Strukturrechnung mit CYANA (2014)
Contents |
Startup
Log in to the Windows system with the username and password of your HRZ account.
The structure calculation will be performed on a Linux server that can be accessed by clicking the icon labeled BCC Linux Terminal-Service.nxs. The username and password for the Linux server are the same as for the Windows XP computer, and the data in the home directories are shared between the two systems.
Download the data for the practical: Data.tgz and store it in your home directory.
In a terminal window, unpack the data and start the bash shell
tar zxf Data.tgz bash
Three subdirectories will be created:
- cyana1: structure calculation using only the assigned 15N-resolved NOESY peak list
- cyana2: structure calculation using the assigned 15N-resolved NOESY peak list and the unassigned 13C-resolved NOESY peak list
- cyana3: structure calculation using the unassigned 15N-resolved and 13C-resolved NOESY peak lists
Enter the corresponding subdirectory. For example
cd cyana1
and copy your data to this directory. For example, for the 'cyana1' calculation, copy your chemical shift list (shifts.prot) and your assigned peak list from the 15N-resolved NOESY spectrum (n15assigned.peaks).
Run the CYANA structure calculation
Start CYANA, and execute the CYANA script CALC.cya (in 'cyana1') or AUTO.cya (in 'cyana2' and 'cyana3'):
> cyana
___________________________________________________________________
CYANA 3.0 (intel)
Copyright (c) 2002-10 Peter Guntert. All rights reserved.
___________________________________________________________________
Library file "/usr/local/soft/cyana-3.0/lib/cyana.lib" read, 38 residue types.
Sequence file "sequence.seq" read, 114 residues.
cyana> CALC
Analyze the results of the structure calculation
The results of the structure calculation are the structural statistics in the overview file, cyana.ovw ('cyana1') or final.ovw ('cyana2' and 'cyana3'), and the structure itself, which is represented by a bundle of 10 conformers whose coordinates are stored in the PDB file, cyana.pdb ('cyana1') or final.pdb ('cyana2' and 'cyana3').
The overview file has at least three parts. The file starts with a table of the target function values and restraint violation statistics. For example:
Structural statistics:
str target upper limits van der Waals torsion angles
function # rms max # sum max # rms max
1 1.69 2 0.0076 0.36 4 5.6 0.34 0 0.3302 3.23
2 1.74 2 0.0077 0.36 5 5.9 0.34 0 0.3272 3.30
3 1.75 1 0.0075 0.36 5 5.7 0.34 0 0.3695 3.45
4 1.87 1 0.0075 0.37 7 6.3 0.34 0 0.3159 2.69
5 1.95 1 0.0075 0.37 5 6.7 0.37 0 0.3185 3.00
6 2.12 2 0.0084 0.36 6 6.6 0.34 0 0.3745 3.56
7 2.19 2 0.0100 0.50 7 6.8 0.34 0 0.3257 3.29
8 2.35 2 0.0096 0.36 8 7.0 0.34 0 0.3748 3.50
9 2.40 2 0.0088 0.36 5 8.4 0.35 0 0.4152 3.44
10 2.49 2 0.0090 0.36 9 7.6 0.33 0 0.3494 3.20
Ave 2.06 2 0.0084 0.38 6 6.7 0.34 0 0.3501 3.26
+/- 0.28 0 0.0009 0.04 2 0.8 0.01 0 0.0308 0.25
Min 1.69 1 0.0075 0.36 4 5.6 0.33 0 0.3159 2.69
Max 2.49 2 0.0100 0.50 9 8.4 0.37 0 0.4152 3.56
Cut 0.20 0.20 5.00
This table has one row for each structure, containing
- the rank of the structure sorted by target function value
- the target function value
- three columns for each type of conformational restraints:
- the number of restraints that are violated by more than the cutoff value given in the last row (“Cut”)
- the root-mean-square (RMS) violation calculated over all, violated and fulfilled, restraints of this type
- the maximal violation
The five bottom lines of the Table give the average value, the standard deviation, the minimum value, and the maximum value of the corresponding quantity over the individual structures, as well as the cutoff value for significant violations.
Restraints that are violated in a significant number of structures by more than the corresponding cutoff value are reported in the second part of the overview file:
Constraints violated in 3 or more structures:
# mean max. 1 5 10
Upper QB LEU 17 - QB PRO 108 3.69 3 0.10 0.50 ++ * peak 1009
Upper HB ILE 85 - H ASP 86 3.80 10 0.36 0.37 +++*++++++ peak 803
VdW N ILE 81 - HD2 PRO 82 2.45 10 0.34 0.37 ++++*+++++
VdW CG2 ILE 81 - C ILE 81 2.90 6 0.20 0.21 + + ++* +
2 violated distance restraints.
0 violated angle restraints.
Each line identifies a violated restraint, and gives the number of structures in which the restraint is violated by more than the aforementioned cutoff value (column labeled “#”), its maximal violation (column “max.”), and the structures in which the violations occur (a one-character column for each structure that is analyzed). Structures in which the restraint is violated by more than the cutoff are marked with “+”, or with a “*” for the structure in which the maximal violation occurs. If available, the number of the cross peak from which the restraint originated is given at the end of the line.
At the end of the overview file, root-mean-square deviation (RMSD) values for the atom positions after optimal superposition of the individual conformers onto the mean coordinates are given:
RMSDs for residues 10..100: Average backbone RMSD to mean : 0.58 +/- 0.11 A (0.45..0.82 A; 10 structures) Average heavy atom RMSD to mean : 1.08 +/- 0.11 A (0.93..1.25 A; 10 structures)
The residue range used for the superposition is indicated. RMSD values are computed for the backbone and heavy atoms of the given residues. The average value, the standard deviation, and the minimal and maximal values of the RMSDs between the analyzed structures and their mean coordinates are calculated.
Visualize the structure
The program MOLMOL can visualize the bundle of conformers that represents the solution structure of the peptide with the command
molmol -r 7-37 cyana.pdb
to start the program MOLMOL and to show a superposition of the 10 conformers whose coordinates are stored in the PDB file cyana.pdb. The option "-r 7-37" indicates MOLMOL to optimally superimpose the backbone atoms of residues 8-21.
Alternatively, other molecular viewers such as 'pymol' can be used.
