As described in the paper:

Prediction of sterically induced alignment in a dilute liquid crystalline phase:
aid to protein structure determination by NMR.

Markus Zweckstetter and Ad Bax
J. Am. Chem. Soc. , 122, (2000) 3791-3792
Contact: zweckste@speck.niddk.nih.gov

DOWNLOAD

Sparc Solaris 5.6 version
SGI Irix 6.2 version
RedHat Linux 6.0 version

The download provides a compressed tar archive with a PALES executable and example files. The archive can be unpacked with a command like the following:


   zcat pales.linux.tar.Z | tar xvf -

Users are encouraged to email the author to be informed about updates and related software.

What is PALES?
Components of the PALES Software
Changes to SSIA
How to Use PALES
Data Format
Test Files
A PALES Example Session
About the Name PALES

What is PALES?

PALES is a software for analysis of residual dipolar couplings. Its main component is the PALES (Prediction of ALignmEnt from Structure) simulation that predicts the magnitude and orientation of a sterically induced alignment tensor from a solute's (protein/nucleic acid/oligosaccharide) three-dimensional shape. This can be used to validate the correctness of derived structures, to distinguish monomeric from multimeric structures and to evaluate multiple-conformer models for flexible proteins. In addition, features for analysis of experimental dipolar couplings and dipolar coupling tensors are available, such as best-fitting a dipolar coupling tensor to its corresponding 3D structure.
pales cartoon 2

Components of the PALES Software

The PALES software consists of five modules:

prediction of molecular alignment/residual dipolar couplings from first principles (PALES; standard PDB format)
PALES for free format 3D structures
best-fit of measured dipolar couplings to 3D structure
basic analysis of dipolar coupling tensors
basic analysis of dipolar couplings

These components allow a rapid analysis of weak alignment that can be observed when a macromolecule is dissolved in a dilute liquid crystalline phase (N. Tjandra & A. Bax, Science , 278, (1997) 1111-1114). The components comprise the following:

Prediction of molecular alignment/residual dipolar couplings from first principles (PALES)
This module comprises the prediction of alignment from structure approach as previously distributed under the name SSIA. PALES predicts the magnitude and orientation of a solute's alignment from its three-dimensional shape. This can be used to validate the correctness of derived structures, to distinguish monomeric from multimeric structures and to evaluate multiple-conformer models for flexible proteins. The only required input is a PDB structure. Using the PDB structure the alignment tensor will be simulated assuming purely steric interaction between the molecule of interest and the liquid crystal particles. From the alignment tensor dipolar couplings can then be predicted. Liquid crystal systems having a dominant steric interaction include bicelles (M. Ottiger & A. Bax, J. Biomol. NMR , 12, (1998) 361-372) and poly(ethylene glycol)-based systems (M. Rueckert & G. Otting, J. Am. Chem. Soc. , 122, (2000) 7793-7797). In cases where electrostatic repulsion plays a major role the orientation of a solute's alignment can also reliably be predicted.
PALES for free format 3D structures
With this module a PALES simulation based on first principles can be performed for any array of three-dimensional coordinates. No standard PDB format is required. Simulation parameters are accessible in the same way as in 1).
Best-fit of measured dipolar couplings to 3D structure
The alignment tensor predicted based solely on the three-dimensional structure of a molecule can be compared to one back-calculated from a set of measured residual dipolar couplings. For such a best-fit of observed dipolar couplings to a 3D structure singular value decomposition and non-linear minimization are available. The order matrix analysis of residual dipolar couplings via singular value decomposition is performed as described by J.A. Losonczi, M. Andrec, M.W.F. Fischer and J.H. Prestegard, J. Magn. Reson. , 138, (1999) 334-342. Singular value decomposition only needs a minimum of 5 dipolar couplings, whereas non-linear minimization allows fixing of any of the five parameters describing the alignment tensor during minimization. In addition, this module offers the possibility to predict residual dipolar couplings from a user supplied alignment tensor. The required input is a PDB file and a minimum of five dipolar couplings.
Basic analysis of dipolar coupling tensors
This module allows manipulation of dipolar coupling tensors. Simple mathematical operations are possible for two dipolar coupling tensors such as addition or multiplication. In addition, back-calculation of Euler angles from an arbitrary symmetric and traceless matrix is possible as well as comparison of two alignment tensors in three and five dimensions. The minimum required input is a matrix with five independent elements.
Basic analysis of dipolar couplings
This module allows manipulation of two sets of dipolar couplings. Simple mathematical operations are possible for sets of dipolar couplings such as addition or multiplication. In addition, two sets of dipolar couplings can be compared statistically. The required input is at least one table with dipolar couplings.

Changes to SSIA

Besides the new name there are several changes compared to the original SSIA version of the program.

PALES is now fully usable for proteins, nucleic acids and oligosaccharides.
Any type of dipolar coupling can be used.
Parameters for a PALES simulation are controlled by command line arguments.
All standard PDB files can be used including multiple-chain molecules. A broad range of functions (written by Frank Delaglio) for selecting certain parts of a molecule were incorporated. In addition, a separate module is available for performing a PALES simulation on non-standard PDB files.
The input and output format has been unified with the NMRWish/DYNAMO and DC programs by Frank Delaglio.
PALES and the other programs now use the same internuclear distances for the N-HN, CA-HA, CA-C and N-C vectors (M. Ottiger & A. Bax, J. Am. Chem. Soc. , 120, (1998) 12334-12341).
All observed dipolar couplings are now supplied as isotropic - aligned splitting. The negative gyromagnetic ratio of 15N is taken into account within PALES.
Additional features for analysis of experimental dipolar couplings and dipolar coupling tensors were included.

How to Use PALES

Various options are available for the different PALES modules. To start off type

pales -help

This gives you the list of available command line arguments.

The basic use of the currently five available modules is as follows:

Prediction of molecular alignment/residual dipolar couplings from first principles (PALES) [default]
```
-stPales
```
- Basic simulation of magnitude and orientation of a solute's alignment (SSIA mode 1)
```
pales -pdb 1IGD_H_sDC.pdb
```
  (Default simulation parameters are used and results are written to standard output; the magnitude of alignment scales linearly with concentration and for molecules with dimensions of roughly the rod radius alignment tensors for the wall and the rod model are collinear.)
- Simulation of alignment tensor and prediction of dipolar couplings (SSIA mode 2)
```
pales -inD dc_1IGD.tab -pdb 1IGD_H_s.pdb -outD ssiaB.tbl \
      -s1 2 -a1 7 -pdbRot rot.pdb
```
  ( '-s1' and '-a1' are used for specifying an offset of 5 between the PDB file and the sequence number of the measured residual dipolar couplings; with '-pdbRot' a PDB file is written out that is rotated in such a way that the axes of the alignment tensor are parallel to the laboratory frame.)
- As above but specifying various simulation parameters
```
pales -inD dc_1IGD.tab -pdb 1IGD_H_s.pdb -outD ssiaC.tbl \
      -s1 2 -a1 7 -r1 10 -rN 60 \
      -bic -wv 0.03 -dGrid 0.3 -dot 133 -digPsi 23 -rM 33.0 \
      -lcS 0.83 -rA 0.3 -H -nosurf
```
  ('-r1' and '-rN' are used for selection of residues 10 to 60 in the simulation.)

PALES for free format 3D structures

-stPalesFree

Basic simulation of magnitude and orientation of a solute's alignment
```
pales -stPalesFree -pdbF 1IGD_H_s.pdb -outA ssiaF.tbl
```

As above but specifying various simulation parameters

pales -stPalesFree -pdbF 1IGD_H_s.pdb -outA ssiaF.tbl \
      -bic -wv 0.03 -dGrid 0.3 -dot 133 -digPsi 23 -rM 33.0 \
      -lcS 0.83 -rA 0.3 -H -nosurf

Best-fit of measured dipolar couplings to 3D structure

-bestFit

Singular value decomposition
```
pales -bestFit -inD dc_1IGD.tab -pdb 1IGD_H_s.pdb \
      -outD svd.tbl -s1 2 -a1 7 -pdbRot rot.pdb   \
      -map 500 -outMap worldmap.txt
```
(The '-map' flag is used for mapping the deviation of alignment tensor orientations. These are written to 'worldmap.txt'. 500 iterations are done.)

Back-calculation of dipolar couplings from a user supplied order matrix

pales -bestFit -inD dc_1IGD.tab -pdb 1IGD_H_s.pdb \
      -outD saupePred.tbl -s1 2 -a1 7 \
      -saupe -9.2042e-05  2.3990e-04  3.8255e-04 -4.4549e-04  3.9788e-04

(The order of matrix elements is Szz, Sxx-yy, Sxy, Sxz and Syz.)

Back-calculation of residual dipolar couplings for Fixed Da, Dr, and Rotations.

pales -bestFit -inD dc_1IGD.tab -pdb 1IGD_H_s.pdb \
      -outD dadrFixed.tbl -s1 2 -a1 7 \
      -fixed -da -4.135779e-04 -dr -6.901196e-05  \
      -psi -43.23 -theta 149.43 -phi 81.40

Non-linear minimization for best alignment orientation with fixed Da & Dr

pales -bestFit -inD dc_1IGD.tab -pdb 1IGD_H_s.pdb \
      -outD dadrOnlyFixed.tbl -s1 2 -a1 7 \
      -dadr -da -4.135779e-04 -dr -6.901196e-05

Basic analysis of dipolar coupling tensors

-anA

pales -anA -outA anA.tbl \
      -inS1 -8.9631e-05  2.4300e-04  3.8479e-04 -4.4164e-04  3.9631e-04 \
      -inS2 -1.3042e-05  4.1560e-04  3.5832e-04 -4.6099e-04  4.0923e-04

Basic analysis of dipolar couplings

-anDC

pales -anDC -outD anDC.tbl -inD1 dc_1IGD.tab -inD2 dc_1IGD.tab \
      -s1 20 -sN 40

Data Format

Input

PDB file

All standard PDB files can be used (including MOLMOL files).

IMPORTANT: All atoms in the PDB file will be used including pseudo atoms (ANI), if no appropriate selection command line arguments are specified.

Dipolar coupling input

The protein sequence should be given as shown by one or more "DATA SEQUENCE" lines. Space characters in the sequence will be ignored.
The table must include columns for residue ID, three-character residue name and the atom name for both atoms that are involved in the dipolar coupling as well as the dipolar coupling itself, its error and a weighting factor. The atom notation must match that of the PDB file. Segment ID and Chain ID are optional.
The table must include a "VARS" line that labels the corresponding columns of the table.
The table must include a "FORMAT" line that defines the data type of the corresponding columns of the table.
Lines with a '#' sign as first character as well as empty lines are ignored.

Example dipolar coupling table (excerpts):

DATA SEQUENCE MQIFVKTLTG KTITLEVEPS DTIENVKAKI QDKEGIPPDQ QRLIFAGKQL
DATA SEQUENCE EDGRTLSDYN IQKESTLHLV LRLRGG

VARS   RESID_I RESNAME_I ATOMNAME_I RESID_J RESNAME_J ATOMNAME_J D      DD    W
FORMAT %5d     %6s       %6s        %5d     %6s       %6s    %9.3f   %9.3f %.2f

    2    GLN      N      2    GLN     HN     -15.524     1.000 1.00
    3    ILE      N      3    ILE     HN      10.521     1.000 1.00
    4    PHE      N      4    PHE     HN       9.648     1.000 1.00
    5    VAL      N      5    VAL     HN       6.082     1.000 1.00

    1    MET      C      2    GLN     HN       3.993     0.333 3.00
    2    GLN      C      3    ILE     HN      -5.646     0.333 3.00
    3    ILE      C      4    PHE     HN       1.041     0.333 3.00
    4    PHE      C      5    VAL     HN       0.835     0.333 3.00

    1    MET      C      2    GLN      N       2.651     0.125 8.00
    2    GLN      C      3    ILE      N      -3.768     0.125 8.00
    3    ILE      C      4    PHE      N       1.463     0.125 8.00
    4    PHE      C      5    VAL      N      -1.726     0.125 8.00

    2    GLN      N      2    GLN     HN     -15.524     1.000 1.00
    3    ILE      N      3    ILE     HN      10.521     1.000 1.00
    4    PHE      N      4    PHE     HN       9.648     1.000 1.00
    5    VAL      N      5    VAL     HN       6.082     1.000 1.00

    1    MET     HA      1    MET     CA     -38.341     1.000 0.50
    2    GLN     HA      2    GLN     CA      11.662     1.000 0.50
    3    ILE     HA      3    ILE     CA      18.424     1.000 0.50
    4    PHE     HA      4    PHE     CA      26.733     1.000 0.50

Output

If no output files are specified results are written to standard output.

Following parameters/values are reported:
- Simulation parameters used.
- Order matrix, its irreducible representation and the corresponding norm.
- Eigensystem from diagonalization.
- Euler angles for clockwise rotation about z, y', z''.
- Euler angles for rotation about three independent axes.
- Magnitude of alignment in a three-dimensional representation as given by Da=1/2Szz and Dr=1/3(Sxx-Syy) (G. M. Clore, A. M. Gronenborn & A. Bax, J. Magn. Reson. , 133, (1997) 216-221).
- Statistics of observed and predicted dipolar couplings.
- Input dipolar couplings, predicted dipolar couplings and their difference in the same format as the input dipolar coupling table (see above). The columns with the predicted dipolar couplings and the difference follow that of the observed ones (see below).
To assess a prediction look at the following parameters:
- Magnitude of alignment
  The degree of alignment can be judged from the norm of the irreducible representation or from the magnitude of the largest eigenvalue Szz of the order matrix. Using the latter, however, can be misleading in the case of high rhombicity.
  (To get the magnitude of alignment in Hz multiply Da by the dipolar interaction value given in the output table.)
- Orientation of alignment
  The orientation of dipolar coupling tensor is described by three Euler angles, defined as follows:
  
  alpha Clockwise rotation around z, leading to new system x',y',z'
  
  beta Clockwise rotation around y', leading to new system x'',y'',z''
  
  gamma Clockwise rotation around z''
  
  Limits: 0 <= alpha < 360, 0 <= beta < 180, 0 <= gamma < 360.
  Four equivalent Euler orientations are reported, due to the sign ambiguity of the eigenvectors.
- Statistics
  Statistics were computed according to methods given in: W.H. Press, S.A. Teukolsky, B.P. Flannery and W.T. Vetterling: Numerical Recipies in C Cambridge, Cambridge University Press, 1988

Test Files

1IGD_H_s.pdb	PDB file of the protein G domain (shortened by 5 residues and protons added with MOLMOL )
dc_1igd.txt	File with measured D(N-HN) dipolar couplings
chkPALES2.1.com	Shell script for testing the different modules

In addition, output files from test runs are included. For visualization of alignment tensor distributions the file 'mapTemplate.tab' with the mapping coordinates of the map itself is included ( J.A. Losonczi, M. Andrec, M.W.F. Fischer and J.H. Prestegard, J. Magn. Reson. , 138, (1999) 334-342). The maps can be viewed with the free program Xmgr.

A PALES Example Session

Program call

pales -pdb 1IGD_H_sDC.pdb -outD ssia.tbl

Output (ssia.tbl)

REMARK Molecular Alignment Simulation.

REMARK Simulation parameters.

DATA PALES_MODE STERIC

DATA PALES LC_TYPE               	wall
DATA PALES LC_CONCENTRATION 	0.050
DATA PALES ORIENT_SPHERE       	100
DATA PALES ORIENT_PSI          	18
DATA PALES GRID_SPACING        	0.200
DATA PALES MODEL_RADIUS        	20.000
DATA PALES LC_ORDER            	0.800
DATA PALES ATOM_RADIUS         	0.000
DATA PALES SEL_SIMPLE_FLAG     	0
DATA PALES SURF_FLAG           	1

REMARK Order matrix.

DATA SAUPE  -8.9631e-05  2.4300e-04  3.8479e-04 -4.4164e-04  3.9631e-04

DATA IRREDUCIBLE REPRESENTATION (A0,A1R,A1I,A2R,A2I)   -3.0583e+00  1.2304e+01  1.1041e+01  3.3849e+00 -1.0720e+01
DATA IRREDUCIBLE GENERAL_MAGNITUDE   2.8438e+01

REMARK Eigensystem & Euler angles for clockwise rotation about z, y', z''.

DATA EIGENVALUES (Axx,Ayy,Azz)    3.0889e-04  5.1593e-04 -8.2482e-04
DATA EIGENVECTORS
DATA EIGENVECTORS XAXIS  1.2036e-01  7.6741e-01  6.2976e-01
DATA EIGENVECTORS YAXIS  8.5264e-01  2.4500e-01 -4.6150e-01
DATA EIGENVECTORS ZAXIS -5.0845e-01  5.9250e-01 -6.2483e-01

DATA Q_EULER_SOLUTIONS    ALPHA     BETA    GAMMA
DATA Q_EULER_ANGLES  1   323.77   128.67    49.37
DATA Q_EULER_ANGLES  2   143.77   128.67    49.37
DATA Q_EULER_ANGLES  3   216.23    51.33   229.37
DATA Q_EULER_ANGLES  4    36.23    51.33   229.37


REMARK Euler angles (psi/theta/phi) for rotation about x, y, z.

DATA EULER_SOLUTIONS 2
DATA EULER_ANGLES  -43.48  149.44   81.97
DATA EULER_ANGLES  136.52   30.56  261.97

DATA Da -4.124083e-04
DATA Dr -6.901216e-05


REMARK Dipolar couplings.

DATA N                    		41
DATA RMS                  		1.222
DATA Chi2                 		61.223
DATA CORR R               	0.996
DATA CORNILESCU Q         	0.120
DATA REGRESSION OFFSET    	-0.739 +/- 0.148 [Hz]
DATA REGRESSION SLOPE     		0.977 +/- 0.015 [Hz]
DATA REGRESSION BAX SLOPE 	0.981 +/- 0.010 [Hz]

VARS    RESID_I RESNAME_I ATOMNAME_I RESID_J RESNAME_J ATOMNAME_J DI D_OBS D D_DIFF DD W
FORMAT  %4d %4s %4s %4d %4s %4s %9.2f %9.3f %9.3f %9.3f %.2f %.2f

    2  THR   HN    2  THR    N -21523.10    1.4640    0.0857    1.3783  1.0000 1.00
    3  TYR   HN    3  TYR    N -21523.10    7.0570    7.8340   -0.7770  1.0000 1.00
    4  LYS   HN    4  LYS    N -21523.10    8.6540    7.3118    1.3422  1.0000 1.00
    5  LEU   HN    5  LEU    N -21523.10   12.1800   10.9437    1.2363  1.0000 1.00
    7  LEU   HN    7  LEU    N -21523.10   12.6910   10.0725    2.6185  1.0000 1.00
    8  ASN   HN    8  ASN    N -21523.10    5.2020    4.5600    0.6420  1.0000 1.00
   12  LEU   HN   12  LEU    N -21523.10   11.6770   10.3776    1.2994  1.0000 1.00
   14  GLY   HN   14  GLY    N -21523.10   10.5530   11.0063   -0.4533  1.0000 1.00
   15  GLU   HN   15  GLU    N -21523.10   11.1540    9.9303    1.2237  1.0000 1.00
   16  THR   HN   16  THR    N -21523.10   11.0090   10.3052    0.7038  1.0000 1.00
   17  THR   HN   17  THR    N -21523.10    8.8740    8.1451    0.7289  1.0000 1.00
   18  THR   HN   18  THR    N -21523.10    5.5190    5.7485   -0.2295  1.0000 1.00
   19  GLU   HN   19  GLU    N -21523.10    5.6510    6.1800   -0.5290  1.0000 1.00
   20  ALA   HN   20  ALA    N -21523.10    4.1730    4.9946   -0.8216  1.0000 1.00
   21  VAL   HN   21  VAL    N -21523.10    4.7990    4.5163    0.2827  1.0000 1.00
   22  ASP   HN   22  ASP    N -21523.10   -4.2940   -4.4661    0.1721  1.0000 1.00
   23  ALA   HN   23  ALA    N -21523.10   -2.3260   -2.5724    0.2464  1.0000 1.00
   24  ALA   HN   24  ALA    N -21523.10  -13.4330  -15.2750    1.8420  1.0000 1.00
   25  THR   HN   25  THR    N -21523.10  -10.5960  -12.4346    1.8386  1.0000 1.00
   26  ALA   HN   26  ALA    N -21523.10   -5.4130   -6.6161    1.2031  1.0000 1.00
   28  LYS   HN   28  LYS    N -21523.10  -16.1580  -16.7218    0.5638  1.0000 1.00
   30  PHE   HN   30  PHE    N -21523.10   -8.6620   -8.3276   -0.3344  1.0000 1.00
   31  LYS   HN   31  LYS    N -21523.10  -13.9680  -14.9181    0.9501  1.0000 1.00
   32  GLN   HN   32  GLN    N -21523.10  -16.0390  -16.6349    0.5959  1.0000 1.00
   33  TYR   HN   33  TYR    N -21523.10  -12.0490  -12.6305    0.5815  1.0000 1.00
   34  ALA   HN   34  ALA    N -21523.10   -9.6080  -10.3609    0.7529  1.0000 1.00
   35  ASN   HN   35  ASN    N -21523.10  -15.6960  -16.4080    0.7120  1.0000 1.00
   36  ASP   HN   36  ASP    N -21523.10  -15.0910  -14.7718   -0.3192  1.0000 1.00
   37  ASN   HN   37  ASN    N -21523.10   -3.6870   -3.9808    0.2938  1.0000 1.00
   38  GLY   HN   38  GLY    N -21523.10   -8.0650   -6.7237   -1.3413  1.0000 1.00
   44  THR   HN   44  THR    N -21523.10   11.6760    9.1670    2.5090  1.0000 1.00
   45  TYR   HN   45  TYR    N -21523.10   11.8120   10.9110    0.9010  1.0000 1.00
   46  ASP   HN   46  ASP    N -21523.10   10.7600    9.6707    1.0893  1.0000 1.00
   47  ASP   HN   47  ASP    N -21523.10   11.0430   10.4336    0.6094  1.0000 1.00
   49  THR   HN   49  THR    N -21523.10    2.4570    0.0918    2.3652  1.0000 1.00
   50  LYS   HN   50  LYS    N -21523.10    8.5140    7.4738    1.0402  1.0000 1.00
   51  THR   HN   51  THR    N -21523.10    9.3680    9.1265    0.2415  1.0000 1.00
   52  PHE   HN   52  PHE    N -21523.10   11.5930   10.7652    0.8278  1.0000 1.00
   53  THR   HN   53  THR    N -21523.10   10.1490    8.8621    1.2869  1.0000 1.00
   54  VAL   HN   54  VAL    N -21523.10   12.2240    9.9134    2.3106  1.0000 1.00
   56  GLU   HN   56  GLU    N -21523.10   11.6220    9.1445    2.4775  1.0000 1.00

About the Name PALES

Pales is the Roman patron goddess of shepherds and flocks. Pales also presides over the health and fertility of the domestic animals. Her festival is the Palilia (also called the Parilia) and was celebrated by shepherds on April 21, the legendary founding date of Rome. On that day large fires were made through which they drove the cattle.

roman coin

Subsequently, an asteroid (discovered September 19th 1857) and a butterfly ( Boloria pales ) was named after the goddess.

alpha	Clockwise rotation around z, leading to new system x',y',z'
beta	Clockwise rotation around y', leading to new system x'',y'',z''
gamma	Clockwise rotation around z''