Docstoc

NMR studies of biomolecular complexes

Document Sample
NMR studies of biomolecular  complexes Powered By Docstoc
					 RECOORD

           REcalculated COORdinates Database

Jurgen Doreleijers
Center for Eukaryotic Structural Genomics
University of Madison-Wisconsin
jurgen@bmrb.wisc.edu


Aart Nederveen
Bijvoet Center for Biomolecular Research
Utrecht University
a.j.nederveen@chem.uu.nl

Wim Vranken
Macromolecular Structure Database
European Bioinformatics Institute
wim@ebi.ac.uk
    Aim

•    Recalculation of protein structures based on
     deposited NMR restraints using state of the art
     methods

•    Goals:
     •    decrease user- and software-dependent biases
     •    allow a better comparison between structures
     •    comparison between different structure calculation programs
     •    provide a database for the development and assessments of
          validation tools and calculation protocols
           Overview recalculation project

                           1                     2               EBI/UU: 3
restraint       PDB:           BMRB:                             Generation of
                -coordinates   STAR files
manipulation                                                     consistent
                -restraints    Doreleijers et al. 2003
                                                                 STAR files




                                               CNS           4                   CYANA 5
recalculation                                  -topology                         -sequence
design of RECOORD                              -MD SA                            -MD SA
                                               -refinement                       -…




analysis                                                         analysis      6
                                                                 -improvement?
                                                                 -correlations?
                                                                 -…
Databases now publicly available

• DOCR/FRED (BMRB)
  databases containing converted and filtered restraints
  http://www.bmrb.wisc.edu/servlets/MRGridServlet



• RECOORD (EBI)
  database containing recalculated coordinates
  http://www.ebi.ac.uk/msd/recoord
                                                          1                      2
                                               PDB:           BMRB:
                                               -coordinates   STAR files
                                               -restraints    Doreleijers et al. 2003

    Selection
• Formats      (if distance restraints available):
      •   CNS/XPLOR
      •   DIANA/DYANA/CYANA
      •   DISCOVER/MSI

• PDB entries selected:
      • only proteins
      • no HET atoms
      • multimers allowed (not yet re-calculated)
      • at least 20 residues
•    Finally 545 monomers were selected
                                                                  EBI/UU: 3
                                                                  Generation of
                                                                  consistent
    Conversion issues                                             STAR files



• Data is converted to formats readable by calculation
     software (e.g. XPLOR/CNS and CYANA) by the
     FormatConverter available within CCPN software (Wim
     Vranken, EBI).

Problems:
• Differences between coordinate and restraint data:
      • e.g. 1 chain in pdb entry, 2 chains in restraint list
      • residue numbering can differ in PDB entry and restraint   list
      • restraints for residues not present in PDB entry…
•    Nomenclature in restraint list
                                               CNS           4   CYANA       5
                                               -topology         -sequence
                                               -MD SA            -MD SA

 Building topology
                                               -refinement       -…



• Starting script: generate_easy.inp from CNS
• Automated detection in original ensemble of:
   •   Disulfide bridges (<3Å S-S distance in original first models)
   •   CIS peptides (if |w|<25º in original first models)
   •   Protonation state of histidines (use CNS patches HISD, HISE)


• CYANA: sequence based on CNS topology
   •   Add CYSS, HIST, HIST+, cPRO in sequence
   •   Automated generation of disulfide restraints
                                       CNS           4   CYANA       5
                                       -topology         -sequence
CONDOR computer cluster CS             -MD SA            -MD SA
                                       -refinement       -…
University Madison

  • More than 800 processor used
  • Total CPU time: 31,169 hours (3.5 years on single
    workstation)


  • Example 2EZM, calculation of 1 model
    (101 a.a. & 2.2 GHz P4 computer)
     CYANA         31 seconds
     CNS        340 seconds
                                         analysis     6
                                         -improvement?
                                         -correlations?
Evaluation of structure quality          -…



• Agreement with experimental restraints
• Improvement?
• Comparison CNS and CYANA
• Relation NMR data quality and structural   quality
                                                                   analysis     6
                                                                   -improvement?
                                                                   -correlations?
            Distance restraints violations                         -…




                                                        ORG: 0.08 Å (0.14 Å)
                                                        original entries
frequency




                                                        CNW: 0.04 Å (0.05 Å)
                                                        recalculated in CNS
                                                        and refined in water




               RMS distance restraints violations (Å)
                                                                       analysis     6
                                                                       -improvement?
                                                                       -correlations?
            Dihedral restraints violations                             -…




                                                            ORG: 1.6° (4.6°)
                                                            original entries
frequency




                                                            CNW: 0.5° (0.5°)
                                                            recalculated in CNS
                                                            and refined in water




             RMS dihedral restraints violations (degrees)
                                                            analysis     6
                                                            -improvement?
    Results: quality indicators                             -correlations?
                                                            -…
    performance CNS vs. CYANA (no water refinement yet)




Average value over 545      Original PDB       CNS           CYANA
entries
                                           recalculation   recalculation
RMS distance restraints     0.08 ± 0.14    0.04 ± 0.06     0.04 ± 0.05
violations (Å)
RMS dihedral restraints      1.6 ± 4.6      0.5 ± 0.7       0.5 ± 0.7
violations (degrees)
Packing quality (Z-score)   -3.5 ± 1.9      -4.1 ± 1.9     -4.3 ± 1.8
WHATCHECK
Bumps per 100 residues        73 ± 63         11 ± 9         86 ± 37
% most favoured               69 ± 14        69 ± 13         61 ± 14
PROCHECK
                                                            analysis     6
                                                            -improvement?
    Results: quality indicators                             -correlations?
                                                            -…
    performance CNS before and after water refinement




Average value over 545      Original PDB       CNS         CNS + water
entries
                                           recalculation    refinement
RMS distance restraints     0.08 ± 0.14    0.04 ± 0.06     0.04 ± 0.05
violations (Å)
RMS dihedral restraints      1.6 ± 4.6       0.5 ± 0.7      0.5 ± 0.5
violations (degrees)
Packing quality (Z-score)   -3.5 ± 1.9      -4.1 ± 1.9     -2.5 ± 2.0
WHATCHECK
Bumps per 100 residues        73 ± 63         11 ± 9         10 ± 7
% most favoured               69 ± 14         69 ± 13        76 ± 11
PROCHECK
                                                                     analysis     6
    Improvement:                                                     -improvement?
                                                                     -correlations?
         packing and Ramachandran Z-scores                           -…



                                                     Improvent Z-score:
improvement Ramachandran




                                                     DZ=Zrefined - Zoriginal

                                                     For ~ 5 % of entries no
                                                     improvement possible
                                                     because of missing NMR
                                                     data compared to authors

                           missing data



                               improvement packing
                                                                               analysis     6
                                                                               -improvement?
            In search of correlations                                          -correlations?
            (Pearson coefficient)                                              -…

                                                      (correlations higher)        refined
                    data      RMS          circular    packing     Ramachandran     bumps
                    density   violations   variance    (Z score)   (Z score)

data density                  -0.23        -0.46       0.35        0.31             -0.03

RMS                 -0.11                  0.22        -0.25       -0.37            0.58
violations
circular            -0.32     0.00                     -0.60       -0.67            0.25
variance
packing             0.32      -0.06        -0.49                   0.69             -0.39
(Z-score)

Ramachandran        0.16      -0.11        -0.48       0.48                         -0.51
(Z-score)

bumps               0.04      0.04         0.07        -0.21       -0.47



                   original   (correlations lower)
                                                                                   analysis     6
                                                                                   -improvement?
            In search of correlations                                              -correlations?
            (Bumps)                                                                -…

                                                                                       refined
                       data        RMS       circular   packing     Ramachandran         bumps
                      density   violations   variance   (Z score)      (Z score)

data density                     -0.23        -0.46      0.35           0.31             -0.03

RMS                   -0.11                   0.22      -0.25          -0.37              0.58
violations
circular              -0.32       0.00                  -0.60          -0.67              0.25
variance
packing                0.32      -0.06        -0.49                     0.69             -0.39
(Z-score)

Ramachandran           0.16      -0.11        -0.48      0.48                            -0.51
(Z-score)

bumps                  0.04       0.04        0.07      -0.21          -0.47



                  original
                                                                                analysis     6
                                                                                -improvement?
            In search of correlations                                           -correlations?
            (NMR data density)                                                  -…

                                                                                    refined
                    data        RMS       circular   packing     Ramachandran         bumps
                   density   violations   variance   (Z score)      (Z score)

data density                  -0.23        -0.46      0.35           0.31             -0.03

RMS                 -0.11                  0.22      -0.25          -0.37              0.58
violations
circular            -0.32      0.00                  -0.60          -0.67              0.25
variance
packing             0.32      -0.06        -0.49                     0.69             -0.39
(Z-score)

Ramachandran        0.16      -0.11        -0.48      0.48                            -0.51
(Z-score)

bumps               0.04       0.04        0.07      -0.21          -0.47



                  original
                                                               analysis     6
                       Correlation NMR data density            -improvement?
                                                               -correlations?
                       Ramachandran Z-score                    -…




                                                      r=0.31
Ramachandran Z-score




                                NMR data density
                                                              analysis     6
                  Correlation NOE completeness and            -improvement?
                                                              -correlations?
                  packing Z-score                             -…




                                             r=0.20


                                             NMR data-based indicators
                                             cannot yield any indication of
packing Z-score




                                             the normality of the
                                             structures




                       NOE completeness
                                                                                 analysis     6
                                                                                 -improvement?
            In search of correlations                                            -correlations?
            (Precision)                                                          -…

                                                                                     refined
                     data        RMS       circular   packing     Ramachandran         bumps
                    density   violations   variance   (Z score)      (Z score)

data density                   -0.23        -0.46      0.35           0.31             -0.03

RMS                  -0.11                  0.22      -0.25          -0.37              0.58
violations
circular             -0.32      0.00                  -0.60          -0.67              0.25
variance
packing               0.32     -0.06        -0.49                     0.69             -0.39
(Z-score)

Ramachandran          0.16     -0.11        -0.48      0.48                            -0.51
(Z-score)

bumps                 0.04      0.04        0.07      -0.21          -0.47



                   original
                                                       analysis     6
          Correlation between precision and data       -improvement?
                                                       -correlations?
          density                                      -…



                                             r=-0.46
circular variance




                      NMR data density
                                                                         analysis     6
                    Correlation between precision and                    -improvement?
                                                                         -correlations?
                    Ramachandran                                         -…



                                                              r=-0.67


                                                              Protein with high
                                                              Ramachandran normality
circular variance




                                                              will have small circular
                                                              variance

                    1SUT




                     Ramachandran plot appearance (Z-score)
                                                                analysis     6
                    Correlation between RMSD and structural     -improvement?
                                                                -correlations?
                    uncertainty (QUEEN)                         -…




                                                 r=-0.69
backbone RMSD (Å)




                                                 Structural uncertainty
                                                 imposes lower limit to the
                                                 RMSD




                        structural uncertainty
    Conclusions I

• NMR-STAR files made consistent for 545 out of ±1700
     entries
•    Protocols and scripts available for recalculation in CYANA
     and CNS
•    Validation database available for testing of new protocols
•    Improvement compared to original data: 1 standard
     deviation closer to X-ray db
      • violations in original data do no limit recalculation effort
      • refinement in water required
      • 5 % no improvement: data missing
    Conclusions II

•    Correlations higher after recalculation and
     refinement, though most of them still weak

•    Highest correlation: precision vs. Ramachandran
     score & structural uncertainty (QUEEN)
    Acknowledgements
•    Utrecht University          Alexandre Bonvin
                                 Rob Kaptein
•    EBI Cambridge               Wim Vranken
•    CESG/BMRB                   Jurgen Doreleijers
                                 Zachary Miller
                                 Eldon Ulrich
                                 John Markley
•    Radboud University Nijmegen Chris Spronk
                                 Sander Nabuurs
•    RIKEN Japan                 Peter Güntert
•    Institut Pasteur Paris      Michael Nilges

				
DOCUMENT INFO
Shared By:
Categories:
Tags:
Stats:
views:5
posted:9/1/2012
language:English
pages:26