Protein evolution proceeds through genetic mechanisms, but selection acts on biological assemblies. I define a protodomain as a minimal independently evolving unit with conserved structure. Protodomain rearrangements have minimal impact on biological assemblies, so they represent a valid evolutionary path through fold space. These slides are from my Candidacy Exam on Jan 28, 2013 at University of California, San Diego. It discusses my current research in Philip Bourne's lab, as well as proposes research for my thesis over the next two years. An audio version is available at http://www.scivee.tv/node/57082

1. FOLLOWING THE EVOLUTION
OF NEW PROTEIN FOLDS VIA
PROTODOMAINS
Spencer Bliven
January 28, 2013
Advancement to Candidacy Exam

3. CATH:
http://www.cathdb.info/browse/bro
wse_hierarchy
http://scop.mrc-lmb.cam.ac.uk/scop/

4. CONTINUITY
Sadreyev, R. I., Kim, B.-H., &Grishin, N. V.
(2009). Discrete-continuous duality of protein
structure space. Current Opinion in Structural
Biology, 19(3), 321–328.
Grishin. J Struct Biol (2001) vol. 134 (2-3) pp. 167-85

5. MODELS OF FOLD SPACE
β
 Orengo, Flores, Taylor, Thorn
ton. Protein Eng (1993) vol. 6 α α/β
(5) pp. 485-500
 Holm and Sander. J Mol Biol
(1993) vol. 233 (1) pp. 123-38 α+β
 Holm and Sander. Science
(1996) vol. 273 (5275) pp. 595-
603
 Shindyalov and Bourne.
Proteins (2000) vol. 38 (3) pp.
247-60
 Hou, Sims, Zhang, Kim.
PNAS (2003) vol. 100 (5) pp.
2386-90
 Taylor. Curr Opin Struct Biol
(2007) vol. 17 (3) pp. 354-61
 Sadreyev et al. Curr Opin
Struct Biol (2009) vol. 19 (3)
pp. 321-8

6. BIG QUESTIONS
 Is fold space discrete or continuous?
 Where do new folds come from?
 What insights can we gain by studying fold
space?

7. DEFINITIONS

8. BIOLOGICAL ASSEMBLIES
Sesbania mosaic
virus [1VAK]
Asymmetric Unit Biological Assembly
Hemoglobin
[1hv4]

9. DOMAINS
 Compact Geometry
 Independently Folding
Non-contiguous domains Multi-chain domains
Squalene-HopeneCyclase Kunitz-type trypsin
[1SQC] inhibitor [1r8o]

10. FOLD
 Group of domains with
 Same major secondary structural elements
 Same mutual orientation
 Same connectivity

11. PROTODOMAINS
 A protodomain is a minimal, independently
evolving protein unit with a conserved
structure.
 Defined through evolution, but usually observed
as structural motif
 Coined by Philippe Youkharibache

12. PROTODOMAINS
 A protodomain is a minimal, independently
evolving protein unit with a conserved structure.
GTP binding
Glyoxalase I from regulator from
Clostridium Thermotoga
acetobutylicum[3 maritima [1VR8]
HDP]
Glyoxalase I in E. Pseudomonas
coli [1F9Z] 1,2-dihydroxy-
naphthalene
dioxygenase
[2EHZ]

13. PROPOSAL

14. SPECIFIC AIMS
1. Improve algorithms to identify conserved
protodomains globally across the PDB.
2. Identify structurally similar and potentially
homologous protodomains across fold space.
3. Integrate protodomain arrangements with
domain and quaternary structure information
to create a parsimonious model of fold evolution
across the tree of life.
4. Apply protodomain principles to understanding
the evolution of specific protein families.

15. AIM 1
 Improve algorithms to identify conserved
protodomains globally across the PDB.
Preliminary Research:
a) Circular Permutation with CE-CP
b) Symmetry with CE-Symm
Proposed Research:
a) Improve CE-Symm algorithm
b) Create algorithms for other types of
protodomain rearrangements
c) Run algorithms globally across the PDB
d) Create non-redundant catalogue of
protodomains

16. CIRCULAR PERMUTATION
 Spencer Bliven and Andreas Prlić. Circular
Permutation in Proteins. PLoSComputBiol (2012)
8(3): e1002445.

17. CIRCULAR PERMUTATION EVOLUTION
Fission & Fusion Permutation by Duplication

18. CE-CP
 A Prlić, S Bliven, P Rose, J Jacobsen, PV Troshin, M Chapman, J
Gao, CH Koh, S Foisy, R Holland, G Rimša, ML Heuer, H.
Brandstätter–Müller, PE Bourne, and S Willis. BioJava: an open-
source framework for bioinformatics in 2012. Bioinformatics (2012).
 http://www.rcsb.org/pdb/workbench/workbench.do
N
C
C
N
Molybdate-binding protein Regulator of G protein
Concanavalin A [1NLS.A]
[1ATG.A] vs. OpuAC signaling 10 [2IHB.A] vs.
vs. Pea Lectin [1RIN.A+B]
[2B4L.A] vaccinia H1-related
phosphatase[1VHR.A]

19. DETECTING CIRCULAR PERMUTATIONS

20. SYMMETRY
Beta Propeller
Goodsell, D. S., & Olson, A. J. (2000).
Structural symmetry and protein function.
Annual Review of Biophysics and Biomolecular
Structure, 29, 105–153.

21. SYMMETRY
 Functionally important FGF-1
3JUT
 Protein evolution (e.g. beta-trefoil)
 DNA binding
 Allosteric regulation
 Cooperativity TATA Binding
Protein
1TGH
 Widespread (19% of proteins)
Hemoglobin
4HHB

22. SYMMETRY EVOLUTION
 Start with perfectly symmetric homomer
 Duplications & Fusions
 Symmetry lost to drift

23. INTERMEDIATES TO BETA-TREFOIL
FGF-1 [3JUT]
Lee, J., &Blaber, M. (2011). Experimental support for the evolution of symmetric protein architecture
from a simple peptide motif. PNAS, 108(1), 126–130.

24. CE-SYMM WISHLIST
 Find alignments for all valid rotations
 Refine alignments based on isomorphism
constraints
 Utilize crystallographic symmetry more
efficiently for biological assemblies Triose Phosphate
Isomerase [8TIM]
 Detect multiple axes of symmetry
5-enol-pyruvyl shikimate-3-phosphate
(EPSP) synthase [1G6S]

25. CE-SYMM
 Andreas Prlić, Spencer E. Bliven, Peter W.
Rose, Philippe Youkharibache, Douglas Myers-
Turnbull, Philip E. Bourne. On Symmetry and
Pseudo-Symmetry in Proteins. In preparation.
FGF-1 [3JUT] AmtB [3C1G]

26. CE-SYMM
FGF-1

27. CE-SYMM
FGF-1

28. ADDITIONAL METHODS FOR DETECTING
PROTODOMAINS
 Changes in Quaternary Structure
 Protodomain searches (Douglas Myers-Turnbull)
 Domain Swapping

29. AIM 2
 Identify structurally similar and potentially
homologous protodomains across fold space.
Preliminary Research
a) All-vs-all comparison of chains & domains
b) Clustering & network analysis
Proposed Research
a) Run all-vs-all comparison of protodomains
b) Build protodomain similarity network
c) Correlate network with existing properties:
ligand binding, symmetry order, enzymatic
activity, and distribution across organisms, etc

30. ALL-VS-ALL STRUCTURAL ALIGNMENT
 Andreas Prlić, Spencer Bliven, Peter W
Rose, Wolfgang F. Bluhm, Chris Bizon, Adam
Godzik, Philip E. Bourne. Precalculated Protein
Structure Alignments at the RCSB PDB website.
Bioinformatics (2010) vol. 26 (23) pp. 2983-2985

31. ALL-VS-ALL STRUCTURAL ALIGNMENT
 Use sequence clustering to get representative
chains with <40% sequence identity (currently
23410)
 Split into domains by SCOP or PDP
 All chains and domains compared using FATCAT
 Use Open Science Grid (OSG)
 Client/Server architecture for aggregating results
…
Scores
…

32. NETWORK FROM TRANSPORTER
CLASSIFICATION DATABASE (TCDB)
Primary Active Transporters
Channels/Pores
Transmembrane Electron Carriers
Group Translocators
…

33. BETA PROPELLERS
Symmetry
C4
C5
C6
C7

34. CROSS-CLASS EXAMPLE
 3GP6.A
 PagP, modifies lipid A
 f.4.1 (transmembrane
beta-barrel)
 1KT6.A
 Retinol-binding
protein
 b.60.1 (Lipocalins)

35. AIM 3
 Integrate protodomain arrangements with domain
and quaternary structure information to create a
parsimonious model of fold evolution across the tree
of life.
Preliminary Research
a) Classification of biological assemblies by quaternary
symmetry & chain stoichiometry
b) Model for evolution via protodomains
Proposed Research
a) Determine the protodomain content of each
biological assembly
b) Identify BAs with conserved protodomain
architecture but different chain architecture, or vice
versa
c) Integrate data with model of protodomain evolution

36. QUATERNARY STRUCTURE
 Find symmetry & pseudosymmetry within
biological assemblies
 Functions at chain level
 Can use various thresholds to determine
stoichiometry (95% sequence, CE alignment, etc)
Rhinovirus 2 [3DPR] GTP Cyclohydrolase I Hemoglobin [4HHB]
I (60,60,60,60,60) [1A8R] D5 (10) C2 (2,2)

37. EVOLUTIONARY MODEL
1. Local Mutation
2. Protodomain fusion
3. Protodomain fission
4. Loss of Interface
5. Gain of Interface
6. New Protodomains

38. CONNECTION TO FOLD SPACE
 Mostly local mutations = continuous regions
 Protodomain creation & rearrangement =
discrete regions
 Identifying evolutionary events allows
quantitative comparison of the frequencies of
each mechanism
 Biologically rather than geometrically motivated

39. AIM 4
 Apply protodomain principles to understanding
the evolution of specific protein families.
 Qualities
 Have good structural coverage
 Contain multiple members with symmetry at either
domain or quaternary structure level.
 Contain circularly permuted members
 Span a diverse set of folds
 Ion Channels
 Beta Propellers
AmtB [3C1G]

40. SODIUM/ASPARTATESYMPORTER FROM
PYROCOCCUSHORIKOSHII(GLTPH)
cytoplasm
Top Side
[2NXW]
Forrest, L. R., Krämer, R., & Ziegler, C. (2011). The structural basis of
secondary active transport mechanisms. Biochimica et
BiophysicaActa, 1807(2), 167–188.

41. CONCLUSIONS
 Biological Assemblies are the functional unit of
structure
 Protodomains can rearrange without modifying
the biological assembly
 Separating changes in biological assembly from
genetic changes can provide evolutionary
perspective on fold space
 Local Changes = Continuous Evolution
 Protodomain rearrangements = Discrete Transitions

42. TIMELINE

43. PUBLICATIONS
 A Prlić, S Bliven, PW Rose, WF Bluhm, C Bizon, A Godzik, PE
Bourne. Precalculated Protein Structure Alignments at the RCSB
PDB website. Bioinformatics (2010) vol. 26 (23) pp. 2983-2985
 Spencer Bliven and Andreas Prlić. Circular Permutation in
Proteins. PLoSComputBiol (2012) 8(3): e1002445.
 A Prlić, S Bliven, P Rose, J Jacobsen, PV Troshin, M Chapman, J
Gao, CH Koh, S Foisy, R Holland, G Rimša, ML Heuer, H
Brandstätter–Müller, PE Bourne, and S Willis. BioJava: an open-
source framework for bioinformatics in 2012. Bioinformatics
(2012).
Intended:
 CE-Symm method
 Evolutionary model & examples of protodomain evolution
 Structural similarity network analysis
 Use of model for specific protein family

44. ACKNOWLEDGMENTS
Committee Collaborators
Philip Bourne Philippe Youkharibache
Milton H. Saier Jean-Pierre Changeux
Russell F. Doolittle BiojavaContributors
Michael K. Gilson
Adam Godzik
The lovely Christine
Bourne Lab/PDB Bliven
Andreas Prlić
Peter Rose
Douglas Myers-Turnbull
Lab & PDB members

45. This work is licensed under a Creative Commons Attribution-ShareAlike 3.0
Unported License.