The Romesberg team at The Scripps Research Institute (TSRI) recently created an unnatural DNA base pair that can be incorporated into the genome of E. coli. While in nature the genetic code only contains two DNA base pairs, A-T and C-G, these researchers have made a bacterium that can “stably harbor DNA containing three” (1).
Creating this unnatural base pair posed many problems for the researchers at TSRI. First, the pair must fit into the DNA double helix and exhibit binding properties similar to those of natural base pairs in order for the DNA double helix to be stable. In addition, DNA polymerases must be able to recognize, incorporate, and properly match the unnatural base pair when synthesizing new DNA. Perhaps the most difficult challenge, these foreign nucleotides must be able to evade highly efficient DNA repair machinery in the cell. Despite these obstacles, in 2008 the team synthesized DNA and transcribed strands containing a new synthetic base pair in vitro (2).
Romesberg’s new base pair may have far-reaching effects, ranging from expression of novel proteins to new therapeutic treatments. Source: http://upload.wikimedia.org/wikipedia/commons/1/16/DNA_orbit_animated.gif
After finding success with the base pair d5SICS and dNaM in vitro, the team decided to attempt expression in vivo in E. coli, but this expression posed many problems as well (2). The unnatural nucleoside triphosphate building blocks cannot be made from material already inside the bacterium, meaning researchers had to provide the bacteria with the right building blocks in the solution surrounding the cell. In addition, to get the building blocks into the cell, nucleotide triphosphate transporters had to be added to the media (1).
After another year of work, researchers found that the bacteria grew at a normal rate using transporters from algae to facilitate intake of the unnatural base pair. When the molecular building blocks and transporters for dNaM and d5SICS were no longer provided in solution, however, the bacteria replaced the synthetic base pairs with natural A-T and C-G pairs. According to Denis A. Malyshev, one of the lead researchers on the project, “these two breakthroughs provide control over the system,” as researchers can determine when the synthetic pair is expressed by manipulating the surrounding media (1).
The findings from The Scripps Research Institute have significant implications for future research. Romesberg’s team hopes that this synthetic genetic material will be used to re-engineer cells and produce new, unnatural amino acids that can be used in novel therapeutics and nanomaterials (2).
References:
1. Denis A. Malyshev, Kirandeep Dhami, Thomas Lavergne, Tingjian Chen, Nan Dai, Jeremy M. Foster, Ivan R. Corrêa, Floyd E. Romesberg. A semi-synthetic organism with an expanded genetic alphabet. Nature, 2014; DOI: 10.1038/nature13314
2. Scripps Research Institute. “Semi-synthetic organism: Scientists create first living organism that transmits added letters in DNA ‘alphabet’.” ScienceDaily. ScienceDaily, 7 May 2014. <www.sciencedaily.com/releases/2014/05/140507132129.htm>.