CAREER: Development of DNA polymerases capable of high fidelity modified DNA synthesis

点击翻译按钮获取中文摘要

DNA is widely known as the material that encodes information in all life forms. Besides this role, DNA is also one of the most valuable tools in modern biotechnology. Foundational technologies, such as the polymerase chain reaction and DNA sequencing, as well as emerging technologies, such as DNA-encoded drug discovery, rely on the unique functions of DNA. However, the usefulness of DNA-based tools is limited by DNA's susceptibility to destruction in a biological environment. With this award, the Chemistry of Life Processes Program in the Chemistry Division is funding Dr. Aaron Leconte from the W. M. Keck Science Department of Claremont McKenna, Pitzer, and Scripps Colleges to develop proteins that can synthesize chemically-modified forms of DNA that are more stable than natural DNA in a biological environment. Consequently, the modified DNAs have the potential to expand the applications of DNA-based tools, such as high throughput DNA sequencing and synthetic biology used in a number of therapeutic and clinical diagnostics. Such research may benefit society by improving human health. Professor Leconte integrates research experiences in laboratory courses for undergraduate students to prepare these students for STEM graduate studies and careers. He is also developing videos that document the research experiences of current and former undergraduate students that can be used to inspire and attract more students to STEM studies. This research project seeks to develop mutant DNA polymerases capable of high fidelity synthesis of modified forms of DNA called M-DNA. This DNA is resistant to nucleases. M-DNAs are generally not amplified by native DNA polymerases. Mutant DNA polymerases capable of synthesizing long, modified DNAs have been identified only recently. While these mutant DNA polymerases represent an exciting step forward, these enzymes make frequent errors, limiting their application. The research in the Leconte laboratory focuses on the characterization and engineering of a recently discovered mutant DNA polymerase, SFP1. Preliminary data suggest that this enzyme is capable of M-DNA synthesis and that it makes far fewer errors than other previously characterized M-DNA polymerases. The Leconte group develops high throughput sequencing methods that can quantitatively evaluate M-DNA synthesis. The high-throughput sequencing data obtained by these methods inform engineering efforts to produce DNA polymerases that have a lower error frequency during M-DNA synthesis. The new polymerases that can perform modified DNA synthesis with higher fidelity have the potential to enable an array of applications in fields that range from clinical diagnostics to synthetic biology.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.