Axel O. Vera
Axel O. Vera
Axel O. Vera, a senior chemistry major at University of Puerto Rico, Río Piedras Campus, synthesized and characterized small-molecule nucleases for miniature genome editors.
CRISPR-Cas9 has furnished transformative technologies for gene editing and functional genomics, making gene editing a point of interest in treating diseases. The Broad Institute is home because of its collaborative environment. Here, I gained many mentors who escalated the impact of my research. I also met many scientists whom I consider my heroes, and learned how to be successful from them. After such an amazing experience, a huge goal for me is to return to the Broad.To accomplish this, we need to deliver Cas9 in vivo through the cell and nuclear membrane to reach the DNA target sequence, which is challenging due to the large size of Cas9 (around 200 kDa). Therefore, alternative smaller systems should be explored to circumvent this delivery problem. Several small molecules that cleave DNA exist, and we are using them in conjunction to DNA readers to design miniature genome editors that are 70% smaller than Cas9. Here, we report the syntheses and biological assays of several metal complexes that cleave DNA by hydrolysis, reminiscent of those encountered in endonucleases. Other mechanisms of DNA cleavage exist, but hydrolytic strand-breakers are desirable since radicals involved in oxidative cleavage may induce mutations or off-target effects in the DNA. We synthesized linear, branched, and macrocyclic ligands and complexed them to metals such as La3+, Fe3+, Zn2+, and Cu2+. We used liquid chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy to confirm the structures and assess the purity of the synthesized ligands. Furthermore, we used gel electrophoresis to measure their DNA cleavage activity and we identified two complexes that were potent at low concentrations. These small-molecule nucleases will be conjugated to DNA readers such as peptide nucleic acids (PNA) to form a synthetic miniature genome editor that will be much easier to deliver than Cas9.
Project: Synthesis and characterization of small-molecule nucleases for miniature genome editors
Mentors: Sophia Lai and Amit Choudhary, Chemical Biology Program