1. Hydrogen Production by Cyanobacteria Dr. Teresa Thiel University of Missouri – St. Louis

2. Light micrograph of filaments of Anabaena sp. with heterocysts

3. Functions of heterocysts and vegetative cells

5. Expression of nitrogen fixation genes only in heterocysts Green cells are heterocysts that are able to fix nitrogen and make hydrogen. Red cells are vegetative cells that cannot fix nitrogen and make hydrogen. Arrows indicate heterocysts identified by brightfield microscopy. Bar=10  M

6. Nitrogen fixation and hydrogen production in heterocysts

7. Schindelin, H.,Kisker, C.,Schlessman, J.L.,Howard, J.B.,Rees, D.C. Nature 387:370-376, 1997.

8. Nitrogenase

9. Building the dinitrogenase components- alpha

10. Building the dinitrogenase component and now beta

11. Building the dinitrogenase component - the second alpha

12. Building the dinitrogenase component- and the second beta

13. Adding the dinitrogenase reductase components the first of four subunits

14. Adding the dinitrogenase reductase components - the second

15. Adding the dinitrogenase reductase components - the third

16. Adding the dinitrogenase reductase components the fourth

17. Half of one nitrogenase molecule with both dinitrogenase and dinitrogenase reductase components

18. The other half of one nitrogenase molecule with both dinitrogenase and dinitrogenase reductase components

19. Building half of one nitrogenase molecule - starting with the dinitrogenase reductase components

21. Building half of one nitrogenase molecule - adding the dinitrogenase components

22. Finishing half of one nitrogenase molecule - adding the second dinitrogenase component Half of Nitrogenase

24. 8 [H + + e - ] Reduces nitrogenase reductase Reduced nitrogenase reductase

25. 8 [H + + e - ] Reduces nitrogenase reductase N N

26. Functions of heterocysts and vegetative cells

27. Modified from Lindberg, P. Cyanobacterial Hydrogen Metabolism. Dissertation. University of Uppsala.2003. Anabaena produces hydrogen in a bioreactor

28. H 2 fueling station http://www.actransit.org/environment/hyroad_richmond.wu

30. Engineer nitrogenase to make more hydrogen

31. Engineer heterocysts to express an efficient hydrogenase http://www.chemistry.montana.edu/~john.peters/gallery/structure1.html CP1 hydrogenase from Clostridium pasteurianum

32. Engineer a strain with a very high frequency of heterocysts A high frequency of heterocysts as depicted here (artistic rendering, not actual photo) could allow cultures to produce large amounts of hydrogen using nitrogenase or hydrogenase.

33. Advantages of Anabaena system Requires only sunlight, air and water to grow and produce hydrogen In contrast to other algal systems, it produces hydrogen in cells growing in air No fossil fuels are required and oxygen is the only byproduct Has been demonstrated to produce ¼ of the volume of a bioreactor of hydrogen per day (Tsygankov, A. A., et al. Biotechnol. Bioeng. 80, 777, 2002)

34. 8 [H + + e - ] Reduces nitrogenase reductase N N Dinitrogenase reduces dinitrogen to ammonia and hydrogen