One of the most important outcomes of the last quarter-century of synthetic biology is the recognition that the biopolymers that have been delivered to us by 4 billion years of biological evolution are not the only molecules that might support genetics, inheritance, evolution, and catalysis. Developing new biopolymers and characterizing their properties in living organisms not only has significance for testing our ideas about the functional optimization of the existing "molecules of life" but also opens new opportunities in biotechnology. Such work also permits insights into questions of the uniqueness of terrean biology and whether life in other parts of the universe could be constructed using alternate chemistries. Considering just DNA and RNA (collectively xNA), it is now clear that the four distinct "standard" building blocks (A, G, C, T and its equivalent U) for xNA do not exhaust the constraints imposed by the two rules guiding Watson-Crick pairing in natural nucleic acids. For example, the number of nucleobase pairs can be increased from two to six by merely rearranging hydrogen bond donor and acceptor groups. Efforts to implement this observation in practice using chemical synthesis, have (so far) led to two "generations" of novel heterocycles that can be incorporated into precursors suitable for use in automated xNA synthesis, thereby yielding artificially expanded genetic information systems (AEGIS). Although exploiting altered patterns of hydrogen bonding to obtain novel nucleobase pairs that are (in principle) "orthogonal" to A:T and G:C appears straightforward, the practical realization of these ideas has proven surprisingly problematic. For example, some potential heterocycles have highly populated tautomeric forms with altered hydrogen bonding patterns; these can base pair with standard nucleobases in either duplex DNA or within the active sites of polymerases, thereby giving rise to unanticipated mutations or the loss of the AEGIS nucleobases during replication. Being able to predict tautomer populations in solution or within enzyme active sites prior to chemical synthesis would be a significant step in improving the efficiency with which new nucleobase pairs can be discovered. Even were these "design" problems to be resolved, little is known about how the incorporation of these non-natural nucleobases into xNA affects the conformational preferences and dynamical properties of these complex molecules, which are fundamental to the interaction of "standard" xNA with proteins, such as polymerases, and transcription factors. We note that there has been a dearth of studies aimed at understanding how AEGIS nucleobases, which have altered electrostatic properties (dipole moments, charge distribution), might perturb xNA structure in both free solution and when bound within polymerase active sites. Finally, the validation of the force field parameters needed to model AEGIS nucleobases by, for example, comparing calculated free energies of interaction between xNA and proteins with experimental measurements has not yet been reported. Work in this project will therefore seek to address the problems outlined above by (i) developing and validating new computational methods for determining the populations of tautomeric forms of AEGIS nucleobases in water and in protein environments, and (ii) using advanced MD-based methods to understand how the incorporation of AEGIS nucleobase pairs affects the conformational and dynamical properties of duplex DNA and its interactions with DNA-binding proteins and polymerases. In particular, free energy perturbation methods will be used to study how replacing "standard" Watson-Crick bases by an AEGIS nucleobase pair changes the affinity of the DNA-binding domain of the human SETMAR transcription factor. The successful accomplishment of this aim will lay a foundation for obtaining novel endonucleases capable of cleaving AEGIS-containing duplex DNA.

This proposal describes a programme of research on single-particle and collective radiation-beam-plasma interactions at high field intensities, production of high-brightness particle beams with femtosecond to attosecond duration, new sources of coherent and incoherent radiation that are both compact and inexpensive, new methods of accelerating particles which could make them widely available and, by extending their parameter range, stimulate new application areas. An important adjunct to the proposal will be a programme to apply the sources to demonstrate their usefulness and also provide a way to involve industry and other end-users. The project builds on previous experiments and theoretical investigations of the Advanced Laser Plasma High-energy Accelerators towards X-rays (ALPHA-X) project, which has demonstrated controlled acceleration in a laser-plasma wakefield accelerator (LWFA), initial applications of beams from the LWFA and demonstrations of gamma ray production due to resonant betatron motion in the LWFA. The programme will have broad relevance, through developing an understanding of the highly nonlinear and collective physics of radiation-matter interactions, to fields ranging from astrophysics, fusion and nuclear physics, to the interaction of radiation with biological matter. It will also touch on several basic problems in physics, such as radiation reaction in plasma media and the development of coherence in nonlinear coupled systems.