Next-Generation Fluorescent Nucleosides and Structure-Photophysics Relationships

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In this project, funded by the Chemical Structure, Dynamics and Mechanisms B Program of the Chemistry Division, Professor Byron Purse of the Department of Chemistry and Biochemistry at San Diego State University (SDSU) is developing fluorescent nucleoside analogues as molecular probes with new capabilities for biophysical studies of nucleic acids. In addition to storing the genetic code, nucleic acids can undergo chemical modification, adopt folded structures, and interact with proteins and enzymes for the regulation of gene expression and metabolism. Fluorescent nucleoside analogues are powerful probes for studying these processes, but existing analogues have limited capabilities. Insufficient knowledge of the factors controlling their fluorescence hinders the rational design of better probes. This project seeks to overcome these limitations and expand the toolkit of fluorescent nucleoside analogue probes with significant, novel capabilities. The design of the new analogues is aided by detailed mechanistic studies of how the local environment in DNA/RNA influences the fluorescence of the probes. This cross-disciplinary research provides excellent training opportunities for students at multiple levels. In addition to graduate student and undergraduate involvement, a Course-based Undergraduate Research Experience (CURE) in the SDSU Physical Chemistry teaching lab and an outreach program involving computational work with community college and high school students will broaden participation in the research.Probably the greatest challenge in designing fluorescent probes that mimic the structure of biomolecules is that limited ability to predict photophysical properties currently necessitates a trial-and-error approach. In this project, photophysical measurements, computation, NMR, and x-ray structure determination are combined to develop a detailed, predictive understanding of the relationships between nucleoside analogue structure and photophysics, especially in the base stack of DNA and RNA. By harnessing these trends and synthesizing new fluorescent nucleoside analogue designs, novel probes with (a) brightness matching conventional fluorophores, (b) absorption and emission at the red end of the visible spectrum, (c) strong fluorescence turn-on responses to specific biomolecular recognition events, and (d) the ability to report on chemical modifications to nucleobases, e.g. from methylation or DNA damage are being developed.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.