Computational Exploration of Metabolic Networks with Pathway Tools Part 1: Overview & Representations Suzanne Paley Bioinformatics Research Group SRI International

Computational Exploration of

Metabolic Networks with Pathway ToolsPart 1: Overview & Representations

Suzanne PaleyBioinformatics Research Group

SRI International

[email protected]://BioCyc.org/

SRI InternationalBioinformatics

Motivation: Theories of Cellular Function Too Large for One Mind to Grasp Example: E. coli metabolic network

160 pathways involving 744 reactions and 791 substrates Example: E. coli genetic network

Control by 97 transcription factors of 1174 genes in 630 transcription units

Past solutions: Partition theories across multiple minds Encode theories in natural-language text

We cannot compute with theories in those forms Evaluate theories for consistency with new data: microarrays Refine theories with respect to new data Compare theories describing different organisms

SRI InternationalBioinformaticsSolution:

Biological Knowledge Bases

Store biological knowledge and theories in computers in a declarative form

Amenable to computational analysis and generative user interfaces

Establish ongoing efforts to curate (maintain, refine, embellish) these knowledge bases

A high quality comprehensive knowledge base enables us to ask and answer important new questions

SRI InternationalBioinformaticsTerminology

Model Organism Database (MOD) – DB describing genome and other information about an organism

Pathway/Genome Database (PGDB) – MOD that combines information about

Pathways, reactions, substrates Enzymes, transporters Genes, replicons Transcription factors, promoters,

operons, DNA binding sites

BioCyc – Collection of 15 PGDBs at BioCyc.org

EcoCyc, AgroCyc, HumanCyc

SRI InternationalBioinformaticsPathway Tools Software

PathoLogic Prediction of metabolic network from genome Computational creation of new Pathway/Genome Databases

Pathway/Genome Editors Distributed curation of genome annotations Distributed object database system Interactive editing tools

Pathway/Genome Navigator WWW publishing of PGDBs Graphic depictions of pathways, chromosomes, operons Analysis operations

Pathway visualization of gene-expression data Global comparisons of metabolic networks

SRI InternationalBioinformaticsPathway Tools Software

Pathway/ Genome Databases

Pathway/GenomeNavigator

PathoLogic Pathway

Predictor

Pathway/GenomeEditors

SRI InternationalBioinformaticsPathway/Genome Database

Chromosomes,Plasmids

Genes

Proteins

Reactions

Pathways

Compounds

CELL

Operons,Promoters,DNA Binding Sites

SRI InternationalBioinformaticsPathway Tools Algorithms

Visualization and editing tools for following datatypes

Full Metabolic Map Paint gene expression data on metabolic network;

compare metabolic networksPathways

Pathway predictionReactions

Balance checkerCompounds

Chemical substructure comparisonEnzymes, Transporters, Transcription FactorsGenesChromosomesOperons

Operon prediction; visualize genetic network

SRI InternationalBioinformaticsDefinitions

Chemical reactions interconvert chemical compounds

An enzyme is a protein that accelerates chemical reactions

A pathway is a linked set of reactions Often regulated as a unit

A conceptual unit of cell’s biochemical machine

A + B C + D

A C E











SRI InternationalBioinformaticsOperations of the

Metabolic Overview

Find pathways, compounds

Find reactions By enzyme name, EC number, substrates, modulation All with isozymes All occurring in multiple pathways By EC class, pathway class

Find genes By name, gene class All regulated by transcriptional regulator protein

SRI InternationalBioinformaticsMetabolic Overview Queries

Species comparison Highlight reactions that are

Shared/not-shared with Any-one/All-of A specified set of species

Overlay expression data Colors reflects expression level and are user-configurable Can show single experiment or animated time series

SRI InternationalBioinformaticsEcoCyc Project

E. coli Encyclopedia Model-Organism Database for E. coli Began in 1992 as collaboration between Karp and Riley Over 3500 literature citations

Collaborative development via Internet Karp (SRI) -- Bioinformatics architect John Ingraham -- Advisor (SRI) Metabolic pathways Saier (UCSD) and Paulsen (TIGR)-- Transport Collado (UNAM)-- Regulation of gene expression

Ontology: 1000 biological classes Database content: 17,700 instances


EcoCyc = E.coli Dataset + Pathway/Genome

Navigator

Genes: 4,393

Proteins: 4,273

Reactions: 2,760

Pathways: 165

Compounds: 774

http://BioCyc.org/

Transcription Units: 724 Factors: 110

Enzymes: 914Transporters: 162

Promoters: 812TransFac Sites: 956

Citations: 3,508

SRI InternationalBioinformaticsMetaCyc: Metabolic

Encyclopedia

Nonredundant metabolic pathway database Describe a representative sample of every experimentally

determined metabolic pathway

Literature-based DB with extensive references and commentary

Pathways, reactions, enzymes, substrates 460 pathways, 1267 enzymes, 4294 reactions

172 E. coli pathways, 2735 citations Nucleic Acids Research 30:59-61 2002.

Jointly developed by SRI and Carnegie Institution New focus on plant pathways

SRI InternationalBioinformaticsMetaCyc Data

MetaCyc contains one DB object for each distinct pathway

Distinct in terms of reaction steps Each pathway labeled with species it occurs in

MetaCyc pathways are experimentally determined

4218 reactions in MetaCyc 401 lack EC numbers

SRI InternationalBioinformaticsMetaCyc Enzyme Data

Reaction(s) catalyzedAlternative substratesCofactors / prosthetic groupsActivators and inhibitorsSubunit structureMolecular weight, pIComment, literature citationsSpecies

SRI InternationalBioinformaticsMetaCyc Frequent Organisms

Escherichia coli 156

Arabidopsis thaliana 47

Homo sapiens 30

Pseudomonas 21

Bacillus subtilis 20

Salmonella typhimurium 20

Sulfolobus solfataricus 18

Pseudomonas putida 14

Saccharomyces cerevisiae 14

Haemophilus influenzae 13

Glycine max 11

Deinococcus radiourans 10

SRI InternationalBioinformaticsEcoCyc and MetaCyc

Review level databasesData derived primarily from biomedical literature

Manual entry by staff curators Updates by staff curators only

Data validation Consistency constraints Lisp programs that verify other semantic relationships

Unbalanced chemical reactions

SRI InternationalBioinformaticsComputationally-Derived PGDBs

Pathway/GenomeDatabase

Annotated GenomicSequence

Genes/ORFs

Gene Products

DNA Sequences

Reactions

Pathways

Compounds

Multi-organism PathwayDatabase (MetaCyc)

PathoLogic Software

Integrates genome and pathway data to identify

putative metabolic networks

Genomic Map

Genes

Gene Products

Reactions

Pathways

Compounds

SRI InternationalBioinformaticsPathoLogic Input/Output

Inputs: File listing genetic elements

http://bioinformatics.ai.sri.com/ptools/genetic-elements.dat Files containing DNA sequence for each genetic element Files containing annotation for each genetic element MetaCyc database

Output: Pathway/genome database for the subject organism Directory tree for the subject organism Reports that summarize:

Evidence contained in the input genome for the presence of reference pathways

Reactions missing from inferred pathways

SRI InternationalBioinformaticsPathoLogic Functionality

Initialize schema for new PGDBTransform existing genome to PGDB formInfer metabolic pathways and store in PGDBInfer operons and store in PGDBAssist user with manual tasks

Assign enzymes to reactions they catalyze Identify false-positive pathway predictions Build protein complexes from monomers Assemble Overview diagram

SRI InternationalBioinformaticsBioCyc Collection of

Pathway/Genome DBs

Literature-based Datasets:Escherichia coli (EcoCyc) MetaCyc

PGDBs at other sites:Arabidopsis thaliana (TAIR)Methanococcus jannaschii (EBI)Saccharomyces cerevisiae (SGD)Synechocystis PCC6803

Computationally-derived datasets:Agrobacterium tumefaciensCaulobacter crescentusChlamydia trachomatisBacillus subtilisHelicobacter pyloriHaemophilus influenzaeHomo sapiensMycobacterium tuberculosis RvH37Mycobacterium tuberculosis CDC1551Mycoplasma pneumoniaPseudomonas aeruginosaTreponema pallidumVibrio cholerae

Yellow = Open Database

http://BioCyc.org/


HumanCyc: Human Metabolic PathwayDatabase PGDB of human metabolic pathways built using PathoLogic Contains information on 28,700 genes, their products, and the

metabolic reactions and pathways they catalyze (no signalling pathways)

Chromosome and contigs from Ensembl Human genetic loci from LocusLink

Mitochondrion data from GenBank Ensembl and LocusLink gene entries were merged to eliminate

redundancies where possible. Contains links to human genome web sites Plan to hire one curator to refine and curate with respect to literature

over a 2 year period Remove false-positive predictions Insert known pathways missed by PathoLogic Add comments and citations from pathways and enzymes to the literature Add enzyme activators, inhibitors, cofactors, tissue information

Funded by commercial consortium

SRI InternationalBioinformaticsBioCyc and Pathway Tools

Availability

WWW BioCyc freely available to all BioCyc.org Six BioCyc DBs openly available to all

BioCyc DBs freely available to non-profits Flatfiles downloadable from BioCyc.org Binary executable:

Sun UltraSparc-170 w/ 64MB memory PC, 400MHz CPU, 64MB memory, Windows-98 or newer

PerlCyc API

Pathway Tools freely available to non-profits

SRI InternationalBioinformaticsInformation Sources

Pathway Tools User’s Guide aic-export/ecocyc/genopath/released/doc/userguide1.pdf

Pathway/Genome Navigator Appendix A: Guide to the Pathway Tools Schema

aic-export/ecocyc/genopath/released/doc/userguide2.pdf PathoLogic, Editing Tools

Pathway Tools Web Site http://bioinformatics.ai.sri.com/ptools/ Publications, programming examples, etc.

Pathway Tools Tutorial http://bioinformatics.ai.sri.com/ptools/tutorial/

SRI InternationalBioinformaticsPathway Tools Implementation

Details

Allegro Common LispSun and PC platforms

Ocelot object database

250,000 lines of code

Lisp-based WWW server at BioCyc.org Manages 15 PGDBs

SRI InternationalBioinformaticsFrame Data Model

Frame Data Model -- organizational structure for a PGDB

Knowledge base (KB, Database, DB)

Frames

Slots

SRI InternationalBioinformaticsKnowledge Base

Collection of frames and their associated slots, values, facets, and annotations

AKA: Database, PGDB

Can be stored within An Oracle DB A disk file A Pathway Tools binary program

SRI InternationalBioinformaticsFrames

Entities with which facts are associated

Kinds of frames: Classes: Genes, Pathways, Biosynthetic Pathways Instances (objects): trpA, TCA cycle

Classes: Superclass(es) Subclass(es) Instance(s)

A symbolic frame name (id, key) uniquely identifies each frame

SRI InternationalBioinformaticsSlots

Encode attributes/properties of a frame Integer, real number, string

Represent relationships between frames The value of a slot is the identifier of another frame

Every slot is described by a “slot frame” in a KB that defines meta information about that slot

SRI InternationalBioinformaticsProperties of Slots

Number of values Single valued Multivalued: sets, bags

Slot values Any LISP object: Integer, real, string, symbol (frame name)

Slotunits define properties of slots: datatypes, classes, constraints

Two slots are inverses if they encode opposite relationships

Slot Product in class Genes Slot Gene in class Polypeptides

SRI InternationalBioinformaticsPathway Tools Ontology

1064 classes Main classes such as:

Pathways, Reactions, Compounds, Macromolecules, Proteins, Replicons, DNA-Segments (Genes, Operons, Promoters)

Taxonomies for Pathways, Reactions, Compounds

205 slots Meta-data: Creator, Creation-Date Comment, Citations, Common-Name, Synonyms Attributes: Molecular-Weight, DNA-Footprint-Size Relationships: Catalyzes, Component-Of, Product

Classes, instances, slots all stored side by side in DBMS, share a single namespace

SRI InternationalBioinformaticsSlot Links from Gene to

Pathway Frame

Sdh-flavo Sdh-Fe-S Sdh-membrane-1 Sdh-membrane-2

sdhA sdhB sdhC sdhD

succinate + FAD = fumarate + FADH2

Enzymatic-reaction

Succinate dehydrogenase

TCA Cycle

product

component-of

catalyzes

reaction

in-pathway

Chrom

succinate

FAD

fumarate

FADH2

left

right


Enzymatic-reaction frame stores properties of pairing between enzyme and reaction

Sdh-flavo Sdh-Fe-S Sdh-membrane-1 Sdh-membrane-2

sdhA sdhB sdhC sdhD

Succinate + FAD = fumarate + FADH2

Enzymatic-reaction

Succinate dehydrogenase

TCA Cycle

EC#Keq

CofactorsInhibitors

Molecular wtpI

Left-end-position

SRI InternationalBioinformaticsMonofunctional Monomer

Gene

Reaction

Enzymatic-reaction

Monomer

Pathway

SRI InternationalBioinformaticsBifunctional Monomer

Gene

Reaction

Enzymatic-reaction

Monomer

Pathway

Reaction

Enzymatic-reaction

SRI InternationalBioinformaticsMonofunctional Multimer

Monomer Monomer Monomer Monomer

Gene Gene Gene Gene

Reaction

Enzymatic-reaction

Multimer

Pathway

SRI InternationalBioinformaticsPathway and Substrates

Reactant-1

Reaction

Pathway

ReactionReactionReaction

Reactant-2

Product-2

Product-1

in-pathwayleft

right

SRI InternationalBioinformaticsGenetic Network Representation

Describe biological entities involved in control of transcription initiation

Promoters, operators, transcription factors, operons, terminators

Describe molecular interactions among these entities

Modulation of transcription factor activity Binding of transcription factors to DNA binding sites Effects on transcription initiation

SRI InternationalBioinformaticsOntology for

Transcriptional Regulation

One DB object defined for each biological entity and for each molecular interaction

site001

pro001

trpE

trpD

trpC

trpB

trpA

trpL

Int002 RpoSig70

TrpR*trpInt001

trpLEDCBA

trp

apoTrpRComplexation reaction

Int001 (binding of TrpR*trp to site001) inhibits Int002 (binding of RNA Polymerase to promoter) and consequently prevents transcription

of genes in transcription unit.

SRI InternationalBioinformaticsPrinciple Classes

Class names are capitalized, plural

Genetic-Elements, with subclasses: Chromosomes Plasmids

GenesTranscription-UnitsRNAsProteins, with subclasses:

Polypeptides Protein-Complexes

SRI InternationalBioinformaticsPrinciple Classes

Reactions, with subclasses: Transport-Reactions

Enzymatic-Reactions

Pathways

Compounds-And-Elements

SRI InternationalBioinformaticsSlots in Multiple Classes

Common-NameSynonymsNames (computed as union of Common-Name,

Synonyms)

CommentCitations

DB-Links

SRI InternationalBioinformaticsGenes Slots

ChromosomeLeft-End-PositionRight-End-PositionCentisome-PositionTranscription-DirectionProduct

SRI InternationalBioinformaticsProteins Slots

Molecular-Weight-SeqMolecular-Weight-Exp

pILocations

Modified-FormUnmodified-Form

Component-Of

SRI InternationalBioinformaticsPolypeptides Slots

Gene

SRI InternationalBioinformaticsProtein-Complexes Slots

Components

SRI InternationalBioinformaticsReactions Slots

EC-Number

Left, RightSubstrates (computed as union of Left, Right)Enzymatic-Reaction

DeltaG0

Spontaneous?

SRI InternationalBioinformaticsEnzymatic-Reactions Slots

EnzymeReactionActivatorsInhibitorsPhysiologically-RelevantCofactorsProsthetic-GroupsAlternative-SubstratesAlternative-CofactorsReaction-direction

SRI InternationalBioinformaticsPathways Slots

Reaction-ListPredecessorsPrimaries

Documents

Computational Exploration of Metabolic Networks with Pathway Tools Part 1: Overview & Representations Suzanne Paley Bioinformatics Research Group SRI International