1. Macromolecular Visualization or… Where to go when ChemDraw just isn’t enough Martin Case Chem258 2009

2. Macromolecular viewing software Over 150 visualization programs exist http://molvis.sdsc.edu/visres/index.html http://molvis.sdsc.edu/visres/index.html 100 of these are free to academic users Many are supported across all common platforms: Mac OS X Windows Linux/Unix This introduction to visualization uses PyMol

3. PyMol Reads files from in Protein Data Bank format (PDB) You can convert to PDB from any other file format using a program called Babel Renders objects as lines, sticks, CPK, ribbons, surface etc Objects are easily manipulated (spin, translate) with the mouse Includes a toolkit for local protein building, energy minimization etc.

4. PyMol screenshots

5. PyMol basics: loading a PDB file The “raw data” for protein structure viewing and manipulation is the PDB file ASCII text files You can read and edit them with eg WordPad or TextEdit A header with experimental details, references etc xyz coordinates from x-ray crystallography of NMR experiments The Protein Data Bank is at http://www.rcsb.org The ALH receptor complexed with its autoinducer is 2uv0.pdb We’ll use this structure for this tutorial

6. Download the biological unit You might need to unzip/unstuff the file And you might need to change the.pdb1 to.pdb

7. PDB headers HEADER TRANSCRIPTION 08-MAR-07 XXXX TITLE STRUCTURE OF THE P. AERUGINOSA LASR LIGAND-BINDING DOMAIN TITLE 2 BOUND TO ITS AUTOINDUCER COMPND TRANSCRIPTIONAL ACTIVATOR PROTEIN LASR KEYWDS TRANSCRIPTION, ACYL-HOMOSERINE LACTONE RECEPTOR, QUORUM KEYWDS 2 SENSING, TRANSCRIPTION REGULATION, ALPHA-BETA-ALPHA KEYWDS 3 SANDWICH, TRANSCRIPTIONAL REGULATOR, ACTIVATOR, DNA-BINDING EXPDTA X-RAY DIFFRACTION AUTHOR M.J.BOTTOMLEY, E.MURAGLIA, R.BAZZO, A.CARFI JRNL AUTH M.J.BOTTOMLEY, E.MURAGLIA, R.BAZZO, A.CARFI JRNL TITL MOLECULAR INSIGHTS INTO QUORUM SENSING IN THE JRNL TITL 2 HUMAN PATHOGEN PSEUDOMONAS AERUGINOSA FROM THE JRNL TITL 3 STRUCTURE OF THE VIRULENCE REGULATOR LASR BOUND TO JRNL TITL 4 ITS AUTOINDUCER. JRNL REF J.BIOL.CHEM. V. 282 13592 2007 JRNL REFN ASTM JBCHA3 US ISSN 0021-9258 JRNL PMID 17363368 JRNL DOI 10.1074/JBC.M700556200 SEQRES 1 E 175 GLY ALA MSE ALA LEU VAL ASP GLY PHE LEU GLU LEU GLU SEQRES 2 E 175 ARG SER SER GLY LYS LEU GLU TRP SER ALA ILE LEU GLN SEQRES 3 E 175 LYS MSE ALA SER ASP LEU GLY PHE SER LYS ILE LEU PHE SEQRES 4 E 175 GLY LEU LEU PRO LYS ASP SER GLN ASP TYR GLU ASN ALA SEQRES 5 E 175 PHE ILE VAL GLY ASN TYR PRO ALA ALA TRP ARG GLU HIS

8. PDB coordinates (protein) ATOM 1 N GLY E 6 29.335 -0.940 63.260 1.00 17.84 N ATOM 2 CA GLY E 6 29.407 0.297 62.504 1.00 17.95 C ATOM 3 C GLY E 6 28.906 1.490 63.294 1.00 17.84 C ATOM 4 O GLY E 6 27.701 1.663 63.477 1.00 18.09 O ATOM 5 N PHE E 7 29.835 2.315 63.764 1.00 17.68 N ATOM 6 CA PHE E 7 30.323 3.445 62.983 1.00 17.46 C ATOM 7 C PHE E 7 29.628 3.519 61.627 1.00 17.40 C ATOM 8 O PHE E 7 28.718 4.324 61.428 1.00 17.42 O ATOM 9 CB PHE E 7 31.838 3.350 62.792 1.00 17.57 C ATOM 10 CG PHE E 7 32.478 4.642 62.369 1.00 17.61 C ATOM 11 CD1 PHE E 7 33.386 4.670 61.324 1.00 17.36 C ATOM 12 CD2 PHE E 7 32.170 5.827 63.015 1.00 17.59 C ATOM 13 CE1 PHE E 7 33.976 5.857 60.932 1.00 17.50 C ATOM 14 CE2 PHE E 7 32.757 7.017 62.627 1.00 17.62 C ATOM 15 CZ PHE E 7 33.661 7.032 61.584 1.00 17.40 C ATOM 16 N LEU E 8 30.039 2.631 60.727 1.00 17.17 N ATOM 17 CA LEU E 8 29.697 2.728 59.315 1.00 17.22 C ATOM 18 C LEU E 8 28.192 2.768 59.079 1.00 17.06 C ATOM 19 O LEU E 8 27.722 3.478 58.191 1.00 17.28 O ATOM 20 CB LEU E 8 30.322 1.572 58.532 1.00 17.34 C ATOM 21 CG LEU E 8 30.213 0.185 59.167 1.00 17.70 C ATOM 22 CD1 LEU E 8 29.138 -0.640 58.477 1.00 17.98 C ATOM 23 CD2 LEU E 8 31.553 -0.533 59.126 1.00 17.59 C

9. PDB coordinates (non-protein) HETATM 2627 C21 OHN E1169 28.316 19.063 77.652 1.00 7.59 C HETATM 2628 C20 OHN E1169 27.312 19.933 76.888 1.00 6.96 C HETATM 2629 C19 OHN E1169 26.399 19.052 76.036 1.00 6.87 C HETATM 2630 C18 OHN E1169 25.305 19.850 75.325 1.00 7.62 C HETATM 2631 C17 OHN E1169 24.113 20.065 76.244 1.00 6.73 C HETATM 2632 C16 OHN E1169 23.381 18.755 76.511 1.00 6.04 C HETATM 2633 C15 OHN E1169 22.384 18.934 77.643 1.00 4.71 C HETATM 2634 C14 OHN E1169 21.910 17.570 78.135 1.00 2.14 C HETATM 2635 C13 OHN E1169 23.043 16.837 78.847 1.00 2.00 C HETATM 2636 C11 OHN E1169 22.528 15.724 79.762 1.00 2.00 C HETATM 2637 O12 OHN E1169 21.330 15.448 79.830 1.00 2.00 O HETATM 2638 C10 OHN E1169 23.563 14.956 80.595 1.00 2.00 C HETATM 2639 C8 OHN E1169 22.908 13.701 81.181 1.00 2.00 C HETATM 2640 O9 OHN E1169 22.782 12.678 80.517 1.00 2.00 O HETATM 2641 N7 OHN E1169 22.500 13.817 82.437 1.00 2.00 N HETATM 2642 C1 OHN E1169 21.856 12.720 83.157 1.00 2.00 C HETATM 2643 C5 OHN E1169 22.417 12.612 84.569 1.00 2.00 C HETATM 2644 C4 OHN E1169 21.209 12.035 85.304 1.00 2.00 C HETATM 2645 C2 OHN E1169 20.403 13.049 83.438 1.00 2.00 C HETATM 2646 O6 OHN E1169 19.679 13.688 82.679 1.00 2.00 O HETATM 2647 OAP OHN E1169 20.010 12.548 84.638 1.00 2.00 O

10. Loading a PDB file Choose the file 2uv0.pdb

11. It should look something like this… Water molecules are shown as red crosses C green, N blue, O red, S yellow Default rendering is “line” Notice there are 2 molecules in the asymmetric unit Not identical: pseudo C 2

12. Impressive, but not very helpful We’d like to know some simple things, eg How many polypeptide chains per protein? Where is the autoinducer? What contacts does the protein make with the autoinducer?

13. View the chains using the ribbon function 1. hide everything 2. show ribbons If you don’t want to see something, hide it The default is to overlay images

14. Removing one copy of the protein Define chain G as a selection Type at the prompt (bottom left) PyMOL> select junk, chain G PyMOL> remove junk PyMOL> delete junk Recenter:

15. Showing the autoinducer as sticks

16. Turn on the helices and sheets Hide ribbon Show cartoon Play with the colors Run the ray-tracer (Ray)