Part:BBa_K1895004

Porin Overexpression for Microbial Fuel Cell Use

As part of our iGEM project we worked on trying to improve the current output from a microbial fuel cell. Microbial fuel cells which use E. coli depend on a mediator, usually methylene blue to transfer electrons between the electrodes and the organisms electron transport chain. The mediator enters the cell through porins in the cell membrane and 'steals' electrons, becoming oxidised. These electrons are then carried to the cathode, which is positively charged where they are lost, allowing electrons to flow around the circuit.

By increasing the amount of mediator that can enter the cell we hope to be able to increase the electron flow rate, or current, from our microbial fuel cell. This occurs because the electrons can be moved more freely and they're movement via the mediator is less limited by the rate at which the cell can transport the mediator across the membrane.

Usage and Biology

We are exploring two ways of increasing the amount of mediator entering and exiting the cell, the first of these is by expressing a larger porin (BBa_K1895004) the second, this part, is to increase the number of porins. For this purpose we have designed this device to control the overexpression of one of the E. coli natural porins, [http://www.rcsb.org/pdb/explore.do?structureId=2OMF OmpF].

We chose OmpF because it occurs naturally in E. coli and is suitable for the mediator we use in our fuel cell (which is a variant of the [http://www.ncbe.reading.ac.uk/ncbe/materials/METABOLISM/fuelcell.html University of Reading's NCBE kit]). OmpF is permeable for molecules smaller than 600 Da whilst [http://biocyc.org/compound?orgid=META&id=CPD0-1625 methylene blue] is 284 Da.

By adding L-arabinose to the grown media the number of OmpF porins in the E. eoli membrane should increase, allowing increased extracellular transport of methylene blue and faster electron shuttling between the cells and the electrodes. This should correspond to an increase in current. This function is shown below.

Note that over-expressing a membrane protein like OmpF may increase replication stress and lead to reduced growth rate.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 125
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 255
  Illegal BamHI site found at 65
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal AgeI site found at 388
  
• 1000
  COMPATIBLE WITH RFC[1000]