1. Stochastic modelling of fatty acid biosynthesis Elahe Radmaneshfar 17th July 2015 Rothamsted

2. De novo synthesis and biosynthesis of FA Plastid Acyl-CoA pool 16:0-CoA 18:0-CoA 18:1-CoA FA Synthesis Endoplasmic Reticulum LC-PUFA biosynthesis pathway Kennedy pathway TAG

3. Aims of the model Developing a predictive mathematical model which: – can help us understand the mechanism of fatty acid biosynthesis. – Suggest possibility to improve the production of fatty acid of our interest (Omega-3)

4. Challenges for building the model 1.Representation of fatty acids 1 1 1 1 1 1 1 1 1 1 1 1 1 0 1111110111111

5. Elongation and desaturation in this representation 1111111111111 111111111111111 Elongase 1111111111111 1110111111111 Desaturase

6. Challenges for building the model 2. LC-PUFA (Long Chain Fatty Acid) biosynthesis pathway is not completely understood. Tomohito Arao T. and Yamada M., Phaeodactylum Tricornutum, Phytochem 1994;35:1177–1181.

7. Challenges for building the model 2. LC-PUFA (Long Chain Fatty Acid) biosynthesis pathway is not completely understood. Hamilton M.L., et al Metab Eng. 2014 Mar; 22(100): 3–9.

8. Stochastic model 18:0 pool Elongation 20:0 pool

9. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool Desaturation

10. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool 18:1 pool

11. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool 18:1 pool

12. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool 18:1 pool

13. Model Schematic Acyl-CoA pool 16:0-CoA 18:0-CoA 18:1-CoA Endoplasmic Reticulum Stochastic modification of LC-PUFA 18:0 20:0 18:1 20:1 18:2 20:2 18:3 20:3 K influx K TAG K e K d TAG

14. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool 18:1 pool

15. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool 18:1 pool

16. Stochastic model 18:0 pool Elongation 20:0 pool 20:1 pool 18:1 pool TAG

17. Assumptions of the model There is just one type of elongase and one type of desaturase in the model. There is a minimum of 3 carbon distance between each double bonds. Desaturation can continue until a molecule is fully desaturated. Elongation can happen until the chain length of an FA reaches maximum chain length, which is set to 20 for now. Every three FA molecules can form a TAG molecule.

18. Time course of FA production Time evolution of intermediate fatty acids Time evolution of TAG fatty acids

19. Intermediate fatty acid distribution TAG fatty acid distribution

20. Most probable pathways >20% <20%

21. Parameters effect on FAs’ distribution K e /K d =1, K TAG /K influx =0.01 K e /K d =0.01, K TAG /K influx =1

22. Influence of parameters on most probable pathways K e /K d =1, K TAG /K influx =0.01 K e /K d =0.01, K TAG /K influx =1 >20% <20%

23. Parameter influence on the EPA production Intermediate EPA EPA in TAG (K e /K d, K TAG /K influx )

24. Summary A stochastic model for biosynthesis of FA has been developed. The model demonstrates the influence of different rates (for elongation, desaturation, influx and out flux of a molecule) on the abundance of FA. The model can suggest the most probable reactions.

25. Feature works Add 3 (?) different type of desaturase to the model. Implement the feedback from TAG/ER(?) to FA pool outside ER. A more systematic parameter search. Search for enzyme parameters via literature and implement them to the model, maybe adapt the model to prokaryote first?

26. Intermediate fatty acid distribution TAG fatty acid distribution

27. Parameters effect on FAs’ distribution K e /K d =1, K TAG /K influx =0.01 K e /K d =0.01, K TAG /K influx =1

28. Parameters effect on FA biosynthesis