January 2011


Genetically Damaged Bacteria Grow via Synthetic Proteins

SUMMARY: Bacteria which possess genetic damage that normally prevents reproduction in a nutrient-deficient medium can be saved by expression of artificial proteins, an important step towards constructing artificial life.
The general "mainstream" media occasionally hypes news of scientists constructing "artificial life" in the laboratory. It happened when a functional virus was first synthesized (an exact copy of one that already exists), and again last year when one bacterium was reprogrammed with another bacterium's DNA.

These were both important advances. However, neither was an example of artificial life.

Scientists are far away from designing a true living artificial cell. In other words, scientists aren't even close to fabricating a living cell from scratch, one that is comprised of artificial components.

For example, scientists aren't able to construct a fully artificial genome, i.e. the chemical "blueprint" of a cell. Initial advances towards this goal have been made recently, e.g. primitive frameshift reading of a synthetic polymer, but the genomic complexity seen in real life, such as epigenetics, is currently far beyond artificial construction.

Scientists have been reprogramming cells for some time now, altering their DNA such that they produce useful materials, e.g. fluorescent metal nanocrystals and synthetic polypeptides. This ability may serve as a useful starting point towards a fully artificial cell.

Specifically, one can envision bridging the divide between fully artificial cells (which have not been constructed) and reprogrammed cells (which have been constructed) by reprogramming bacteria to synthesize artificial proteins that rescue them from otherwise growth-preventing genetic damage. This research goal has been achieved by Michael Hecht (Princeton University, United States) and coworkers.

Initial inspiration.

It turns out that the proteins in living cells are comprised of only a very small number of the possible compositions that are theoretically possible. This in turn suggests that life could possibly be sustained by a far larger variety of protein compositions than are found in nature.

The scientists tested this hypothesis by searching for artificial proteins that sustain the growth of genetically damaged cells. They didn't screen proteins at random; in addition to taking an impossibly long length of time, many of these hypothetical proteins would not be able to fold into a stable three-dimensional structure (a typical, but not absolute, requirement for protein function).

They instead screened a database of over a million proteins designed to stably fold based upon their water-repelling (hydrophobic) and water-attracting (hydrophilic) composition. Specifically, each of the proteins was 102 subunits long, and was designed to possess 4-helix bundles.

Note that no function was "built into" these proteins. Although the protein design requirements were structural, not functional, function in living cells was still found with some of the proteins, as discussed next.

Rescuing bacteria.

The basic idea was that the scientists genetically engineered their bacteria to express one of their artificial proteins. The bacteria were also genetically engineered to lack a gene normally required for growth in a specific nutrient-deficient medium.

When the bacteria don't grow, the artificial protein clearly doesn't "save" the bacteria. When the bacteria do grow, the artificial protein at least partially compensates for the normally expected loss of bacterial function (further necessary control experiments, which the scientists did perform, are not discussed here).

A number of successful proteins were found, each of which rescued the function of a bacterial cell with one of four genetic deletions. A search of relevant biochemical databases confirms that none of these proteins matches known proteins in nature.

Further experiments suggest that at least 0.1% of the bacterium's genome, and 1% of that otherwise necessary for growth in the scientists' nutrient-deficient medium, is replaceable via the artificial proteins (gene products). This is especially impressive, given that the artificial proteins were designed with structure, not function, in mind.

Almost none of these artificial proteins enable bacterial growth that's as fast as with the normal biological protein. Again, the fact that there's any growth at all is notable, especially given that the artificial proteins were designed with structure, not function, in mind.

This brings up the question of the function of these artificial proteins. What do these proteins do in the bacteria to enable growth?

Artificial protein function.

The scientists deleted bacterial genes important in serine synthesis, glutamate synthesis, isoleucine synthesis, and iron uptake. How do the artificial proteins make up for such limitations?

The short answer is that the scientists don't know. So far they've ruled out the possibility that the artificial proteins generate the exact same end product as the natural proteins (an unlikely proposition to begin with), and the possibility that the artificial proteins generated a generic compensatory stress response.

They've largely ruled out the possibility that the artificial proteins alter the expression or function of some other known protein. Further experiments to determine the proteins' function are complicated by the fact that the function is at low levels, and unknown molecules may be required for their function (not necessarily, and probably not, the same molecules required for the function of the natural protein).

Final comments.

Bacterial growth can at times be sustained via artificial proteins which are far simpler in structure than the proteins commonly found in nature. Again, "artificial life" has not been demonstrated with these experiments, but they're nevertheless an important, early step in that direction.

NOTE: The scientists' research was funded by the National Science Foundation.

Fisher, M. A., McKinley, K. L., Bradley, L. H., Viola, S. R., & Hecht, M. H. (2011). De Novo Designed Proteins from a Library of Artificial Sequences Function in Escherichia Coli and Enable Cell Growth PLoS ONE, 6 (1) DOI: 10.1371/journal.pone.0015364