“Creating the right chemistry” in Pathways Master Seminar Fall 2006 M.A. Ott (CMBI)

Main concept ”Creating the right chemistry” in Pathways M.A. Ott Enzymes don’t do new chemistry; they do ‘old’ chemistry faster Enzymes don’t perform magic; they perform known (organic) chemistry

Contents ”Creating the right chemistry” in Pathways M.A. Ott Enzymes and Metabolites Metabolic Pathways and Reactions Representing Chemistry BioMeta Database

Ongoing progess in “omics” ”Creating the right chemistry” in Pathways M.A. Ott Genomics…going strong Transcriptomics…good progress Proteomics…some progress Metabolomics…less progress Systems biology…not in ten years

Metabolite and pathway databases ”Creating the right chemistry” in Pathways M.A. Ott Kyoto Encyclopedia of Genes and Genomes BRENDA enzyme database Chemical Entities of Biological Interest ExPASy biochemical pathway EMP enzymology and metabolism data MetaCyc metabolic pathways EcoCyc E. coli genome and metabolism PathDB biological pathways and networks B-NET - Biochemical Network BioPath database ERGO comprehensive genome analysis Reactome biological processes IUBMB enzymes

Metabolism and friends ”Creating the right chemistry” in Pathways M.A. Ott Metabolism: The reactions in organisms Anabolism: Synthesis of larger biomolecules from smaller ones » Usually requires energy input Catabolism: Break down of larger molecules into smaller ones » Usually releases energy

Enzymes ”Creating the right chemistry” in Pathways M.A. Ott Virtually all metabolic reactions require enzymatic catalysis Rate enhancement factor can be up to Enzymes are highly reaction-specific, catalyzing only one conversion Enzymes are substrate-specific, limiting their action to only one compound or to related compounds

The metabolic network ”Creating the right chemistry” in Pathways M.A. Ott

Some metabolites ”Creating the right chemistry” in Pathways M.A. Ott “Housekeeping” metabolites: Water, NADH, ATP, Phosphate, … End products of pathways: Vitamin B 12, Urea, Penicillin, Ecdysone, … Intermediates in pathways: Glucose-6-P, Lanosterol, Precorrins, Shikimate, …

Housekeeping metabolites: ATP / ADP ”Creating the right chemistry” in Pathways M.A. Ott Full structures: Shorthand:

Housekeeping metabolites: NADH / NAD + ”Creating the right chemistry” in Pathways M.A. Ott Full structures: Shorthand:

Housekeeping metabolites and chemical distance ”Creating the right chemistry” in Pathways M.A. Ott What is the chemical distance between B and D?

Typical housekeeping metabolites ”Creating the right chemistry” in Pathways M.A. Ott Water Ammonia Phosphate ATP, ADP NADH, NAD + NADPH, NADP + FAD, FADH2 FMN, FMNH2 SAM, SAH Coenzyme A Glutamate/2-Oxoglutarate Phosphorylation Reduction/oxidation Methyl group transfer Acyl group transfer Amino/ketone group exchange

Cholesterol biosynthesis outline Pathways and Molecules M.A. Ott

Cholesterol biosynthesis from squalene Pathways and Molecules M.A. Ott

Metabolic disorders Pathways and Molecules M.A. Ott Inability to synthesize compounds: Absence of important compounds Inability to degrade compounds: Accumulation of undesired waste products E.g., in phenylketonuria

Phenylketonuria (error in phenylalanine catabolism) Pathways and Molecules M.A. Ott

The BioMeta Database ”Creating the right chemistry” in Pathways M.A. Ott

The BioMeta Database ”Creating the right chemistry” in Pathways M.A. Ott

BioMeta database structure ”Creating the right chemistry” in Pathways M.A. Ott

BioMeta database structure ”Creating the right chemistry” in Pathways M.A. Ott Molecules Reaction 1: Mol_1 + Mol_2  Mol_3 + Mol_4 Reaction 2: Mol_2 + Mol_3  Mol_5 + Mol_6 Reaction 3: 2 Mol_6  Mol_7 R-M Links Reactions

Sample database query ”Creating the right chemistry” in Pathways M.A. Ott ec_nr | enzyme_name | rxn_id |rev | role |stoich| name | oxalate oxidase | MR | ir | s | 1 | Oxalic acid,, |,, |,, |,, | s | 1 | O2,, |,, |,, |,, | p | 1 | H2O2,, |,, |,, |,, | p | 2 | CO2

Molecular structure identification ”Creating the right chemistry” in Pathways M.A. Ott Mesomerism (resonance forms) Tautomerism (variable positions of hydrogens) Other “chemical” flexibility Protonation state

Molecular structures and reactions ”Creating the right chemistry” in Pathways M.A. Ott Reactions:balancing atoms & charges adding direction/reversibility Molecules:adding stereochemical configurations “canonicalizing” tautomeric form No stereochemistry Phosphorus imbalance Direction Carbon imbalance

And now what’s really going on… ”Creating the right chemistry” in Pathways M.A. Ott

Accuracy required! ”Creating the right chemistry” in Pathways M.A. Ott Without accurate chemical structures and balanced reaction descriptions, a biochemical pathways database is unsuitable for “chemical computation”: Graph-theoretical considerations for creating a network Idem for describing fluxes Docking applications need correct stereochemistry

Validation results (structures) ”Creating the right chemistry” in Pathways M.A. Ott Type of Error# in KEGG# in BioMeta# Corrected Structure missing Valence violation(s) 76 0 Incorrect constitutionunknown 107 Total (constitution) Undefined stereo double bond(s) Invalid sp3 stereocenter(s) Ambiguous sp3 stereocenter(s) 46 0 Undefined sp3 stereocenter(s) Unspecified enantiomer Undefined sp3 stereochemistry Incorrect stereochemistryunknown 69 Total (stereochemistry) Total corrected 1468

Validation results (reaction balance) CountPerc. Total Unknown Balanced Unbalanced Corrected * ”Creating the right chemistry” in Pathways M.A. Ott DifferenceCount [H+] *801 2[H+] *35 H 2 O *57 O98 H2H2 17 H 2 – 2[H+]41 2H 2 17 H 2 – [H+]18 CH 2 30 O 2 – H 2 O17

Some unresolved issues ”Creating the right chemistry” in Pathways M.A. Ott

BioMeta database ”Creating the right chemistry” in Pathways M.A. Ott Based on freely available data (KEGG Ligand database) Augmentation, completion and correction of small-molecule information: - Adding missing stereochemistry to structures - Balancing atoms and charges in reactions - Establishing the reversibility and direction of reactions Search for alternative pathways between metabolites Search for “missing” enzymes

Alternative pathways ”Creating the right chemistry” in Pathways M.A. Ott

BioMeta: things being done ”Creating the right chemistry” in Pathways M.A. Ott Enhancing compound/reaction structures Establishing directionality/reversibility of reactions Distinguishing cofactors/“current”/“external”/ “common” metabolites from ‘real’ metabolites Adding more compound data, e.g. pK a (curve) Increasing functionality of WWW interface

BioMeta: things to do ”Creating the right chemistry” in Pathways M.A. Ott Atom-to-atom correspondence (reaction mapping) Tautomer handling Pathway searches Putative/hypothetical reactions for predicting metabolism

Acknowledgements ”Creating the right chemistry” in Pathways M.A. Ott Felix van Diggelen Gert Vriend CMBI