Main

One important benefit of the ongoing nanotech revolution has been the recognition of nucleic acids as a useful and versatile scaffold for building more complex biochemical tools. A variety of specific functional groups can be covalently tacked onto nucleotides, which can in turn be assembled into chains via several chemical reactions. This can be laborious, however, and some scientists are now looking into the possibility of using naturally occurring enzymes as a more efficient alternative.

One such researcher is Michael Famulok, of the University of Bonn, whose group has published several articles demonstrating the feasibility of this approach. More recently, they scaled up their efforts, systematically measuring the incorporation of a variety of nucleotides containing different functional groups—acidic, basic or lipophilic—into oligonucleotide chains using seven different bacterial DNA polymerases. Some enzymes had only a limited ability to work with unorthodox substrates, but two polymerases, Pwo and Vent (exo−) stood apart from the rest, efficiently incorporating every nucleotide variant tested. In subsequent experiments, Famulok's team delineated template sequence determinants that may affect the efficiency of incorporation and demonstrated the efficient enzymatic synthesis by these two polymerases of oligonucleotides containing variants of all four nucleotide types, as well as the successful PCR amplification of a modified template to produce a similarly functionalized product.

The flexibility of these polymerases could have dramatic implications for future biochemical engineering projects, such as aptamer development. “Additional chemical functionality might give an aptamer more protein-like activities, without losing its advantageous nucleic acid–like properties,” Famulok explains, “and that might be useful in certain cases where you would like to use an aptamer as a diagnostic or a drug.” He also envisions using this approach for the engineering of complex, higher order DNA structures—for example, building modified DNA molecules with altered charge distributions that can self-assemble in a manner similar to the way proteins interact. And although the work still remains to be done, the same principles could apply to the functionalization of RNAs, creating new possibilities for the fine regulation of microRNAs. “You could introduce caged bases into a microRNA in an enzymatic fashion,” suggests Famulok, “and then try to activate this microRNA by a light pulse, or something like that.”

Most of all, Famulok hopes that his group's work will provide a valuable reference for other researchers with innovative ideas: “If people would want to use high-density functionalized DNA for anything—for ideas that I'm too stupid to come up with—then they know how to make them, and they can find very important guidelines in our paper.”