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Research

Summary

The research conducted in our laboratory focuses on biological and physical organic chemistry. In the biological chemistry area, the reactivity of nucleobases and nucleotides, including enzymes for which nucleic acids are substrates, is explored using methods such as mass spectrometry (MS) and computer modeling. Goals of this research include the improved understanding of DNA reactivity and enzyme catalysis toward the ultimate end of improved drug design. In the physical organic area, synthesis, MS, and theoretical methods are used to explore pericyclic rearrangements. Goals include identifying the root of acceleration in various reactions, toward the end of increasing the synthetic utility of these rearrangements.

DNA DUPLEX STABILITY

The factors, including hydrogen bonding, pi-pi stacking, and solvation, which control the stability of duplex DNA are under investigation. State-of-the-art experimental techniques, including electrospray ionization, liquid chromatography mass spectrometry (LCMS) and Fourier transform mass spectrometry (FTMS), are used to explore the binding affinities of DNA double-stranded oligomers in the gas phase, in the absence of solvent. Because solvation plays a role in stabilizing the double helix, from affecting base pairing to mediating binding events, the study of naked DNA to examine intrinsic behavior is of fundamental importance. Our overall goal is twofold: 1) to undertake the systematic examination of DNA base pairs, base-stacked dimers, and longer oligonucleotides in order to lend insight into the intrinsic stability and behavior in the absence of solvent; and 2) to establish a novel method for the rapid assay of gas-phase and solution-phase binding affinities using mass spectrometry. Since biological processes are regulated, fundamentally, through specific DNA-protein recognition, and DNA protein recognition is controlled by DNA conformation, the study of duplex DNA structure and stability is highly significant.

ENZYME CATALYSIS

The mechanisms by which enzymes catalyze reactions are of interest from a purely scientific, physical organic perspective, but also from an applied point of view: understanding an enzyme mechanism has implications for inhibitor design and potentially, rational drug design. Two nucleotide-related reactions that are particularly suitable for directed computational and gas-phase experimental studies are the decarboxylation of orotidine 5-monophosphate (OMP) and the cleavage of an erroneously inserted uracil in DNA. The decarboxylation of OMP is catalyzed by a highly proficient enzyme, OMP decarboxylase; the mechanism remains unknown. Computational and experimental studies are used to uncover the mechanisms by which the enzyme can effect catalysis. Uracil-DNA glycosylases cleave the uracil-ribose bond in DNA (where a deoxyuridine has been erroneously inserted). The study of the mechanism can be reduced to a question of the acidity of uracil. The effects of protonation and nearby cations on uracil acidity are currently being explored computationally, while the gas-phase acidity of uracil is being examined experimentally.

PERICYCLIC REARRANGEMENTS

The effect of media on reactivity is a classic concern for organic chemists. Whether one is considering the solvent in which to conduct a reaction, or the effect of a low dielectric interior in an enzyme active site, the medium has profound effects on reactivity and binding. The elimination and decarboxylation reactions of benzisoxazoles and 3-carboxybenzisoxazoles are extremely medium-sensitive, and occur more rapidly in aprotic solvents. A combined experimental and theoretical investigation is being undertaken to explore these reactions in the gas phase and with individual-molecule solvation in order to understand how solvation and biological catalysis affect reaction energetics and transition structures. Medium also plays a role in the rearrangements of anionic Cope substrates. Anion-accelerated Cope rearrangements, of which the anionic oxy-Cope is the most well-known, are predicted to proceed very rapidly in the gas phase. Probe reagents are used that can differentiate between the starting material and product, which have the same m/z ratio, in order to discern whether rearrangement has occurred. Theory is utilized to explore transition structures and to lend insight into how and why these reactions experience such great acceleration.

Awards & Honors

Wepukhulu, W. O.; Smiley, V. L.; Vemulapalli, B.; Smiley, J. A.; Phillips, L. M.; Lee, J. K. "Evidence for Pre-Protonation in the Catalytic Reaction of OMP Decarboxylase: Kinetic Isotope Effects using the Remote Double Label Method," Biochemistry 2007, submitted.

Sun, X.; Lee, J. K. "The Acidity and Proton Affinity of Hypoxanthine in the Gas Phase versus in Solution: Intrinsic Reactivity and Biological Implications," J. Org. Chem. 2007, 72, 6548-6555.

Tantillo, D. J.; Lee, J. K. "Reaction Mechanisms: Pericyclic Reactions," Annu. Rep. Prog. Chem., Sect. B. 2007, 103, 272-293.

Pan, S.; Sun, X.; Lee, J. K. "DNA Stability in the Gas versus Solution Phases: A Systematic Study of Thirty-One Duplexes with Varying Length, Sequence, and Charge Level," J. Am. Soc. Mass Spectrom. 2006, 17, 1383-1395.

Pan, S.; Sun, X.; Lee, J. K. "Stability of Complementary and Mismatched DNA Duplexes: Comparison and Contrast in Gas versus Solution Phases," Int. J. Mass Spectrom. 2006, 253, 238-248.

Pan, S.; Verhoeven, K.; Lee, J. K. "Investigation of the Initial Fragmentation of Oligodeoxynucleotides in a Quadrupole Ion Trap: Charge Level-Related Base Loss," J. Am. Soc. Mass Spectrom. 2005, 16, 1863-1865.

Phillips, L. M; Lee, J. K. "Theoretical Studies of the Effect of Thio Substitution on Orotidine Monophosphate Decarboxylase Substrates," J. Org. Chem. 2005, 70, 1211-1221.

Lee, J. K. "Insights into Nucleic Acid Reactivity through Gas Phase Studies," Int. J. Mass Spectrom. 2005, 240, 261-272.

Sharma, S.; Lee, J. K. "Gas Phase Acidity Studies of Multiple Sites of Adenine and Adenine Derivatives," J. Org. Chem. 2004, 69, 7018-7025.

Lee, J. K., Editor. "Orotidine Monophosphate Decarboxylase: A Mechanistic Dialogue," Topics in Current Chemistry 2004.

Lee, J. K.; Tantillo, D. J. "Computational Studies on the Mechanism of Action of Orotidine Monophosphate Decarboxylase," Adv. Phys. Org. Chem. 2003, 38, 183-218.

Haeffner, F.; Houk, K. N.; Schulze, S. M.; Lee, J. K. "Concerted Rearrangement versus Heterolytic Cleavage in Anionic [2,3]- and [3,3]-Sigmatropic Shifts. A DFT Study of Relationships Between Anion Stabilities and Mechanisms and Rates,"J. Org. Chem. 2003, 68 2310-2316.

Kurinovich, M. A.; Phillips, L. M.; Sharma, S.; Lee, J. K. "The Gas Phase Proton Affinity of Uracil: Measuring Multiple Basic Sites and Implications for the Enzyme Mechanism of Orotidine 5-Monophosphate Decarboxylase," Chem. Commun. 2002, 2354-2355.

Sharma, S.; Lee, J. K. "The Acidity of Adenine and Adenine Derivatives and Biological Implications. A Computational and Experimental Gas Phase Study," J. Org. Chem. 2002, 67, 8360-8365.

Kurinovich, M. A.; Lee, J. K. "The Acidity of Uracil and Uracil Analogs in the Gas Phase: Four Surprisingly Acidic Sites and Biological Implications", J. Am. Soc. Mass. Spectrom. 2002, 13, 985-995.

Phillips, L. M.; Lee, J. K. "Theoretical Studies of Mechanisms and Kinetic Isotope Effects on the Decarboxylation of Orotic Acid and Derivatives," J. Am. Chem. Soc. 2001, 123, 12067-12073.

Schulze, S. M.; Santella, N.; Grabowski, J. J., Lee, J. K. "The Secondary and Tertiary Anionic Oxy-Cope Alkoxides Rearrange in the Gas Phase," J. Org. Chem. 2001, 66, 7247-7253.

Houk, K. N.; Lee, J. K.; Tantillo, D. J.; Bahmanyar, S.; Hietbrink, B. N. "Crystal Structures of Orotidine Monophosphate Decarboxylase: Does the Structure Reveal the Mechanism of Natures Most Proficient Enzyme?," ChemBioChem 2001, 2, 113-118.

Kurinovich, M.A.; Lee, J, K. "The Acidity of Uracil from the Gas Phase to Solution: The Coalescence of the N1 and N3 Sites and Implications for Biological Glycosylation," J. Am. Chem. Soc. 2000,122, 6258-6262.

Singleton, D. A.; Merrigan, S. R.; Kim, B. J.; Beak, P.; Phillips, L. M.; Lee, J. K. "¹³C Kinetic Isotope Effects and the Mechanism of the Uncatalyzed Decarboxylation of Orotic Acid," J. Am. Chem. Soc. 2000, 122, 3296-3000

Chen, J.; McAllister, M. A., Lee, J. K., Houk, K. N. "Short, Strong Hydrogen Bonds in the Gas Phase and in Solution: Theoretical Exploration of pK_a Matching and Environmental Effects on the Strengths of Hydrogen Bonds, and their Potential Roles in Enzymatic Catalysis," J. Org. Chem. 1998, 63, 4611-4619.

Yoo, H. Y.; Houk, K. N.; Lee, J. K.; Scialdone, M. A.; Meyers, A. I. "A New Paradigm for Anionic Heteroatom Cope Rearrangements," J. Am. Chem. Soc. 1998, 120, 205-206.

Lee, J. K.; Houk, K. N. "A Proficient Enzyme Revisited: The Predicted Mechanism for Orotidine Monophosphate Decarboxylase," Science 1997, 276, 942-945. Reported in "News of the Week": Rouhi, A. M. "Carbenes May Be Key to Enzymes Power," Chemical and Engineering News 1997, 75, 12.

Lee, J. K.; Houk, K. N. "Cation Cyclization Selectivity: Variable Structures of Protonated Cyclopropanes and Selectivity Control by Catalytic Antibodies," Angew. Chem. Int. Ed. Engl. 1997, 36, 1003-1005.

Houk, K. N.; Lee, J. K. "Physical Organic in the 21st Century: Evanescent or Transcendent?," Pure Appl. Chem. 1997, 69, 237-239.

Houk, K. N.; Beno, B. R.; Nendel, M.; Black, K.; Yoo, H. Y.; Wilsey, S.; Lee, J. K. "Exploration of Pericyclic Reaction Transition Structures with Quantum Mechanical Methods: Competing Concerted and Stepwise Mechanisms," J. Mol. Struct. (Theochem.) 1997, 398-399, 169-179.

Lee, J. K.; Grabowski, J. J. "Anion Structure Determination in the Gas Phase: Chemical Reactivity as a Probe," J. Org. Chem. 1996, 61, 9422-9429.

Wu, Y.-D.; Lee, J. K.; Houk, K. N.; Dondoni, A. "Theoretical Study of a Termolecular Mechanism for the Reaction of (Trimethyl)silylthiazole with Carbonyl Compounds," J. Org. Chem. 1996, 61, 1922-1926.