The broad, long term goal of this research is to elucidate the role of conformational change in catalysis and enzyme regulation. Our research will emphasize the allosteric regulation and inter- and intramolecular communication of riboucleotide reductases (RNRs). These amazing enzymes utilize organic radical-based chemistry to convert nucleotides to deoxynucleotides. Allosteric regulation of RNR enzymes serves to maintain an appropriate supply of each of four deoxynucleotides (dNTPs) needed for DNA replication and repair. As a result of these cellular activities, RNRs are actively pursued as targets for antibacterial, antiviral, and antitumor therapies. Gemcitabine (2',2'-difluoro-2'-deoxycytidine), a mechanism-based inhibitor of RNRs, was recently approved by the FDA for treatment of pancreatic cancer and demonstrates the medical importance of these enzymes. The proposed research is intended to identify the conformational changes involved in RNR re-reduction by thioredoxin/thioredoxin reductase/NADPH, to identify the role of conformational change in the allosteric regulation of radical formation, and to identify the structural basis for allosteric regulation of enzyme specificity. These goals will be achieved by crystallographic analysis of the least complex members of the ribonucleotide reductase family, the B12-dependent class II enzymes. We will complement structural data with biochemical characterization and single crystal spectroscopic techniques.
| Setser, Jeremy W; Lingaraju, Gondichatnahalli M; Davis, C Ainsley et al. (2012) Searching for DNA lesions: structural evidence for lower- and higher-affinity DNA binding conformations of human alkyladenine DNA glycosylase. Biochemistry 51:382-90 |
| Lingaraju, Gondichatnahalli M; Davis, C Ainsley; Setser, Jeremy W et al. (2011) Structural basis for the inhibition of human alkyladenine DNA glycosylase (AAG) by 3,N4-ethenocytosine-containing DNA. J Biol Chem 286:13205-13 |
| Blasiak, Leah C; Drennan, Catherine L (2009) Structural perspective on enzymatic halogenation. Acc Chem Res 42:147-55 |
| Ryan, Katherine S; Drennan, Catherine L (2009) Divergent pathways in the biosynthesis of bisindole natural products. Chem Biol 16:351-64 |
| Kulik, Heather J; Blasiak, Leah C; Marzari, Nicola et al. (2009) First-principles study of non-heme Fe(II) halogenase SyrB2 reactivity. J Am Chem Soc 131:14426-33 |
| Wong, Cintyu; Fujimori, Danica Galoni?; Walsh, Christopher T et al. (2009) Structural analysis of an open active site conformation of nonheme iron halogenase CytC3. J Am Chem Soc 131:4872-9 |
| Ryan, Katherine S; Chakraborty, Sumita; Howard-Jones, Annaleise R et al. (2008) The FAD cofactor of RebC shifts to an IN conformation upon flavin reduction. Biochemistry 47:13506-13 |
| Hamill, Michael J; Chobot, Sarah E; Hernandez, Hector H et al. (2008) Direct electrochemical analyses of a thermophilic thioredoxin reductase: interplay between conformational change and redox chemistry. Biochemistry 47:9738-46 |
| Ryan, Katherine S; Balibar, Carl J; Turo, Kaitlyn E et al. (2008) The violacein biosynthetic enzyme VioE shares a fold with lipoprotein transporter proteins. J Biol Chem 283:6467-75 |
| Hernandez, Hector H; Jaquez, Orlando A; Hamill, Michael J et al. (2008) Thioredoxin reductase from Thermoplasma acidophilum: a new twist on redox regulation. Biochemistry 47:9728-37 |
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