Seán O’Leary obtained a B.Sc. degree in Chemistry from University College Dublin, and M.S. and Ph.D. degrees in Chemistry and Chemical Biology from Cornell University. His doctoral research, with Prof. Tadhg P. Begley, focused on the enzymology of biochemical reactions involving novel mechanistic motifs, primarily in the metabolism of clinically relevant bacteria. He was a postdoctoral researcher with Prof. Joseph D. Puglisi at Stanford University School of Medicine, where he worked on applying biochemical and biophysical experimental approaches, particularly single-molecule fluorescence microscopy, to study the mechanism of eukaryotic translation initiation. He joined the UCR faculty in 2016. The focus of his laboratory is the elucidation of molecular mechanism in the operation and regulation of complex biochemical systems.

Intricate assemblies and networks of biomolecules are crucial for cellular function. Understanding mechanistically how these operate allows us to not only define fundamental biology, but also to understand cellular dysfunction in disease, and potentially how therapeutics perturb the cell to restore health. Our research aims to quantitatively elucidate molecular mechanisms in multi-step, multi-component biochemical systems, by making biochemical and biophysical measurements. We focus on eukaryotic translation initiation, a key process in the regulation of gene expression. In initiation, twelve or more protein factors are required to guide assembly of an elongation-competent ribosomal particle at the correct start codon of a messenger RNA. One major focus is to understand biomolecular dynamics – the time-evolution of composition and conformation in the pre-initiation complex – and how these dynamics mediate efficient, faithful, and regulated initiation. We use single-molecule fluorescence microscopy with fluorescently labeled initiation components to directly observe initiation as it unfolds. A second focus is the enzymology of initiation – how biomolecular catalysis, including the activity of RNA helicases, is leveraged to drive initiation. This focus also leads to a more general interest in the dynamics of RNA-protein interactions, and echoes a long-standing interest in the organic and physical chemistry of biochemical processes. We investigate how mRNA is prepared for initiation, how the small ribosomal subunit is recruited to the mRNA, how the mRNA is scanned to locate the start codon, and how the ribosomal subunits join to begin elongation. Throughout, the aim is to first mechanistically understand the fundamental biological process, and then to understand how it is perturbed by regulation in health and dysregulation in disease.

Molecular Biophysics Enzymology Physical Biochemistry

Petrov , A., Groseley, R., Chen, J., O’Leary, S. E., Puglisi, J. D. (2016) Multiple parallel pathways of translation initiation on the CrPV IRES. Mol. Cell 62(1), 92– 103. Choi, J., Ieong, K.-W., Demirci, H., Chen, J., Petrov, A., Prabhakar, A., O’Leary, S. E., Dominissini, D., Rechavi, G., Soltis, M., Ehrenberg, M., and Puglisi, J. D. (2016) N6-methyladenosine in mRNA disrupts tRNA selection and translation elongation dynamics. Nat. Struct. Mol. Biol.23(2), 110–115. Chen, J., Coakley, A., O’Connor, M., Petrov, A. O’Leary, S. E., Atkins, J. F., and Puglisi, J. D. (2015) Coupling of mRNA structure rearrangement to ribosome movement during bypassing of non-coding regions. Cell163(5), 1267–1280. Fuchs, G., Petrov, A. N., Marceau, C. D., Popov, L. M., Chen J., O’Leary, S. E., Wang, R., Carette, J. E., Sarnow, P., and Puglisi, J. D. (2015) Kinetic pathway of 40S ribosomal subunit recruitment to hepatitis C virus internal ribosome entry site. Proc. Natl. Acad. Sci. USA 112(2), 319–325. Chen, J., Petrov, A., Johansson, M., Tsai, A., O’Leary, S. E., and Puglisi, J. D. (2014) Dynamic pathways of -1 translational frameshifting. Nature 512(7514), 328–332. Cooper, L. E., O’Leary, S. E., and Begley, T. P. (2014). Biosynthesis of a thiamin antivitamin. Biochemistry 53(14), 2215–2217. Chen, J., Petrov, A., Dalal, R., Tsai, A., O’Leary, S. E., Chapin, K., Cheng, J., Ewan, M., Hsuing, P.-L., Lundquist, P., Turner, S., Hsu, D. R., and Puglisi, J. D. (2013). High throughput platform for real-time monitoring of biological processes by multicolor single-molecule fluorescence. Proc. Natl. Acad. Sci. USA111(2), 664–669. O’Leary, S. E., Petrov, A., Chen, J., and Puglisi, J. D. (2013) Dynamic recognition of the mRNA cap by Saccharomyces cerevisiae eIF4E. Structure 21(12), 2197–2207. Vivona, S., Cipriano, D., O’Leary, S., Li, Y. H., Fenn, T., and Brunger, A. T. (2013) Disassembly of all SNARE complexes by N-ethylmaleimide-sensitive factor (NSF) is initiated by a conserved 1:1 interaction between a-soluble NSF attachment protein (SNAP) and SNARE complex. J. Biol. Chem. 288(34), 24984–24981. Gokulan, K., O’Leary, S. E., Russell, W. K., Russell, D. H., Lalgondar, M., Begley, T. P., Ioerger, T. R., and Sacchettini, J. C. (2013) Crystal structure of Mycobacterium tuberculosis PKS11 reveals intermediates in the synthesis of methyl-branched alkylpyrones. J. Biol. Chem.288(23), 16484–16494. Hicks, K. A., O’Leary, S. E., Begley, T. P., and Ealick, S. E. (2013) Structural and mechanistic studies of HpxO, a novel flavin adenine dinucleotide-dependent urate oxidase from Klebsiella pneumoniae. Biochemistry 52(3), 477–487. Paul, D., O’Leary, S. E., Toms, A., Rajashankar, K., Bu, W., Settembre, E. C., Sanders, J. M., Begley, T. P., and Ealick, S. E. (2010), Glycal formation in crystals of uridine phosphorylase. Biochemistry49, 3499–3509. O’Leary, S. E., Hicks, K. A., Ealick, S. E., and Begley, T. P. (2009) Biochemical characterization of the HpxO enzyme from Klebsiella pneumoniae, a novel FAD-dependent urate oxidase. Biochemistry48(14), 3033–3035. O’Leary, S. E., Jurgenson, C. T., Ealick, S. E., and Begley, T. P. (2008) O-Phospho-l-serine and the thiocarboxylated sulfur carrier protein CysO-COSH are substrates for CysM, a cysteine synthase from Mycobacterium tuberculosis. Biochemistry47(44), 11606–11615.