Aims

• To illustrate the structural variability that can be present in an an ensemble of NMR-derived structures.

Objectives

After this practical you will:

• have seen ensembles of NMR-derived structures;
• have compared structures determined by X-ray crystallography and NMR;
• understand how biological units can be reconstructed from data on a single subunit in a Protein Data Bank file.

Exercises

1. Protein Data Bank entry 1SAE contains an ensemble of 26 structures of human p53 tumor suppressor. Load this file into RasMol, and colour the structure by chain, and then by "temperature". Look at the Protein Data Bank file for entry 1SAE, and read remarks 5-10.

   Further structures are in Protein Data Bank entries 1SAG (26 structures) and 1SAI (25 structures). Protein Data Bank entry 1SAK contains the minimized average structure.

2. Protein Data Bank entry 1C26 contains an X-ray structure of a subunit of human p53 tumor suppressor. The biological unit (seen in Question 1) consists of four copies of this subunit, so download the biological unit in addition to the usual structure file (use the 'gunzip' command to uncompress the file that you download). Compare this structure with the structures seen in Question 1.

3. Suppose that a PDB file containing the biological unit cannot be obtained from the Protein Data Bank, and that only the basic entry containing the single chain present in the crystallographic asymmetric unit is available. Describe how you would generate the coordinates for the biological unit from PDB entry 1C26 (hint: look at remarks 300 and 350 in the PDB file). How do the matrices in remark 350 relate to the transformation code that you have used in the practicals? Describe the transformation represented by each of the four BIOMT matrices.

4. Protein Data Bank entry 1AAF contains an ensemble of 20 structures of HIV-1 nucleocapsid protein. Load this file into RasMol.

   To view a single structure (e.g. structure 1), type "restrict */1". Select the Cys residues in model 1 ("select cys/1") and display these as sticks. Select zinc atoms in this model ("select zn/1") and display these in the spacefill style.

5. Read the description of the NMRCORE program. Summarise the purpose of that program, and the algorithm that is uses.

6. NMRCLUST was introduced in an earlier lecture simply as a program for clustering structural fragments. Look at that paper again, and think about the task of clustering structures in the context of NMR structure determination.