DISCOVERY AND DEVELOPMENT OF ANTHRAQUINONE PHOTONUCLEASE

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THIS IS A SHANNON AWARD PROVIDING PARTIAL SUPPORT FOR THE RESEARCH PROJECTS THAT FALL SHORT OF THE ASSIGNED INSTITUTE'S FUNDING RANGE BUT ARE IN THE MARGIN OF EXCELLENCE. THE SHANNON AWARD IS INTENDED TO PROVIDE SUPPORT TO TEST THE FEASIBILITY OF THE APPROACH; DEVELOP FURTHER TESTS AND REFINE RESEARCH TECHNIQUES; PERFORM SECONDARY ANALYSIS OF AVAILABLE DATA SETS; OR CONDUCT DISCRETE PROJECTS THAT CAN DEMONSTRATE THE PI'S RESEARCH CAPABILITIES OR LEAD ADDITIONAL WEIGHT TO AN ALREADY MERITORIOUS APPLICATION. THE APPLICATION BELOW IS TAKEN FROM THE ORIGINAL DOCUMENT SUBMITTED BY THE PRINCIPAL INVESTIGATOR. Broadly, the plan is to develop a set of anthraquinone derivatives as photofootprinting agents, focusing on two areas: i) Discovery of quinones which can image major and/or minor groove-bound ligands. ii) Develop time-resolved photofootprinting systems for the study of actively transcribing DNA-RNA polymerase complexes. The reactions of these quinones with nonduplex DNA and RNA forms will be explored to determine if they are able to react selectively with unique DNA and RNA structural features such as loops and hairpins. The quinones will be conjugated with recognition elements, particularly oligonucleotides and peptide nucleic acids, to assess their ability to perform sequence-specific ds cleavage of DNA. And the recently established collaboration with PNA Diagnostics to assess the ability of these quinones to sterilize blood products when they are conjugated with PNAs will be continued. The anthraquinones have special promise as agents for the cleavage of DNA. They can be selectively and rapidly activated by near-UV light. Because of their planar, aromatic skeleton the quinones normally bind to DNA by intercalation between adjacent base pairs. However, subtle structural changes switch them to minor-groove binders and change their reactions with DNA. Excited-state anthraquinone derivatives are good oxidants. Cleavage of DNA is initiated by oxidation of the poly(sugar- phosphate) backbone or by electron transfer from a base. Most importantly, the quinones are stable to repeated cycling through oxidized and reduced forms. Thus the quinones are catalytic and each molecule can cleave numerous DNA strands. The reactions of the quinones will be studied by laser spectroscopy to assess mechanisms for transport of base radical cations (holes) through ds DNA to a site remote and to probe the nature of the initial oxidation step. Analysis of the final cleavage products will be used to reveal the precise site of attack by the quinone. The binding mode of the quinone derivatives will be studied by high resolution NMR spectroscopy.