheet sandwiched between two sets of alpha helices. Look at the pattern of secondary structures in the overall fold. Looking just at amino acids 190-240 may help you see the pattern. Briefly describe how the fold is made up. The fold starts with a beta sheet, turns to form an alpha helix, turns back to another beta sheet, turns again to an alpha helix, and the final part is another beta sheet. The three beta strands form a parallel beta sheet. 9. (2 pts) Visualize the Gly173, Gly175, Gly 320, and Gly321. What do you notice about the protein structure around these amino acids? Propose a brief explanation for what you see. Glycine is shown to be in positions where the alpha helix bends. This interruption in the normal alpha helix bend may be due to glycine being the smallest amino acid, creating a disruption in the interactions between the sidechains of other amino acids in the helix.  10. (3 Pts) Insert a picture of the view in pymol you used to answer question #9. The answer to Question 9 should be clearly visible. Hint: Rotate around the structure so that you can see all of chain A (and ONLY chain A) along with the glycines you visualized in question 9. Make sure the glycines are clearly visible, not just labeled. Save a copy of the image and insert it here:  Part C (5 pts): Visualizing the effect of ligand-protein interactions In this section you are going to investigate two forms of pyruvate dehydrogenase (PDH) Kinase, an enzyme that we will study in more detail later in the quarter. PDH Kinase is inhibited by pyruvate. Below you will investigate two structures of PDH kinase, one with a pyruvate analog bound (2BU8) and one without (2BU2). PDH Kinase transfers a phosphate group from ATP to pyruvate dehydrogenase. In the active site the backbone carbonyl of G317 and F318 can interact with either the ATP (substrate) or ADP (product). Open structure 2BU8: Visualize G317, F318 and the ADP bound to the structure. Using the measurement tool determine the distances between the oxygen of the backbone carbonyl of each amino acid and the closest oxygen to it attached to the β -phosphate (units are Angstroms): 1.   (1 pt) Between G317 and ADP: 3.9 Angstroms 2.   (1 pt) Between F318 and ADP: 2.8 Angstroms Open structure 2BU2: Visualize G317, F318 and the ATP bound to the structure. Using the measurement tool determine the distances between the oxygen of the backbone carbonyl of each amino acid and the closest oxygen to it attached to the β -phosphate (units are Angstroms): 3.   (1 pt) Between G317 and ATP: 6.4 Angstroms 4.   (1 pt) Between F318 and ATP: 7.4 Angstroms 5.   (1 pt) Other than the distances what is the difference between these amino acids in the different structures? The difference between these amino acids is that in 2BU2, it binds to ATP and in 2BU8, it binds to ADP. Extra Credit (2 pts): Visualize the pyruvate analog TF4 in the 2BU8 structure. Propose a mechanism for how binding of pyruvate may lead to the differences you saw between the two structures, and how this can contribute to changes in activity of PDH kinase. TF4 is the binding site for the pyruvate, and the binding of the pyruvate will phosphorylate the ADP into an ATP since the pyruvate has a phosphate group. ADP and pyruvate, which are part of 2BU8, is an inhibitor for the PDH. However, since 2BU2 has ATP, it may serve as an activator for PDH.
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