1. #1 Bottleneck is Psychological!!!
Structural Biology in the 3rd Millennium
Structural Genomics
Structural Proteomics
Beyond Structure—Function and Mechanism!
#1 Bottleneck is Psychological!!!

2. High-Throughput Structure Discovery

3. • Synchrotron Radiation
1960
Today
PDB Entries
~20500 1 Mb Hb
Lyso
• Molecular Biology
• Detectors (IP, CCD)
• Software
• Synchrotron Radiation

4. 1999 Protein Structure Initiative (PSI)
9 National Centers with the mission to solve
10,000 structures in the next 10 years (5+5).
~20,500 Structures
4,000 unique (<90%)
1,500 Families
~700 distinct folds
PDB contains:
Vastly increase # of
unique structures and
families
Approach 2-3K distinct
folds in Nature
***
10K new structures:
(if selected from distinct families)

5. NY Structural Genomics Research Consortium
Albert Einstein College of Medicine
Brookhaven National Laboratory
Columbia University
Weill Medical College of Cornell University
UCSF
Structural GenomiX Inc.***

6. SUMMARY OF NYSGRC PROGRESS!!
~60 Structures Solved
~2500 Models
~45% of ALL Structures at X9A/B
Goal: structures/yr.

7. Swaminathan, et al.

9. • Mevalonate pyrophosphate decarboxylase
~3,000 Å2 interface
Burley, et al.

10. MPD Sequence and Secondary Structure Alignment
white green
30% %

11. Detection of a Conserved Cleft
Dimer
Interface
white green
30% %

12. Opportunities/Target Selection:
Functional Studies:
Kinases (122)
Phosphatases (~30)
Regulatory Domains
Nucleic acid modifying Enzymes
Biologically Driven:
Neurobiology
Motile Processes
Immunology
Pathogens and Virulence Factors
Individual Pieces Fit into the Larger Complex Puzzle

13. function with putative essential roles
Systematic examination of essential genes in pathogens:
259 ORFs of unknown
function with putative
essential roles

14. Cell Motility/Actin Cytoskeleton

15. Las17 Arp2/3 Pan1 Abp1 Ent1 Ent2 End3 Yap180 Yor284w Srv2 Rvs167 Cla4
JCB 154, (2001)
Ent1
Ent2
End3
Yap180
Yor284w
Srv2
Rvs167
Cla4
Ynl094w
Sla1
Yhr133c
Ypl246c
Arp2/3
Pan1
Abp1
Las17

16. ABP1
??????
Cofilin Homology
SH3
Strokopytov, et al

17. Srv2/Cyclase Associated Protein (CAP)
PPPPP 1
168
370
526
Adenyl Cyclase
SH3
Actin/Oligomerization
Fedorov, et al.

18. Las17 Arp2/3 Pan1 Abp1 Ypl246c Sla1 Yhr133c Rvs167 Yap180 Ynl094w Srv2?
Aip1
Ent1
Ent2
End3
Yap180
Yor284w
Srv2
Rvs167
Cla4
Ynl094w
Sla1
Yhr133c
Ypl246c
Pan1
Abp1
Las17
Arp2/3

19. Aip1 Nine WD40 Motifs

20. 1.6 Å: Solved in 12 Hours!
Vorobiev, et al.

21. ABP1 ? ? ? ABP1 ABP1 Arp2/3 CAP CAP CAP F - Actin C N C N C N N N C C
Cofilin HD ? ? ? F -
Actin
Cofilin HD
Cofilin HD
Cofilin HD
Cofilin
Cofilin
Cofilin
Cofilin HD HD HD HD
ABP1
ABP1
Arp2/3
SH3
SH3
SH3
SH3 C N PP C N PP
CAP PP PP
SH3
SH3 C N N N PP PP
CAP
CAP
Cofilin C C C C C N N N N
Aip1
Adenylate
Adenylate
Cyclase
Cyclase
Actin
Actin
Actin
Ras
Ras

22. High-Throughput Complex Analysis

23. Tight, stoichiometric
complexes
Rout & Akey; 98

24. New Crystallization Platforms INDUSTRIAL SCALE PROCESS!
Protein limited:
mg from 10s of liters of yeast
New Crystallization Platforms
and/or
INDUSTRIAL SCALE PROCESS!
Absolute requirement for dedicated
Synchrotron access!!!

25. RNAP-DNA Complex ~6.5 Å
Darst, et al.
Tubulin-Stathmin ~4.0 Å
Knossow, et al.
F1F0 ATP Synthase ~4.0 Å
Leslie, Walker, et al.

26. Range of Resolutions High Resolution (X-ray) 3.0 Å or better
Intermediate Resolution (X-ray/EM)
4-10 Å
Modest Resolution (EM)
10-30 Å

27. Footprinting of
Partial Complexes

28. #1 Bottleneck is Psychological!!!
Structural Biology in the 3rd Millennium
Structural Genomics
Structural Proteomics
In vivo structure function relationships
#1 Bottleneck is Psychological!!!