My research is in Nucleic acid chemistry, DNA sequence recognition and application of nucleic acids to nanotechnology, diagnostics and biology. It has produced 300 publications, several patents and 3 start-up companies. My Professorial position at Oxford University is a joint initiative between Chemistry and Oncology.
Aberrant base pairing and implications for DNA repair and mutagenesis
My research is interdisciplinary, focusing on nucleic acid chemistry and its applications in other fields including DNA diagnostics, forensic science and nanotechnology. I have used NMR and X-ray crystallography to study the properties of base pair mismatches in DNA and their implications for DNA repair. Examples are the elucidation of the structures of DNA duplexes containing G.T (Nature 1985, 315, 604) and A.C mispairs (Nature 1986, 320, 552). This approach led us to determine the molecular basis of the mutagenic effect of a number of chemical lesions in DNA, for example O6-methyl guanine (Proc. Natl. Acad. Sci. USA 1990, 87, 9573). This was the first time that the structural basis of a mutation caused by chemical damage to DNA had been defined at the molecular level. Our studies also highlighted the fact that mispaired bases form distinct structures and cause minimal distortion to the overall conformation of the DNA duplex. Therefore enzymatic recognition of modified base pairs in living systems must depend upon subtle differences.
The O(6) methylguanine lesion in DNA mimics a Watson-Crick base pair, whereas the correct partner (cytosine) induces a distorted base pair under physiological conditions.
We have also investigated the molecular basis of the recognition of mismatched and mutagenic nucleobases by DNA repair enzymes. An example of this is a collaboration to elucidate the structural basis of specific base-excision repair by uracil-DNA glycosylase (UDGase) and MUG (Nature 1995, 373, 487, and Nature Struct. Biol. 1998, 5, 697). These essential enzymes reverse the damage that is caused to DNA by oxidation of cytosine bases, recognizing the absence of a single methyl group to initiate excision-repair. UDGase is an important enzyme in pathogens and as such is a target for antiviral therapy.
Diagnostic and Forensic applications of synthetic DNA (oligonucleotides)
The expertise that we gained from these fundamental studies on DNA base pairing was then used in the emerging field of molecular genetics to develop new methods of mutation analysis. In collaboration with AstraZeneca we invented a novel fluorescence-based real-time PCR method for the identification of mutations and single nucleotide polymorphisms (SNPs) in the human genome. This was one of the first successful technologies for rapid mutation detection and was patented, published in Nature Biotechnology (1999, 17, 804) and named “Scorpion Primers”.
The key elements of a Scorpion primer and how it functions
After the development of the new Scorpion Primers technology the key scientists from AstraZeneca set up DxS Ltd to develop mutation detection products. In 2008 DxS launched its first Scorpion-based companion diagnostic product to allow European sales of Amgen’s colorectal cancer drug, Vectibix® which was initially rejected by the European Medicines Agency on the basis of limited efficacy. The DxS kit was used to stratify patients on the basis of their KRAS mutation status, and Vectibix was approved for the KRAS wild-type population. Further diagnostic products followed, with the EGFR kit being used to establish the mutation status of non-small cell lung cancer tumours, to determine likely response to the drugs Iressa® and Tarceva®. DxS was acquired by Qiagen in 2009 and continues to develop diagnostic kits for use in personalised medicine, recently obtaining FDA approval of their KRAS kit in the US for use with the colorectal cancer drug Erbitux®.
I am also co-inventor of “HyBeacons”, novel fluorogenic probes that utilises DNA melting temperature as a means of identifying mutations. HyBeacons are the result of collaboration with the Laboratory of the Government Chemist (LGC). They can be used for the rapid diagnosis of bacterial infections and genetically-related diseases, and are also being developed by us for Forensic applications (New Scientist Jan 14th 2006). We recently demonstrated that such methodology has potential for rapid human identification at crime scenes and in custody suites, with clear implications for crime detection (Forensic Sci. Int. Genetics. 2008, 2, 333, Org. Biomol. Chem. 2008, 6, 4553). A collaboration with Philip Bartlett in Southampton recently led to a new method for identifying mutations in the human genome using a novel combination of SERS detection and electrochemical DNA melting, and a method for forensic analysis using SERS detection and electrochemical DNA melting (J. Am. Chem. Soc. 2008, 130, 15589; Angew. Chem. 2010, 49, 5917; J. Am. Chem. Soc. 2012, 134, 14099; Chem. Sci. 2013, 4, 1625).
Most of our research starts with organic chemistry and the synthesis of chemically modified DNA. For example our advances in genetic and biophysical analysis of DNA are based on the synthesis of novel fluorescent oligonucleotides (e.g. Nature Protocols 2008, 2, 615-623, Nucleic Acids Res., 2002, 30, e39).
We recently developed a click chemistry methodology which has been used to synthesise a DNA catenane (JACS 2007, 129, 6859) and to unravel the DNA binding mechanism of a novel threading intercalator (JACS 2008, 130, 14651).
DNA catenane made by click ligation
We have also used a related approach to construct large biologically active RNA constructs which are beyond the reach of conventional solid-phase synthesis (PNAS 2010, 107, 15329) and to synthesise DNA templates containing modified triazole backbones which are copied by DNA and RNA polymerases and are functional in vivo (JACS 2009, 131, 3958; PNAS 2011, 108, 11338; Chem. Comm. 2011, 47, 12057, Nucleic Acids Res. 2012, 40, 10567, Chem. Sci. 2013, DOI: 10.1039/c3sc51753e). The structural basis of the remarkable biocompatibility of this triazole linkage has recently been elucidated (Chem. Eur J. 2011, 17, 14714) and our work has been reviewed (Accounts Chem. Res. 2012, 45, 1258). Click nucleic acid ligation is currently being used in the chemical synthesis of genes and in several other biological applications, for which we have received substantial funding (BBSRC sLoLa grant “Extending the boundaries if nucleic acid chemistry”).
Artificial click DNA backbone canbe read-through by T7 RNA polymerase and is functional in bacteria
External Appointments and Recognition
I have received several awards including the Royal Society of Edinburgh MakDougall-Brisbane prize for research, the Royal Society of Edinburgh Caledonian Research Fellowship, the Royal Society Leverhulme Senior Research Fellowship, the Royal Society of Chemistry Josef Loschmidt prize, the Royal Society of Chemistry award for Nucleic Acid Chemistry and the Royal Society of Chemistry prize for Interdisciplinary Research. I am a Fellow of the Royal Society of Edinburgh, a Fellow of the Royal Society of Chemistry, President-elect of the Chemical Biology Interface Division committee of the Royal Society of Chemistry, and a member of the editorial board of Chemistry World.