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Why PMB? Well this started as a place to put programs being developed for the Gulf Coast Structural Genomics Consortium (GCSCC). Never heard of the GCSCC? Well that explains why PMB does not include any model-building tools. However, I started to include our automated structure refinement tools in the PMB suite, and the name PMB remains. The murky origin of this software lies deep within the ancient past of protein structure refinement with Alec Hodel and Paul Harkins of Robert Fox's group at Yale. If you use these utilities please cite:

Emily E. Scott, Mark A. White, You Ai He, Eric F. Johnson, C. David Stout, and James R. Halpert, Structure of mammalian cytochrome P450 2B4 complexed with 4-(4 chlorophenyl)imidazole at 1.9 A resolution: Insight into the range of P450 conformations and coordination of redox partner binding J. Biol. Chem 279, 26, 27294-301, 2004. (PDF)

A more detailed description of PMB and the improved CNS routines included in pmb_bncs.f will be available when our Acta Cryst. D paper is published.

Best Regards,
Mark A. White
white@xray.utmb.edu



DOWNLOADS

Image and data processing programs - from our web-server  



PMB_CNS_utils.tar.gz


CNS utilities. Last updated Sept. 26, 2006. A typical installation will be placed in /usr/local/PMB (cd /usr/local ; tar -xzvf PMB_CNS_utils.tar.gz). Then add these lines to your .cshrc (/etc/csh.chrc) files:

setenv PMB_HOME /usr/local/PMB/

setenv PMB_BIN $PMB_HOME/linux #OR#

setenv PMB_BIN $PMB_HOME/irix


The instructions on how to recompile CNS with the new restraint routines are located in the file:

/.../PMB/CNS/other/pmb_bncs.f

PMB_BRUKER_utils.tar.gz

Bruker image and data processing

PMB_DIP2030_utils.tar.gz

Macscience calibration utility

PMB_HKL_utils.tar.gz

HKL utilities



INDEX

An index of programs and utilities in PMB



pmb_sca2cns pmb_CNS pmb_refine pmb_cns2phs pmb_start pmb_resolve pmb_check


PROGRAM DESCRIPTIONS

The most current versions are installed on our main server in $PMB_HOME



pmb_sca2cns scalepack.sca


This program converts a SCALEPACK merged HKL file into a CNS format CV file with a 5% test set. The test set is picked using a THIN SHELL algorithm, which is prefered when there is NCS. The files Fobs.o.cv and Fobs.cv are also generated. This utility also prints a $PMB_HOME/pmb_CNS command line for you to cut and paste or edit, if the parameters need adjustment. Normally only the high_res parameter may need to be changed, but check the syntax of the "Space(Group)" entry.
BUG: Automatically runs pmb_CNS with default values. Rerun pmb_CNS with revised values to overwrite all CNS input files.



pmb_CNS a b c alpha beta gamma "Space(Group)" SG_Number Resolution


This com file sets up ALL the CNS 1.1 input files using the given command line inputs. It also copies the necessary PMB refinement utilities into your current directory. The BINDIVIDUAL refinement script utilizes the improved B-factor restraints developed at UTMB by Mark A. White (Acta Cryst D submitted, 2006). Note the syntax of the "SpaceGroup" entry, which needs to be in quotes to be recognized as a single text string. {eg. P4(1), P2(1)2(1)2(1), C2(1), P3(1)21 }



pmb_refine -c5 -g -w2 -n -r -a -b -l -s22 -o -q -d -k -h [MODEL.pdb]


An automated CNS refinement program with several command line flags:
FLAGS:

-c{#} (Cycles {opt#cycles})       -Default 1 cycle of refinement
-g (Generate new topology files) - Do this after model building (1)
-w{#} (pick waters {opt#rounds})   - Default 1 round water pick
-n (NCS-shake)       - Relaxes NCS restraints for 1 round: "NCS Annealing"
-r (Rigid body refinement)  - Normally only used after MR
-a (Simulated Annealing)    - Do this with your initial models! (*See pmb_param.inp)
-b (Use H-bond restraints)    - For low resolution models (*See pmb_param.inp)
-l (LocalScale OFF flag)   - Turns OFF local scaling: PHS files are still generated (2)
-s{#} (Stereochem optimize )  - Optimize bond rmsd to Engh & Huber (*See pmb_param.inp) (3)
-o (A composit Omit Map)     - Generate a Composit Omit Map after refinement
-q (Occupancy refinement)  - qindividual of selected atoms!
-d{%} (Density modification)  - Use RESOLVE to generate a MAP (CCP4) (4)!
-k (ChecK structure)       - Use PROCHECK & SFCHECK to check model(CCP4) (4)!
-h (HELP)         - View this help message
Each refinement cycle continues until the freeR value stops decreasing. This routine utilizes the Czerwinski/Matthews/Hynes local scaling program to correct for absorption and anisotropic scattering. It generates an XtalView format PHS file for generating a “map” in xfit or COOT. The MTZ file new_sa.mtz generated with the -d flag (using pmb_resolve) is recommended for use in COOT.

(1) Run ./pmb_start on your rebuilt model OR specify your MODEL.pdb as the last entry on the command line.
(2) Local scaling should be turned off when the model is incomplete (<80%).
(3) See pmb_param.inp for bond_trgt and refined wa_scale value. Enter 0 to automatically determine or a bond_trgt value in thousanths of an A (3-25). A negative value will reset the wa_scale to CNS defaults.
(4) This option requires CCP4! RESOLVE: Input %Solvent or else let CNS determine.



pmb_cns2phs


A com file which launches the Czerwinski/Matthews/Hynes local scaling program pmb_cns2phs.I65, written in FORTRAN. This program uses the output of the CNS script FoFc_calc.inp (fo.sfo.fc.fob), which calculates Fc, Fbulk and phases, to produce an XtalView PHS format file (pmb_lscale.phs) and a corrected Fobs.cv (fobs_lscale.fob) file. The inclusion of the local scaling correction and bulk solvent component produces superior quality maps!



pmb_start PDB_file


Configures your PDB file for generate to create the connectivity and stereo chemical restraints list. This splits your PDB file into the necessary components for generate.inp: start.protein.pdb, start.ion.pdb, start.ligand.pdb, and start.water.pdb. The start.ion.pdb and start.ligand.pdb files may need manual editing to insert the necessary TER cards between each ion or ligand. The pmb_start com file may be edited to do this automatically for you. For example this the line to parse the TRIS (TRS) ligand, and automatically add the TER card after the last ATOM of each molecule "O3 TRS":

awk '/ TRS / {print $0 }; /O3 TRS/{printf"TER\n"}' $1 >> start.ligand.pdb

This function is now called if you specify the PDB file at the end of the \"pmb_refine -g\" command.



pmb_resolve


Create a solvent-flipped map (CCP4+RESOLVE). This runs RESOLVE using your latest model (minimize.pdb) and phases (pmb_lscale.phs). The MTZ files new_sa.mtz and resolve.mtz generated by pmb_resolve are recommended for use in COOT. This script is run after refinement with the pmb_refine -d flag.
BUG: Requires CCP4.



pmb_check


Check the Quality of your Model (CCP4) This runs both PROCHECK and SFCHECK on your latest model (minimize.pdb). This script is run after refinement with the pmb_refine -k flag.
BUG: Requires CCP4.



File Naming Conventions


generate.mtz(pdb), minimize.pdb, bindividual.pdb

The output files of generate.inp, minimize.inp and bindividual.inp. They are used as the input files for iterative minimization/b-factor refinement. The pmb_param.inp file can override the defaults set in these input files: (minimize, bindividual, anneal)

Fobs.o.cv, Fobs.cv

The ORIGINAL CV format data file, and the locally scaled file used by most refinement scripts.

start.pdb, start.ion.pdb, start.ligand.pdb, start.water.pdb

The input files for generate containing your protein model, the ions, any ligand, and the waters.

ligand.top, ligand.param

These ligand parameter and topology files must exist in your working directory. They may be empty if you do not have a ligand. (Look in $PMB_HOME/CNS/other for some examples)



HOW TO

A roughly cronological order of how to use the PMB utilities



WHAT TO DO

HOW TO DO IT.

Convert Scalepack to CNS

Run $PMB_BIN/pmb_sca2cns ScalePack.sca to generate your Fobs.cv file. This utility also copies the output file "ScalePack.sca.cv" to the two required CV files for you:Fobs.o.cv and Fobs.cv

Make the CNS input files

Run $PMB_HOME/pmb_CNS a b c alpha beta gamma “S(G)” SG_No. Res to automatically produce the required CNS input files and copy all the PMB utilities into your current directory.



Set up PDB for CNS

Then run "pmb_start My_Model.pdb" to set up your CNS input models. The start.ion.pdb file may need manual editing to insert the necessary TER cards between each ion. The script pmb_start may be edited to add your ligand to the list of automatically recognized ones:
For example this the line to parse the TRIS (TRS) ligand, and automatically add the TER card after the last ATOM of each molecule "O3 TRS":
awk '/ TRS / {print $0 }; /O3 TRS/{printf"TER\n"}' $1 >> start.ligand.pdb



Refine your model in CNS

To refine the model and generate a map run:
"pmb_refine -g |tee LogFile.log"
The pmb_refine program uses the current directory name as the PROJECT name. It creates data and log files with a date/project stamp for easy project management. The pmb_param.inp file can override the defaults set in these input files: (minimize, bindividual, anneal). This program has many flags to modify its behavior. To see the current list of flags type: pmb_refine -h



Build the Model

XtalView may be run using the "sharefonts;xtalmgr &" command.
You will need to create a project name and crystal data entry from the XTALMGR menu before using XFIT "xfit minimize.pdb pmb_lscale.phs" to examine your model there are links to local copies of the XtalView HowTo webpages on our web-server under SOFTWARE. To use COOT for your model building it is recommended that you run pmb_resolve in order to generate a MTZ file (new_sa.mtz or resolve.mtz). (Use the -d flag in pmb_refine) COOT can display PHS files, but there is a problem using the resulting map with COOT's refinement tools.



Use NCS and Solvent Flattening in RESOLVE/CCP4

Edit pmb_resolve to have the correct CELL and SG number. Pick suitable residues to use as the basis of NCS and edit the default selection criteria accordingly. Note that you do not want too many nor too few atoms to be selected. Note that this script requires that the model has been refined at least once in order to generate PHIC.



   

EXAMPLES

   



WHAT TO DO

A cut and paste example of how to do it.

Convert Scalepack to CNS CV

$PMB_BIN/pmb_sca2cns My_Data.sca

Set up files for CNS

$PMB_HOME/pmb_CNS 47.812 47.812 62.134 90.000 90.000 90.000 "P4(1)" 76 1.7

Prepare PDB model for CNS

pmb_start 1STN.pdb

Refine your model in CNS

pmb_refine -g -r -a -s 1STN.pdb |tee MMDDHH_My_Project_log

Build the Model in XTALVIEW

sharefonts;xtalmgr &
xfit minimize.pdb pmb_lscale.phs
xfit MMDDHHx.Project.pdb MMDDHHx.Project.phs

Use NCS and Solvent Flattening in RESOLVE

pmb_resolve



 

 

 

Mark A. White, white@xray.utmb.edu
X-RAY | SCSB | UTMB
Department of Biochemistry and Molecular Biology
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