Protein Assignment 2

Assignments Index

The Secondary Structure of Your Protein

General Remarks

This course is self-paced, so PLEASE DON'T FEEL GUILTY if you haven't done assignment 1. But DON'T skip it. A very large number of errors are made because people don't know enough about what the constitution of their protein is. (I've come across crystallographers more than once who weren't precisely sure what the sequence was of the structure they'd solved. And you can't tell by looking at the density whether you've got an Asp or and Asn...). And crystallographers produce very high class data compared with some other disciplines ...

All questions refer to YOUR protein. Confine your answers to SECONDARY structure (i.e. helices, sheets, turns) and not the tertiary structure (complete fold). If your protein has no helices, for example, find someone in your group who *has* , and vice versa and exchange information. Similarly, some of the questions may ask for operations that require PROCHECK. There should be someone in your group who has access to it.

(****DO NOT MAIL THE COURSE ADMIN, BUT USE YOUR GROUP OR THE pps96-proteins LIST***)

Brookhaven PDB ANNOTATIONS

(Only use the pre-ATOM cards for this information!)

1. Do any of the REMARK cards mention secondary structure?
2. Do you have SHEET, HELIX, TURN cards? Can you make sense of them?
3. Does your protein have DISULphides? Are they intra- or inter-chain? (Oh dear - PM-R spells it -PH-; too old to change :-)
4. Draw out the linear sequence of your protein with the SHEETS, HELIXs, TURNs as annotation. Are there any gaps? Are there subtypes of any of these? Are there any PRO residues in helices? Is this a problem?
5. Are there any independent observations of your protein (NOT homologous proteins). Either in different crystal forms? Or by different authors? Or complexed with different ligands? If there are, pick NOT MORE than 3 and see if they all have the same SHEETs, etc If they differ, why? Would the list be interested?
6. IF there are studies by both NMR and X-ray, tell us about them!
7. What does PDB say about HBONDS?
8. Are there any areas where the secondary structure is uncertain?

   Paper Literature

9. If you are really keen, find a paper about the structure or the abstract in Macromolecular Structures. What do these say about the secondary structure?

   (****DO NOT MAIL THE COURSE ADMIN, BUT USE YOUR GROUP OR THE pps96-proteins LIST***)

   Brookhaven Coordinates

   (Use the structure to make your own deductions about the secondary structure). (Compare these with what you found above)

10. Use RasMol to show where its algorithm decides the secondary structure elements are. Does it differ from the PDB assignment? If so, why?
11. Measure (say) 5 phi/psi torsion angles in (a) some strands (b) some helices (c) some turns. Do the values seem reasonable?
12. Look *along* the axis of the helices. Are they regular? straight? and for the strands?
13. Can you plot a simple helical wheel for your helices? Are they amphipathic? Does this make sense?
14. PROCHECK is a program written at UCL, London which calculates dihedral angles from coordinates, assigns secondary structure and plots a Ramachandran map of the backbone phi/psi angles. If you have not already done so, do this (or find someone who can, or find another program...) for your protein. Click here for a WWW interface to PROCHECK for use by PPS participants.
15. Is there any evidence that parts of your structure are either poorly determined or possibly incorrect/unusual?

    Graphics

16. Make a gif of your protein, showing the secondary structural elements to their best advantage. (For a large protein you might wish to edit out some of the structure and produce more than one picture).

(****DO NOT MAIL THE COURSE ADMIN, BUT USE YOUR GROUP OR THE pps96-proteins LIST***)

Assignments Index