Team:Reading/Transformations

Transformations

Genetic modifications, with the aim to increase the number of electrons transfered to the anode in our fuel cell, are a major part of our project. Here we detail the transformations we plan to carry out with Synechocystis sp. PCC 6803, and the modifications these will produce.

Inducing hyperpilation

Bacterial nanowires are electrical conductive projections, which are a form of modified pili that allow electrons to travel away from the cell surface to reduce the cells surroundings. Nanowires were first identified in the bacterium Geobacter sulfurreducens1, and when the gene PilA1, (coding for pilin, the protein subunit of pili), was deleted, G. sulfurreducens became unable to reduce Fe3+ and Mn4+ in its surroundings1.

Since their initial discovery, bacterial nanowires have been observed in several species of bacteria, including Synechocystis sp. PCC 68032, which holds much importance for our project.

Mountain View

Much research has been done on nanowires, however mainly in Geobacter species, and there has been little research into how nanowires function in Synechocystis sp. PCC 6803. Because of this, the method by which Synechocystis sp. PCC 6803 transfers electrons to the electrodes in biological photovoltaic devices is very much unknown; endogenous mediators such as Quinones or Flavins may be being produced to shuttle electrons, or nanowires may be providing the major route of electron transportation in Synechocystis sp. PCC 68033. This casts doubt over the benefits of attempting to modify or induce the over-expression of nanowires in Synechocystis sp. PCC 6803.


We plan to induce hyperpilation in Synechocystis sp. PCC 6803, as we believe this will have several benefits to our photovoltaic even without the added bonus of bacterial nanowires. Hyperpilation may promote the formation of biofilms on the electrode surfaces in our fuel cell, and will aid adhesion of bacterial cells to the carbon fibre surface. We plan to induce hyperpilation by deleting the gene pilT1, an ATPase which controls pilus retaction5. When deleted, the bacteria exhibit more numerous long surface pili4. We also shall modify Synechocystis sp. PCC 6803 by inserting additional copies of the pilA1 gene, which codes for pilin, the pilus subunit, so that pilA1 is responsible for the biogenesis of pili5. The insertion of pilA1 will induce hyperpilation in the bacterium, and if the nanowires in Synechocystis sp. PCC 6803 are formed and function in a similar manner to those in G. Sulfurreducens, should also induce the production of more bacterial nanowires, which will increase the current of electrons from the bacteria to the anode in our fuel cell.

Modifying the photosynthetic electron transport chain

Potential modifications for the future

References

  1. Reguera, G. et al. Extracellular electron transfer via microbial nanowires. Nature 435, 1098–1101 (2005).
  2. Gorby, Y. A. et al. Electrically conductive bacterial nanowires produced by Shewanella oneidensis strain MR-1 and other microorganisms. Proc. Natl. Acad. Sci. 103, 11358–11363 (2006).
  3. Bradley, R. W., Bombelli, P., Rowden, S. J. L. & Howe, C. J. Biological photovoltaics: intra- and extra-cellular electron transport by cyanobacteria. Biochem. Soc. Trans. 40, 1302–1307 (2012).
  4. Okamoto, S. & Ohmori, M. The Cyanobacterial PilT Protein Responsible for Cell Motility and Transformation Hydrolyzes ATP. Plant Cell Physiol. 43, 1127–1136 (2002).
  5. Yoshihara, S. & Ikeuchi, M. Phototactic motility in the unicellular cyanobacterium Synechocystis sp. PCC 6803. Photochem. Photobiol. Sci. 3, 512–518 (2004).