Strain Engineering Mona Yousofshahi Prof Soha Hassoun Department of Computer Science Prof Kyongbum Lee Chemical amp Biological Engineering Tufts University 1 Production or overproduction by synthetic pathways ID: 1032556
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1. Pathway Identification for Strain EngineeringMona Yousofshahi, Prof. Soha HassounDepartment of Computer ScienceProf. Kyongbum LeeChemical & Biological EngineeringTufts University1
2. Production or overproduction by synthetic pathways 2MotivationBiofuelsAlcoholDieselBioplasticsOrganic plasticsDerived from biomass sources instead of petroleumDrugsAntimalarialAnticancer
3. Pathway identification Identify a coherent set of enzyme-catalyzed reactions from existing databasesIntegration with the host Ensure that the pathway minimally affects growth and other essential functions of the host3Pathway synthesis
4. Probabilistic graph search algorithm based on metabolite connectivityGraph construction begins with a target metabolite and ends in a host Explicitly accounts for cofactorsSearch criteria is metabolite connectivity within the KEGG database:Number of reactions in which a metabolite participatesMore diversity in the search space 4ApproachHostTarget metaboliteDatabase
5. 5Metabolite connectivity in the KEGG databaseP(k) ≃ 3.48 k-2.04
6. Metabolite connectivity:The number of reactions in which a metabolite participatesWeighting of a reaction:Minimum connectivity in a reaction is the bottleneckWR = minimum metabolite connectivity of the metabolites in reaction R (on the side opposite to the parent metabolite)6Calculating reaction probability distribution based on metabolite connectivity6R1R2ABCD
7. Construct the graph recursively starting from the target metaboliteSelect a random reaction based on metabolite connectivitySearch terminationLimit the number of reactionsPerform flux balance analysis on the constructed pathways7Probabilistic graph search algorithmHostTarget metabolite
8. Constructing the tree recursively, starting from the root and by adding all reactions to the treeApplying FBA to rank the constructed pathways8Exhaustive search algorithm
9. Genome-scale model of E. coli (iAF1260) (Feist, Henry et al. 2007) as a hostTarget metabolitesDrug: Isopentenyl diphosphateBiofuels: Biodiesel, Fatty acid methyl esterBiofuel feedstock: TriacylglycerolPolymer: 1, 3-propanediolCompare three search algorithms based on yield resultsProbabilistic, random and exhaustiveYield is defined as the optimal flux of the target metabolite Fixed biomass flux9Test cases
10. Probabilistic algorithmRandom algorithmExhaustive algorithmMetabolite nameNumber of pathwaysMax. YieldNumber of pathwaysMax. YieldNumber of pathwaysMax. YieldIsopentenyl diphosphate111.28141.28151.281,3-Propanediol12.1912.1912.19Biodiesel173.30192.095043.58Fatty acid methyl ester691.25460.7611211.25Triacylglycerol711.94450.4429491.9710ResultsRun times: Exhaustive search for maximum 10 reactions in a pathway: hours Probabilistic and random search: minutes
11. 11Comparing with literature (Martin, Piteral et al. 2003)Identified pathway for isopentenyl diphosphate by probabilistic algorithm:Acetyl-CoA + Acetoacetate (S)-3-Hydroxy-3-methylglutaryl-CoA (R)-Mevalonate (R)-5-Phosphomevalonate (R)-5-Diphosphomevalonate Isopentenyl diphosphate
12. Yield distributionsProbabilistic searchExhaustive search12Triacylglycerol yield distribution
13. 13Number of required iterationsFatty acid methyl ester50 runs for each iteration
14. 14Probabilistic vs. random searchFatty acid methyl ester50 runs for each iteration
15. PathMiner (McShan, Rao et al. 2003) exploring the biochemical state space using a heuristic search based on minimizing the cost of transformationAtom mapping (Blum, Kohlbacher 2008)Optstrain (Pharkya, Burgard et al. August 2004)building a framework for identifying stoichiometrically balanced pathways while maximizing product yieldRequires database curation15Prior works
16. A probabilistic graph search algorithm to identify synthetic pathwaysUsing the notion of the metabolite connectivityDoes not require any database curationReproduce experimentally obtained pathways reported in the literatureFuture work:Integration with the hostGene interactions16Conclusion & future work