1. Protein Structure Database for Structural Genomics Group Jessica Lau December 13, 2004 M.S. Thesis Defense

2. Bioinformatics is Analysis of biological data: gene expression, DNA sequence, protein sequence. Data mining and management of biological information through database systems. At the Northeast Structural Genomics Consortium, database management systems play a large role in its daily operation Data collection and mining of experimental results Track target progress – status milestones Exchange information with rest of the world My thesis presents work in database management systems at the NESG. Part 1: ZebaView Part 2: Worm Structure Gallery Part 3: Prototype of NESG Structure Gallery

3. Zebaview is the official target list of the Northeast Structural Genomics Consortium Display summary table of NESG targets. –Status milestones –Protein properties: DNA and protein sequences, molecular weight, isoelectric point New targets are curated and then uploaded to SPiNE. 11,284 targets from 88 organisms.

4. Family View NESG Families Unfolded Membrane Core 50 Nf-kB

5. Target Summary Statistics Selected  Cloned  Expressed  Soluble  Purified  X-ray or NMR data collection  In PDB 4,418 targets cloned 141 structures 3.4% successful targets

6. GO, Cellular Localization, and SignalP Search for targets that have any of the three GO ontologies defined no GO ontologies defined at all 116 NESG structures do not have Molecular Function defined

7. LOCTarget Secretory proteins require formation of disulfide bonds Oxidative Folding needed for proper native folding 2,132 “Extracellular” NESG targets Bovine ribonuclease A has four disulfide bonds to stabalize its 3-D structure. Mahesh Narayan, et al. (2000) Acc. Chem. Res., 33 (11), 805 -812.

8. SignalP mRNA are translated with signal peptide for cellular localization Peptide is cleaved upon destination SignalP predicts cleavage of signal peptide Removal of signal peptide gives proper native fold Lodish et al. Molecular Cell Biology 4 th edition, Figure 7.1 (2000)

9. Part 2 – Worm Structure Gallery

10. Caenorhabditis elegans –Widely studied model organism 2-3 weeks life span, small size (1.5-mm-long), ease of laboratory cultivation, transparent body Small genome, yet has complex organ systems similar to higher organisms: digestive, excretory, neuromuscular, reproductive systems Donald Riddle et al, C. elegans II (1997) Altun Z F and Hall DH., Atlas of C. elegans Anatomy, Wormatlas (2002-2004)

11. System Components 22,653 C. elegans proteins 42 experimentally determined 4 are from NESG 24 homology models 14 are from NESG 960 C. elegans proteins potentially modeled Uniprot: Pfam domain, Gene name, ORF name PDB Coordinates Structure Validation Report Sequence similarities to proteins in PDB

12. 3-D structures from experimental determination (PDB) and homology modeling (HOMA) 42 experimentally determined structures –4 are from NESG 24 homology models –14 are from NESG

13. Data Captured: Uniprot –Pfam domain –Gene name –ORF name PDB Coordinates Structure Validation Report Alignment of possible models

14. Protein Structure Validation Software Suite of quality validation software –PROCHECK Quality of experimental data Distribution of φ, ψ angles in Ramachandran plot –MolProbity Clashscore Number of H atom clashes per 1,000 atoms With respect to a set of scores from 129 high resolution X-ray crystal structures < 500 residues, of resolution <= 1.80 Å, R-factor <= 0.25 and R-free <= 0.28; Bahattacharya, A et al. to be published

15. Algorithm based on alignment between query and template sequences. –Regions of conserved residues forms a set of constraints for modeling Sequence identity of 40% or more Good quality template Homology Modeling Automatically (HOMA)

16. Bad alignment  Bad model

17. Poor quality template  Poor quality model

18. Quality scores of 3-D structures

19. Search Search for C. elegans proteins in local database. Keyword: “Ubiquitin” in any field Results: 72 C. elegans proteins 2 Experimentally determined structures 1 Homology model 11 Potential models Results: 152 C. elegans proteins 2 Experimentally determined structures 1 Homology model 19 Potential models

20. System Architecture Java, Tomcat, MySQL, Perl. Three-tier architecture Client: Web browser Application: JSP, Logic components, Data access components Data: MySQL

21. Part 3 – NESG Structure Gallery

22. Structure files submitted by individual groups Structure information is entered into SPiNE manually Manually run PSVS and MolScript Structure files submitted by automated pipeline ADIT integrated with SPiNE for uniform format PSVS and images automatically generated Structure information from PSVS directly into SPiNE Archives structure files.

23. Downloads –Structure Validation Report –Structure related files Atomic coordinates NMR constraints NMR peak lists Chemical shifts Structure factor Annotation –Functional annotation provided by other NESG members –Uniprot –PDB coordinates file Reusing Java components from Worm Structure Gallery

24. –Enhance ZebaView performance to handle increased load and functionalities –Integrate annotation from other protein and structure databases. –Make modules available for other java-based applications within structural genomics. –Develop a gallery for other organisms: yeast, fruit fly, human –Continue specifications for the new NESG Structure Gallery

25. Advisor: Dr. Gaetano Montelione Thanks to everyone at the Protein NMR lab and NESG! Aneerban Bhattacharya John Everett All the scientists who solved the structures!