Abstract
DNA polymerases play a central role in biology by transferring genetic information from one generation to the next during cell division. Harnessing the power of these enzymes in the laboratory has fueled an increase in biomedical applications that involve the synthesis, amplification, and sequencing of DNA. However, the high substrate specificity exhibited by most naturally occurring DNA polymerases often precludes their use in practical applications that require modified substrates. Moving beyond natural genetic polymers requires sophisticated enzyme-engineering technologies that can be used to direct the evolution of engineered polymerases that function with tailor-made activities. Such efforts are expected to uniquely drive emerging applications in synthetic biology by enabling the synthesis, replication, and evolution of synthetic genetic polymers with new physicochemical properties.
Highlights
DNA polymerases are an ancient family of enzymes responsible for replicating the genomes of organisms during cell division
We examine the impact of polymerase engineering on the field of synthetic biology
In a striking example of enzyme promiscuity, we recently discovered two naturally occurring DNA polymerases that will faithfully replicate 2′-fluoroarabino nucleic acid (FANA) (Wang et al, 2018b), which is an unnatural genetic polymer that contains 2′-fluoroarabino residues in place of natural ribose or deoxyribose nucleotides (Damha et al, 1998)
Summary
DNA polymerases are an ancient family of enzymes responsible for replicating the genomes of organisms during cell division. Structures of KlenTaq (Klenow-fragment analog of Taq DNA polymerase) containing an abasic site in the template reveal that the conserved gating tyrosine residue (Fig. 4b) can pair opposite an incoming substrate to allow for primer extension, albeit at significantly reduced rates due to the formation of a sub-optimal enzyme active site (Obeid et al, 2010, 2012). In 1987, Benner and coworkers suggested that the functional activity of nucleic acid catalysts could be improved by incorporating additional chemical diversity into DNA and RNA (Benner et al, 1987) Toward this goal of augmenting nature’s genetic alphabet, several non-natural base pairs were envisioned that would allow for novel hydrogen-bonding schemes between the various hydrogen-bond donor and acceptor groups found on the Watson–Crick face of designer nucleobases (Fig. 7). Baar et al (2011) Ghadessy et al (2001) Ghadessy et al (2001) d’Abbadie et al (2007)
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