Systems Biology – Connecting Genotype to Phenotype

Allosteric Interactions and Bifunctionality Make the Response of Glutamine Synthetase Cascade System of

Escherichia coli Robust and Ultrasensitive, Vivek K. Mutalik, Parag Shah and K V Venkatesh, The Journal of

Biological chemistry, [278] (29), 26327-26332, 2003

K. V. VenkateshDept. of Chemical Engineering

IIT BombayIRCC Presentation, August 4, 2010

Genotype to PhenotypeGenotype to Phenotype

GenomeGenome

TranscriptomeTranscriptome

ProteomeProteome

MetabalomeMetabalome

PhenotypePhenotype

Presence of genome does not ensure a phenotypeIt requires a specific state in the hierarchical chain.

Central Dogma of BiologyCentral Dogma of Biology

Firefly GlowsTransgenic Plant made to Glow

Phenotype

Phosphorous releaseusing ATP Metabolic Reaction

Catalyzed by Luciferase Enzyme Protein

Luciferase gene decoded RNA

Luciferase Gene Gene

Descriptional ScienceDescriptional Science

Historically Biology has been a descriptional Historically Biology has been a descriptional sciencescience

Molecular Biology has given molecular level Molecular Biology has given molecular level description of the Hierarchical state.description of the Hierarchical state.

Genetic, signaling/protein and metabolic Genetic, signaling/protein and metabolic networks are the result of reductionism of networks are the result of reductionism of Molecular Biology.Molecular Biology.

Bioinformatics has added more information to Bioinformatics has added more information to this approach.this approach.

Resulting in a highly interconnected network!Resulting in a highly interconnected network!

JCB, 161, 463-464, 2003,

Quantification of SystemsQuantification of Systems

Engineering systems are quantified to a Engineering systems are quantified to a level that they are designed, optimized level that they are designed, optimized and optimally operated.and optimally operated.

In the last decade Biology has rapidly In the last decade Biology has rapidly evolved to be a quantitative science.evolved to be a quantitative science.

Principles of system science can be Principles of system science can be applied to biology: applied to biology: Systems BiologySystems Biology

System Analysis and QuantificationSystem Analysis and Quantification

DesignDesign

OperationOperation

ControlControl

Fault DiagnosisFault Diagnosis

Evolve/AdaptEvolve/Adapt

1250 computers Hundreds of feedback loops Millions of components Design Manual Available

Bottom-up Design of a Complex SystemBottom-up Design of a Complex System

Design in Nature: Top-Down Design in Nature: Top-Down Approach (Approach (Escherichia coliEscherichia coli))

About 4400 genes

Connectivity between genes, mRNA, proteins & metabolites

Thousands of feedback loops

No design principles available

No computation – control & sensing achieved through interactions of biomolecules

Complexity in Engineered and Complexity in Engineered and Natural SystemsNatural Systems

Non-linear dynamicsNon-linear dynamicsMultiple feedback loopsMultiple feedback loopsMultiple interactionsMultiple interactionsCascade structuresCascade structuresFeed forward loopsFeed forward loopsInteractions between modulesInteractions between modulesTimescale separationTimescale separation

Resulting in a Complex system

Variable ObjectiveVariable Objective

RobustnessRobustness

SensitiveSensitive

AdaptiveAdaptive

ControllableControllable

Respond to uncertain environmentRespond to uncertain environment

Noise mitigationNoise mitigation

Memory and learning systemsMemory and learning systems

Nitrogen Assimilation in E. coli

Glucose

Pyruvate

TCA Cycle

AKG Glutamate Glutamine

Ammonia Ammonia

Glutamine Synthetase

Presence of Ammonia : GS is requiredPresence of Glutamine: GS is not required

Either Synthesis can be stopped oractivity can be inhibited

Glutamine Synthetase system

Glutamate Glutamine

ATP + NH3 ADP

NADPH + 2KGNADP

GS

Glutamate synthase

Amino acids

AMP, CTP

Feedback inhibitionGenetic Regulation

Covalent modification

GS is essential for nitrogen assimilation under nitrogen starvation conditions. Adenylylated GS is catalytically inactive

NRI-P Dimer

gln ANR

I-P

NR

I-P

gln L gln G

GS NRII NRI

NR

I-P

NR

I-P

Connecting nutritional signal to gene activation

Results

2KG (µM)a = 5b = 30c = 100d = 300e = 600f = 900

Prof. Ninfa’s LabIn the Department of Biological ChemistryUniversity of Michiganhas experimentally demonstrated our predictions

Results Quantification of Response of GS bicyclic cascade,

NRII –NRI two component system and Ntr operon

Synthesis and activity of GS is independent of all component concentrations and is only a function of Glut and 2KG

Coordination of assimilation of nitrogen & carbon seems to exhibit robustness

( )( ) ( )

( )( ) ( )

2 100

1 2 max2 2 100 100

1 2

K K

Active GS GSK GLN K GLN

� �� �= � �� �

+ +� �� �� �� �

SummarySummaryQuantification is key to Systems BiologyQuantification is key to Systems BiologyDesign principles inherent in Biological structureDesign principles inherent in Biological structureSystem analysis to elucidate role of structure and System analysis to elucidate role of structure and connectivityconnectivityOperational characteristics of networksOperational characteristics of networksFault-diagnosis for characterization of disease state Fault-diagnosis for characterization of disease state Possible sites in the network towards drugs targetsPossible sites in the network towards drugs targetsModular analysis towards Modular analysis towards in silicoin silico cellular cellular representationrepresentationScope for evaluating and using biological structures Scope for evaluating and using biological structures which have interconnected networkswhich have interconnected networksDesign and Engineering of Artificial Biological systemsDesign and Engineering of Artificial Biological systems

Acknowledgements

Vivek Mutalik, Senior Scientist, BioFAB, University of California Berkeley, USA

Parag Shah, Doctoral Candidate, University of Colorado, Boulder, USA.

Prof. Jayadeva Bhat – for all the insights

Prof. Sharad Bhartiya – for system engineering approaches