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Project 3 Metal Transporter

In this project our aim is to transport specific metal ions from a specific location to another on the surface of engineered bacterial cells. In other words, to use cells as metal carriers.

• We didn't want the metal ions to be uptaken by the cells as in usual bioremediation procedures, since we want to "transport" specific metal ions (for example radioactive ones) that could be toxic to cells and also we want to reobtain those metal ions from the cells to keep the cells cycling in the system and for the metal ions to be used in other industrial purposes. So we focused on putting metal-binding domains on the surface of the bacterial cells. This can be done by adding short metal-binding domains to bacterial constitutively expressed transmembrane proteins such as LamB or OmpC.(1)

• For the metal transportation to occur the cells should adsorp metal ions at a specific location and release them at another specific location. In order to achieve this we planned to use the pH dependence of the metal-binding domains' affinity for metals.

• We need at least a semi-solid medium for bacterial cell movement. Thus, we cannot use a constant pH gradient in the medium due to diffusion.

• We decided to use the bacteriorhodopsin/proteprhodopsin molecule, which transports protons into the cells when exposed to light at a specific wavelenght. We think that it will create a pH decrease at the surface of the cells if the cells are illuminated with a specific wavelength of light. This sudden pH decrease will cause the metal-binding domains to release the metals that they have previously bound. So the location where we want the cells to release the metal ions should be illuminated with a specific wavelength.

• We planned to use phototaxis in order to make the cells move from metal ion-rich area to our desired location for release. We did not want to use chemotaxis because of the limitations caused by diffusion. Although, E.coli lacks a phototaxis mechanism, but have a chemotaxis; fortunately, these two taxis ayatems have similar molecular mechanisms except for the receptor parts. E.coli cells can be engineered to give taxis responses to light, by introduction of sensory rhodopsin and its transducer.(2)

• As a result we will have a medium in which metal uptake and release locations and also the path are illumintated by lights at specific wavelenghts. We could imagine that the E. coli cells will act as trucks carrying cargo (metal ions) from one house to another on a road we instantaneously build by using light.

• • • A schematic representation of the system

• Where can we use such a complex system:

- In bioremediation, cleaning heavy metals and also nuclear remnants (There are metal binding proteins for a variety of metals, and even for uranium)

- In high degree purification of metals for industrial purposes

References:

1) Engineering the Escherichia coli Outer Membrane Protein OmpC for Metal Bioadsorption, Norberto Cruz, Sylvie Le Borgne, Georgina Hern´andez-Ch´avez, Guillermo Gosset, Fernando Valle & Francisco Bolivar, 2000, Biotechnology Letters, 22: 623–629

2) An Archaeal Photosignal-Transducing Module Mediates Phototaxis in Escherichia coli', KWANG-HWAN JUNG, ELENA N. SPUDICH, VISHWA D. TRIVEDI, AND JOHN L. SPUDICH, 2001, Journal of Bacteriology, Vol. 183, No. 21, p. 6365–6371

• Bacterial Rhodopsin: Evidence for a New Type of Phototrophy in the Sea, Oded Beja, L. Aravind, Eugene V. Koonin, Marcelino T. Suzuki, Andrew Hadd, Linh P. Nguyen, Stevan B. Jovanovich, Christian M. Gates, Robert A. Feldman, John L. Spudich, Elena N. Spudich, Edward F. Delong, 2000, Science, Vol 289, 15 September 2000, p. 1902-1905
• Proteorhodopsin Phototrophy in the Ocean, Oded Beja, John L. Spudich, Elena N. Spudich, Marion Leclerc and Edward F. Delong, 2001, Nature, Vol 411, 14 June 2001, p. 786-789

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