Research Interests

Synthetic biology encompasses a wide range of molecular biology tools that enable researchers to precisely manipulate the DNA sequence within a gene, gene cluster, or genome. Recent developments, however, have made it increasingly possible to generate man-made enzymes that can perform similar functions on artificial genetic polymers that are distinct from those found in nature (DNA and RNA), but still capable of heredity and evolution. Collectively referred to as xeno-nucleic acids, or XNAs, these genetic polymers have unique physiochemical properties that include resistance to nuclease digestion and expanded chemical functionality. We envision a future where many of the same synthetic biology tools available to manipulate DNA and RNA are available to manipulate XNA. This field of science, termed ‘xenobiology’, is expected to produce novel biological systems whose information is stored in XNA-coded polymers. Such efforts open the door to a vast new world of synthetic genetics, where sequence-defined synthetic polymers can be used to create new tools for biotechnology and medicine, and possibly even improve our understanding of the origin of life itself.

While the possibility of manipulating XNAs in a test tube has enormous potential for new applications in biotechnology and medicine, several challenges must be overcome before such achievements can be realized. The most significant challenges include:

The development of chemical synthesis strategies that deliver XNA substrates on the scales required for enzymatic studies
The creation of new molecular evolution approaches that enable the production of highly efficient XNA enzymes
The elucidation of NMR and X-ray crystal structures that reveal how XNA enzymes function and provide insights into how XNA enzymes can be generated with improved activity

Chemical Synthesis

In order to develop enzymes that function on artificial genetic polymers, researchers must have access to XNA substrates (nucleotide triphosphates and oligonucleotides) that can be produced on the scales required for enzyme evolution and eventual downstream applications. This challenging endeavor requires developing synthetic strategies that enable grams of XNA monomers to be generated in high yield and purity, as well as new methodologies that endow XNA substrates with expanded chemical functionality.

Enzyme Engineering

Xenobiology will require a wide range of enzymes that can modify XNA in different ways. Of these, polymerases represent an important initial function due to their ability to synthesize and replicate genetic information. To help meet this challenge, we have developed a general strategy for evolving XNA polymerases called droplet-based optical polymerase sorting (DrOPS) that employs an optical sensor to monitor polymerase activity inside the microenvironment of uniform synthetic compartments generated by microfluidics.

Structure Determination

The evolution of polymerases that can synthesize and recover genetic information stored in XNA polymers demonstrates that the biology concepts of heredity and evolution are not limited to the natural genetic polymers of DNA and RNA. Examining how these enzymes function at a molecular level and using this information to generate new enzyme variants with improved activity is an important goal that would help expand the XNA toolkit.

Coming soon…

In Vitro Selection (SELEX)

In vitro selection is a powerful method for evolving nucleic acid molecules that can fold into shapes with specific target binding affinity or catalytic activity. Although this approach has been widely applied to DNA and RNA, natural genetic polymers have limited practical value because these polymers are prone to nuclease digestion. However, XNAs, by virtue of their unnatural backbone structures, are highly resistant or, in some cases, recalcitrant to nuclease digestion. Consequently, there exists a significant demand –medically and commercially– for aptamers and catalysts that retain their function in complex biological environments. In addition, such studies have the potential to inform us about nature’s choice of DNA and RNA as the molecular basis of life’s genetic system.