The evolution and structural anatomy of small molecule metabolism pathways in Escherichia coli. Of...

The evolution and structural anatomy of

small molecule metabolism pathways in

Escherichia coli.Of Pathways and ProteinsStuart Rison and Sarah

Teichmann

Questions

• How are homologous proteins (enzymes) distributed in E. coli metabolism?

• How does this distribution fit with theories of pathway evolution?

Pathway evolution

• Norman Horowitz, 1945: ‘On the evolution of biochemical syntheses’, Proc. Nat. Acc. Sci. 31:153-157.

“Retrograde evolution”• Roy Jensen, 1976: ‘Enzyme

recruitment in evolution of new function’, Ann. Rev. Microbiol 30:409-425.

“Patchwork evolution”

Retrograde evolution

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Jensen, 1976: Substrate ambiguity

• ‘Original pool’ of unregulated and enzymatically versatile proteins

• Enzymes recruited from the pool• Ad hoc pathways• Gene duplication and

specialisation leads to regulated, specific and efficient pathways

Patchwork evolution

Why E. coli?

• An extensively studied model organism

• Complete genome available• Most Small Molecule Metabolism

pathways well known and empirically characterised

• A manageable size

• Good associated databases

Strategy

• Identify all SMM proteins and the pathway(s) in which they belong

• Detect homologous proteins by structure or sequence

• Combine these data to analyse homologous protein distribution in SMM

Methods

E. coli

IMPALA

HMM

-BLAST

-BLAST (>75aa)

+ =

EvolutionaryRelationships

PathwaysProteins

566 SMM proteins

442 proteins assignedto 1+ families (78%)

124 unassignedproteins

169 PDB-D families 31 ‘sequence’ domainfamilies

200 domain families

Domain assignments

Chemistry andclose substrateChemistry and

substrate

Glycogen Catabolism

malQ

malS

malZ

pgmmalP

glgPamyA

-amylase, 3.2.1.1

phosphoglucomutase, 5.4.2.2

amylomaltase, 2.4.1.25

-amylase, 3.2.1.1 glycogen phosphorylase

malodextrin phosphorylase

malodextrin glucosidase

-glucosyltransferase

Phosphoglucomutase

-amylase, C-term

Glycosyltransferases

Domains

Internal duplicationIsozymes

Duplications Across Pathways

• 110 out of 200 families occur in more than one pathway

• Can exhibit conservation of chemistry, shared cofactor or minor substrate similarity

• 36 families have close conservation of EC number (Chemistry conserved)

• 74 families conserve 1 or no EC number; 11 are cofactor-binding families (cofactor, minor substrate)

Duplications within and across Pathways

• 710 domains in 200 families510 domains have arisen by

duplication

• 232 duplications within pathways to 278 duplications across pathways

(Assumption: duplication within pathways wherever possible.)

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10

20

30

40

50

60

70

80

90

100

Num

ber o

f pro

tein

s in

volv

ed

Cofactor ChemistryIsozymesInternal

Dup.Substrate

Type of conservation

Conclusion: Structural Anatomy

• 710 domains in 442 proteins of the 566 proteins in E. coli SMM pathways

• 200 families (3.5 members/family)

• Most sizeable families are distributed in several pathways

Conclusion: Recruitment and Conservation

• Duplications have taken place between and within pathways to roughly the same degree

• Duplications occur within most longer pathways:– Isozymes, internal duplications and co-

factor binding most common– Chemistry common– Conservation of substrate binding with

modified chemistry is rare

Conclusions: Pathway evolution

• Data support a “patchwork evolution” model

• Little evidence of “retrograde evolution”

Conclusions: hum…

• Recruitment, duplication and evolution of enzymes are constantly taking place so we are always observing a dynamic system

• Likely to be other evolutionary mechanisms and combinations thereof

Future

• Identification and analysis of novel pathway duplication events

• Focus on order in pathways:– Stepwise analysis– Doublet/triplet analysis

• Analysis domain combination in SMM

Acknowledgements

• Sarah A. Teichmann, Dept. Biochemistry, University College London

• Janet M. Thornton, David Lee, Dept. Crystallography, Birkbeck College and Dept. Biochemistry, University College London

• Monica Riley, Alida Pelegrini-Toole, Marine Biology Laboratory, Woods Hole, USA

• Cyrus Chothia, Julian Gough, MRC Laboratory of Molecular Biology, Cambridge, UK