talk - Phenix

Pipelining Ligands in PHENIX: 

eLBOW and REEL 

Nigel W. Moriarty 

Lawrence Berkeley National Laboratory 

Physical Biosciences Division 

PHENIX User Meeting March 28 th 2008

Ligands in Crystallography 

• Drug design 

• Biological function studies 

Generate 

ligand 

restraints 

Fit ligand to 

density 

Refine 

macromolecule 

and ligand 


e-LBOW Goals 

• Fast, simple and flexible procedure to include 

ligands in refinement 

• Reduce the tedium of building 3D ligand models 

• Automate generation of restraints for ligands 

• Comparison of ligand structures with PDB models 

Chemical input format 

Reflection Data 

Protein Information 

Chemical restraints file (CIF) 

Cartesian coordinates file (PDB) 

phenix.refine 


Ligand pipeline flowchart 

Chemical 

input 

Reflection 

data 

Protein 

model 

eLBOW 

Geometry 

LigandFit 

Ligand 

model 

phenix.refine 

Restraints 

Final 

model 


angles 

elements 

names 

atom 

dihedrals 

connectivity 

bond 

residue 

name 

centres 

chiral 

orders 

bond 

charges 

formal 

planes 

coordinates 

cartesian 

Input Formats 

• SMILES 

• PDB 

• xyz 

• Mol2D, Mol3D 

(v2000, v3000) 

• GAMESS input and 

output files 

• Sequences 

• PRODRG TXT 

format 

• Monomer library 

CIF 

• PDB Ligand 


The Algorithm 


The Algorithm 

• Parse SMILES or other input 

• Post process Molecular object 

– Atomic centres 

– Bonds 

• Determine ring and chiral 

structures 

• Construct a Z-Matrix model of 

heavy atoms 

– Z-Matrix is an internal coordinate 

representation 

– Chemically intuitive 

– Provide direct control of bonds, 

angles and dihedrals 


Z-Matrix 

C1 

H1 C1 1.1 

H2 C1 1.1 H1 109.6 

H3 C1 1.1 H1 109.6 H2 120.0 

H4 C1 1.1 H1 109.6 H2 -120.0 

H1 

Degrees of freedom = 3n-6 

H4 

C1 

H2 

H3 


The Algorithm 

• Optimise Z-Matrix geometry using 

simple force field (requires 

cartesian coordinates) 

• Add hydrogens 

• Optimise just hydrogens using 

simple force field 

• Optimise using AM1 semiempirical 

quantum chemical 

method 

• Check structure and repeat 

optimisation if necessary 

• Generate geometry restraints 


Comparison with experiment 

• MSD Chem ligand library contains 

– approximately 6443 ligands 

– SMILES strings 

– a smaller number of experimental cartesian coordinates 

– approximately 3285 comparable ligands 

• Input SMILES string to eLBOW 

• Compared AM1 geometry to experimental results from 

the PDB 


Refinement Tests 

• Use the restraints for ligand 

obtained from eLBOW in 

phenix.refine 

• Refined at resolution 1.2Å or 

better and 2.5Å (data 

truncated) 

• Compare to structure 

deposited in PDB 

• Legend 

– PDB 

– High 

– Low 

• Calculate RMSD using 

cartesian coordinates, bond, 

angle & dihedrals 

e-LBOW run time – 194 seconds 


Challenges 

• Disordered or weak density 

• Poor deposited ligand geometry 


Map Density Correlation 

• Compare the 

correlation of the 

model density to the 

2mFo-DFc map 

• phenix.refine and 

eLBOW provide 

provide ligand 

geometries that match 

the experimental data 

equally as well as the 

original libraries used 

in refinement 

• Results are also good 

for low resolution, but 

the comparison is 

difficult (PDB 

refinements were at 

high resolution) 


Histograms 

• Mean, Sigma 

– xyz: 0.11, 0.07 

– bond: 0.037, 0.017 

– dihedral: 6.0, 4.8 

• Differences between the original and new geometries are 

within the restraint standard deviations 


Flexibility 

• Simple command line interface 

• Scriptable using Python 

• Covalently bound ligands 

• Automatically treats all ligands in a PDB file 

• Close integration with refinement 


• Start COOT with 

eLBOW in COOT 

– coot --script $PHENIX/elbow/elbow_in_coot.scm 

– coot --script $PHENIX/elbow/elbow_in_coot.py 

• Build a ligand from SMILES or use the 

coordinates from an internal molecule 

• Manipulation of ligand geometry to provide a 

starting geometry 

• AM1 geometry optimisation 



• SMILES 

Simple run 

– phenix.elbow --smiles “c1ccccc1C” 

– phenix.elbow --smiles filename.smi 

– phenix.elbow --msd atp 

• PDB file 

– phenix.elbow --file filename.pdb 

– phenix.elbow filename.pdb 

• Atom naming 

– phenix.elbow --smiles “c1ccccc1C” --template 

names.pdb 

– phenix.elbow --msd atp --template atp.pdb 


Output 

• PDB file contains optimised geometry 

• CIF file contains restraints 

• filename.bonding.py is a Python script to 

change the bonding via --bonding 

• --tripos will output a TRIPOS file 

• --output will change the name of the output 

files 


Adjusting geometry 

• phenix.elbow --smiles “FC=CF” --opt - 

-view pymol 

• --view pymol 

– run “pymol filename.pdb” 

• Edit geometry and save as fixed.pdb 

• eLBOW will read the new geometry as 

starting geometry for AM1 optimisation 


Providing geometry 

• Starting geometry for AM1 optimisation 

– phenix.elbow --initial-geometry file.pdb 

• Final geometry to get a CIF file 

– phenix.elbow --final-geometry file.pdb 

– Provide corresponding SMILES for a better CIF 

file 


PDB options 

• A PDB file can have many ligands 

• To list all residues in a file 

– phenix.elbow file.pdb --all-residues 

• To run eLBOW on all “unknown” ligands 

– phenix.elbow file.pdb --do-all 

• To run eLBOW on a “known” ligand 

– phenix.elbow file.pdb --residue ATP 


PDB options (cont.) 

• To control the auto bond determination 

– -- auto-bond-cutoff=2.5 

• To control hydrogen addition 

– --add-hydrogens=True 

• To use a large PDB file as template 

– phenix.elbow --smiles “O” 

--template large.pdb --residue HOH 


eLBOW is Python scripts 

from elbow.command_line import builder 

molecule = builder.run(smiles=“O”, 

opt=True, 

quiet=True) 

print molecule.Display() 


Misc. features 

• --read-only to exit after reading input 

• --pickle to read topology file 

• --pipe to use | or open a pipe shell 

• --pymol to use progress in PyMOL if PHENIX 

installed 

• --quiet & --silent 

• --name sets ligand name 

• --id defaults to LIG 


eLBOW & phenix.refine 

• Run phenix.elbow ensuring that the atom 

naming is correct using a protein-ligand 

complex PDB file 

• Run phenix.refine using the CIF file 

• Possible to generate CIF link file using the 

CONECT or LINK record in the proteinligand 

complex PDB file 

• Add hydrogens to a PDB containing protein 

model and ligands 


Other eLBOW programs 

• phenix.metal_coordinate 

• elbow.join_cif_files, join_pdb_files 

– elbow.join_cif_files combined.cif file1.cif file2.cif 

• elbow.doc 


eLBOW Summary 

• e-LBOW provides easy to use methods to 

generate novel ligands and known ligands 

• AM1 provides a computationally efficient 

method for geometry optimisation of 

molecules containing any main group 

elements 

• Provides geometries comparable to existing 

methods 

• Readily pipelined into automatic refinement 

using phenix.refine and phenix.ligandfit 


What is REEL? 

• Restraints Editor eLBOW Ligands 

– Loads any restraints file 

– Loads any eLBOW input 

• Restraints Editor Especially Ligands 

– No linking or modifications 

• Restraints Editor Exclusively Ligands 

– Loads a small PDB file 

• Restraints Editor Effectively Ligands 

• “REEL will make a real difference” 

– A. White, Ph.D., Boehringer-Ingelheim GmbH 


Visualisation 


REEL features 

• Generate a geometry for a set of restraints 

• Modify restraints and generate new geometry 

• Fast editing in menu items 

• Highlight atom and restraints are highlighted 

• Multiple ligands simultaneously 

• Highlight restraint and atoms are highlighted 

• Save restraints (CIF) 

• Save geometry (PDB) 

• Run eLBOW 


eLBOW GUI 


REEL flowchart 

Chemical 

input 

eLBOW 

Geometry 

Restraints 

REEL 

Geometry 

Restraints 


Demonstration 


Summary 

• eLBOW converts many inputs to geometry 

and restraints 

• REEL allows restraints editing and several 

eLBOW features 


Acknowledgments 

• Computational Crystallography Initiative 

– Paul Adams 

– Pavel Afonine (phenix.refine) 

– Ralf Grosse-Kunstleve (cctbx, phenix.refine, HySS, ….) 

– Nick Sauter (iotbx, labelit) 

– Peter Zwart (mmtbx.xtriage, phenix.refine) 

• Other PHENIX developers 

– Cambridge University, Los Alamos National Laboratory, 

Texas A&M, Duke University 

• Others 

– CCP4 developers (MTZ library) 

– Alexei Vagin & Garib Murshudov (Monomer Library) 

• Funding: 

– LBNL (DE-AC03-76SF00098) 

– NIH/NIGMS (P01GM063210) 

– NIH/NIGMS (P01GM064692) 

– PHENIX Industrial Consortium

talk - Phenix

You also want an ePaper? Increase the reach of your titles

Delete template?

Save as template?