XNA stands for Xeno nucleic acids were developed by Holliger and colleagues.
The idea of synthetic genomes evolved after Craig Venter’s synthetic cell got published in 2010. The group that invented XNA, a new synthetic artificial DNA like molecule which is capable of storing hereditary information. They focused on the chemistry of the normal DNA backbone, nucleobases and sugars to see if there are possible chemical modifications that could be ade that could substitude bases A,T,G,C to something man-made.
1. Focus on a man-made chemical framework (XNA). XNA can base pair with DNA. Figure 1 shows the 6 artificial bases.
2. Modify polymerase that can make XNA with DNA as template
3. Modify a reverse transcriptase that can make DNA with XNA as template
Strategy 2 and 3 was done by compartmentalised self-tagging (CST) and the efficiency of the polymerase was tested by a polymerase activity assay as shown in the figure. In CST, compartments have water-in-oil emulsions with polymerases, X-NTPs and labeled primers. The polymerases are made by encoded plasmids. Plasmid serve as template. Primers bind plasmids, polymerases made by the plasmids extend. The beads bind to primer-plasmid pairs. After washes, the beads are screened further for polymerase. The polymerase activity assay measures how long it can extend using Xeno-NTPs.
Chemical modification to bases are tolerated in N7 of purines and C5 of pyramidines which extends into the major groove of the DNA . This gives more flexibility in terms of steric hindrances. Many fluorescent analogues have been made but their enzymatic synthesis and maintenance was unsuccessful. Novel base pairs give room for expansion of genetic storage of information . Figure 2 highlights the changes that were made to the sugar, backbone or the base to make new artificial DNA. Figure 3 shows the structure of the duplexes with DNA. TNA (Threofuranosyl nucleic acids) can self -pair and hybidise with DNA and RNA. Therinator polymerase can incorporate 80 TNA triphosphates. LNA have locked ribofuranose rings which works well for PCR,aptaers and aptazymes but the LNA triphosphates seem to be poor substrates. Table 1 shows the polymerases that have been succesfull in incorporating these artificial bases. The team also found that when stressed, these structures evolve into a different form. Under selective pressure, different xNTPs are incorporated.
This discovery could revolutionize synthetic biology, biotechnology and medicine. Oligo-nucleotide based drugs with optimal pharmacokinetics can be designed and custom made. It could also help us explain how life evolved. Also, its ultra cool and sounds like it cae out of a science fiction novel!
1. Science. 2012 Apr 20;336(6079):341-4. doi: 10.1126/science.1217622.
Synthetic genetic polymers capable of heredity and evolution.
2. Curr Opin Chem Biol. 2012 Aug;16(3-4):245-52. doi: 10.1016/j.cbpa.2012.05.198. Epub 2012 Jun 14.
The XNA world: progress towards replication and evolution of synthetic genetic polymers.
3. Nat Biotechnol. 2012 Jul 10;30(7):624-5. doi: 10.1038/nbt.2298.
The sweet allure of XNA.