Associate Professor

Ph.D., University of Arkansas for Medical Sciences
B.S., Henderson State University

E-mailrleoff@uams.edu
Office:  (501)686-8343 – Biomedical Research Center 1 B421E
Lab:  (501)603-1004 – Biomedical Research Center 1 B416
FAX:  (501)686-8169

The “reading” and “writing” of the genetic code underlies many facets of life at the molecular level. The processes of evolution, cellular adaptation/differentiation, and many aspects of pathogenesis are dependent upon how enzymes recognize and process DNA and RNA. Our group is interested in applying quantitative biochemical and biophysical measurements to the study of enzymes and macromolecular complexes that engage nucleic acids. The major focus of our research is in the study of proteins that are involved in DNA damage tolerance and how these elements contribute to genomic instability. The mechanism of action for Y-family DNA polymerases and interactions that are important for facilitating their biological functions are of particular interest to our group. These specialized polymerases appear to be important for preventing replication fork stalling in the face of damaged template bases or unusual secondary structures (e.g. G4 tetraplex structures, hairpins, and/or Z-DNA). We are interested in understanding at the atomic scale how these enzymes catalyze DNA synthesis when other polymerases are inhibited and how protein-protein interactions between the Y-family polymerases and other replisome-associated proteins, such as the Werner’s syndrome protein, function to alter the efficiency and outcome of specialized DNA synthesis events. Because of their association with various cancers, as well as the unique structural and functional properties used by these enzymes, we are attempting medium-throughput screening of chemical libraries to identify small molecules and/or nucleoside analogues that specifically modulate Y-family DNA polymerase activity.

In addition to understanding the kinetic and structural features defining specialized DNA synthesis events, we are motivated to investigate how DNA damage tolerance mechanisms contribute to the initiation and progression of highly malignant primary brain tumors. Recent evidence has shown that both low- (i.e. grade II) and high-grade (i.e. grades III and IV) primary gliomas exhibit constitutively active DNA damage signaling. The relevance of Y-family DNA polymerases to glioma development was highlighted by a study reporting that two Y-family DNA polymerases (pol iota and kappa) were over-expressed in glioma specimens from a cohort of Chinese patients. One of these enzymes (pol kappa) was shown to be a statistically significant prognostic indicator for survival (i.e. more enzyme yielded a shorter survival time). We are currently performing proteomic analysis of glioma specimens obtained at UAMS, with a specific focus on enriching for DNA damage response elements and identifying changes in the PTMs associated with replication fork stress. These results will provide valuable insight into how damage signaling is altered in primary brain tumors as the lesion progresses to the highly malignant and invasive glioblastoma multiforme (grade IV) and may help us better understand and perhaps attenuate cellular mechanisms that contribute to chemo- and radio-resistance.

The core techniques employed by our group use biochemical/molecular biological approaches with particular emphasis on structural and functional analysis of enzymes and macromolecular complexes involved in the pathways described above. We study the kinetic and biophysical characteristics of protein-protein and protein-nucleic acid complexes using rapid chemical quench and stopped-flow methods, as well as x-ray crystallography and mass spectrometric-based methods, such as hydrogen-deuterium exchange mass spectrometry. Cell-culture experimental systems are also employed to address critical questions concerning the involvement of specific polymerases in mutagenic events, macromolecular complex assembly/function (i.e. replication fork dynamics and RNA processing machinery) and in testing the efficacy of compounds that modulate the activity of specific enzymes. For a more detailed discussion of research objectives feel free to contact me by phone, email or visit us at www.eofflab.org

 

Selected Publications

Eoff, R.L., McGrath, C.E., Maddukuri, L., Salamanca-Pinzon, G.S., Marquez, V.E., Marnett, L.J., Guengerich, F.P., and Egli, M. (2010) “Selective modulation of DNA polymerase activity by fixed conformation nucleoside analogues” Angew. Chem. Int. Ed. Engl. 49, 7481-7485

Eoff, R.L., Choi, J-Y., and Guengerich, F.P. (2010) “Mechanistic studies with DNA polymerases reveal complex outcomes following bypass of DNA damage” J. Nucleic Acids 2010

Maddukuri, L., Eoff, R.L., Choi, J-Y., Rizzo, C.J., Guengerich, F.P., and Marnett, L.J. (2010) “In vitro bypass of the major malondialdehyde- and basepropenal-derived DNA adduct by human DNA Y-family polymerases kappa, iota, and Rev1” Biochemistry 49, 8415-8424.

Eoff, R.L., and Raney, K.D. (2010) “Kinetic mechanism for DNA unwinding by multiple molecules of Dda helicase aligned on DNA” Biochemistry 49, 4543-4553.

Eoff, R.L., Ponce-Sanchez, R., and Guengerich, F.P. (2009) “Conformational changes during nucleotide selection by Sulfolobus solfataricus DNA polymerase Dpo4” J. Biol. Chem. 284, 21090-21099.

Irimia, A.*, Eoff, R.L.*, Guengerich, F.P., and Egli, M. (2009) “Structural and functional elucidation of the mechanism promoting error-prone synthesis by human DNA polymerase – opposite the 7,8-Dihydro-8-oxo-2′-deoxyguanosine adduct” J. Biol. Chem. 284, 22467-22480