Abstracts

Keynote Session Abstracts#

Lester Andrews Graduate Research Symposium Keynote

21st Century Solar Army

Harry Gray
California Institute of Technology

The sun is a boundless source of clean energy, but it goes down every night. We and many others are trying to design solar-driven molecular machines that could be used on a global scale to store solar energy by splitting water into its elemental components, hydrogen and oxygen. Hydrogen is a clean fuel that could be used directly or combined with carbon dioxide to produce methanol, a liquid fuel. We are investigating the structures and mechanisms of hydrogen evolving catalysts made from Earth abundant elements such as cobalt, iron, nickel, and molybdenum. We also are employing pulsed laser ablation for synthesis of metal- oxide nanoparticles that will be deployed as catalysts on photoanodes such as tungsten oxide. To aid our research, we have recruited hundreds of students to join a Solar Army whose mission is the discovery of mixed- metal oxides for testing on the photoanodes of our solar water splitters.

Oral Presentation Abstracts#

Nanoscale Patterning of Proteins: Toward Biosensor Development

Perera, Y. Randika,Wang, Ailin, Hughes, Alex, and Fitzkee, Nicholas C.
Department of Chemistry, Mississippi State University, Mississippi State, 39762

Understanding the interactions of gold nanoparticles (AuNPs) with biological macromolecules is becoming increasingly important. This is in part due to the potential applications in drug delivery, bio sensing, diagnostics, and imaging. Our long-term goal is to use protein-functionalized AuNPs as a general tool for molecular sensing and drug delivery. We hypothesize that electrostatic interactions play an important role in the protein-AuNP interaction, since citrate-stabilized AuNPs carry a net negative charge. Using an NMR-based approach developed by our group, we have monitored apparent binding capacity of two similarly structured, but differently charged proteins. Previous work on the GB3 protein hypothesized that positively-charged lysine residues are involved in protein-AuNP binding, and a potential binding site was proposed involving three specific lysine residues, K4A, K19A, and K50A. To test this hypothesis, we mutated the lysine residues to alanine one at a time using site directed mutagenesis. NMR was then used to observe how the binding capacity of these variants changed relative to the wild-type protein. In particular, the K4A, K13A and K50A variants significantly reduced binding, while other variants did not. This result supports our original hypothesis, and suggests that GB3 adopts a specific orientation on the AuNP surface. As the next study we methylated the GB3 protein when it’s bound to AuNP hypothesizing if above mentioned residues are involved in binding, they shouldn’t be methylated. In the future, we believe that experiments like these will result in a better understanding of protein-nanoparticle interactions.

Investigation of the Tsuji-Trost Winstein-Masamune intramolecular phenolic allylation for the synthesis of tricyclic scaffolds found in Lycopodium alkaloids

Nicholas Jentsch, Erin Realini, Dr. Matthew Donahue
Department of Chemistry and Biochemistry, University of Southern Mississippi

The total synthesis of natural products, especially Lycopodium alkaloids, remains a vigorous area of research. With the advancement of new reaction technologies even the most daunting molecular structures are being synthesized with a pace unrivaled in history. Pursuant to these endeavors we have initiated a research program aimed at transforming simple aromatic phenols into increasing complex spirocyclic compounds. The intramolecular para-alkylation of a phenol with a carbon chain bearing a suitable leaving group, known as the Winstein-Masamune reaction, results in a spiro[4.5]deca-6,9-dien-8-one. Of notable consequence is the synthesis of a sterically congested spirocyclic quaternary carbon. This particular motif is a substructure found in the AC rings of Lycopodium alkaloids such as magellanine, which has captured the imagination of many synthetic organic groups as a proving ground for demonstrating new strategies and methods. In this talk, we will discuss the merging of this reaction with the Tsuji-Trost allylic alkylation to afford a spirocycle containing a pendant alkene. In addition, we will discuss the utilization of the alkene functional handle for elaboration of the spiro[4,5]decane to a tricyclic core via free radical conditions. To this end, we are taking an AC to ACB approach to the construction of Lycopodium alkaloids with the spirocyclic quaternary carbon as the focal point.

Capillary Electrophoretic Enzyme Assay of Acetyl-Coenzyme A Carboxylase

Thu H. Nguyen¹, Alexandra Evans², Grover Waldrop², and S. Douglass Gilman^1
1Department of Chemistry, Louisiana State University, Baton Rouge, LA 70803
²Division of Biochemistry and Molecular Biology, Louisiana State University, Baton Rouge, LA 70803

A simple, direct, off-column capillary electrophoretic (CE) assay was developed and optimized to study enzyme activity and inhibition of acetyl-coenzyme A carboxylase (ACC). This enzyme catalyzes the first committed step and rate-limiting step in fatty acid synthesis in bacteria, plants, and animals. Malonyl-CoA, a product of ACC plays a critical role in fatty acid synthesis and regulation of fatty acid oxidation. Therefore, ACC is a potential target for treatment of obesity, diabetes, and other metabolic diseases as well as for antibiotic drug development. In CE assay, spectral interference is minimized by separation of substrates, products, inhibitors, and assay matrix. This assay is unlikely susceptible to inhibitor interference because coupling enzymatic reactions are not needed. The developed CE assay was applied to study inhibition of ACC. Several synthetic and natural compounds that inhibit ACC-catalyzed reaction were identified.

Cooperative binding in boronic acid modified PAMAM dendrimers

Xiaoli Liang, Marco Bonizzoni
Department of Chemistry, The University of Alabama

Cooperativity in molecular binding process of macromolecules with multiple binding sites is essential yet poorly understood. So far, no detailed binding behaviors for compounds with more than four binding sites are well understood. We studied the binding cooperativity in systems with multiple interaction sites, with the intention of developing general models. G3 poly(amidoamine) (PAMAM) dendrimers containing 32 terminal amine groups are ideal for these studies. We first decorated their surface with 3-formylphenylboronic acids to turn them into receptors with multiple boronic acid binding sites. The coverage of boronic acids at the surface was controlled to 28%, 56% and 78% of the available amine groups on the surface, which gave us access to a series of hosts with various cooperativity. Cis-diol groups reversibly bind to boronic acids, so we used pyrogallol red (PG), a dye with two set of cis-diol groups, as a cooperative guest. As the boronated PAMAM receptors were titrated into PG, the binding was monitored via UV-Vis absorption and fluorescence emission spectroscopy. As a control experiment, the PG dye was titrated with a physical mixture of G3 PAMAM dendrimer and different amounts of free 3-formylphenylboronic acids. The binding isotherms for all boronated PAMAM receptors revealed significant cooperativity when compared to the physical mixtures. Much less boronated PAMAM was required to achieve binding saturation than it was the case with the physical mixture, indicating that the covalently bound system has a cooperative advantage. Models for complex binding systems with multiple binding sites will be built up for further studies.

Discrimination of metal cation mixtures in water

Michael Ihde and Marco Bonizzoni
Department of Chemistry, The University of Alabama

Many metal ions are known to have severe health and environmental impacts, particularly in aqueous environments. Therefore, we have investigated the detection of divalent metal ions in water using array sensing techniques.

A multivariate dataset obtained by exposing an array containing two water soluble commercially available dyes (xylenol orange and methylthymol blue) to multiple analytes was interpreted using pattern recognition algorithms such as linear discriminant analysis (LDA). The array has been shown to discriminate nine divalent metal cations with excellent resolution. We have also optimized our array by taking advantage of responses from a variety of metal-dye stoichiometries to map metal ion concentrations of four environmentally relevant divalent metal ions (Hg^II, Pb^II, Cd^II, and Cu^II) quantitatively at concentrations as low as 1 μM in neutral water.

Encouraged by the excellent results obtained with single metals using our array, we also successfully used our array to discriminate binary and ternary metal cation mixtures, a particularly valuable accomplishment for a pattern recognition system, which otherwise typically struggle with the “problem of mixtures”. To validate our array’s utility as a calibration plot, we have displayed a projection of unknown metal cation mixtures on our original scores plot with exceptional predictive ability.

Radical Chain Reduction of CCl₄ Initiated by Illumination of SPEEK Solutions

Md S Islam, J. R. Black and G. Mills
Department of Chemistry and Biochemistry, Auburn University, Alabama, 36849, United States

Illumination of sulfonated poly(ether etherketone), SPEEK, together with polyvinyl alchohol, PVA or sodium formate in aqueous solution have been found to effectively reduce CCl₄ to Cl^–. The photoreduction process involves polymeric α–hydroxyl (SPEEK•) and CO₂^– radicals where the polyketone acted as a sensitizer and PVA or HCO₂^– served as H-tom donor. Formation of Cl^– proceeded via a zero-order rate law for extended times after an initial short induction period. For air-free solutions r(Cl-) increased slightly with light intensity (I₀) whereas the quantum yields of Cl^– generation, 𝜙(Cl^–), decreased with increasing I₀. Experiments in which irradiation was periodically interrupted showed that post-irradiation formation of Cl^– took place. In most cases, the quantum yields, 𝜙(Cl^–) obtained in solutions with HCO₂^– ions were between 10 to 15 times higher than the 𝜙(Cl^–) values determined in solutions of PVA as HCO₂^– ion produce CO₂^– radicals which reduces CCl₄. Highest values of 𝜙(Cl^–) were determined at 6 ≤ pH ≤ 8, coinciding with the range where SPEEK• is most efficiently photogenerated. However, a sharp maximum of (Cl^–) was determined at a pH of 7.3; DSC results suggest that aggregation of the polyelectrolyte under such conditions may influence the photoreaction.

The anions in the citric acid cycle: tackling the differentiation of minimally diverse species using hyperbranched polymeric receptors

Yifei Xu, Marco Bonizzoni
Department of Chemistry, The University of Alabama, Tuscaloosa, AL 35487-0336

The carboxylic acids present in the citric acid cycle have similar structures and are thus difficult to differentiate, so they present an interesting analytical challenge. Our group has had previous success in using poly(amidoamine) (PAMAM) dendrimers as supramolecular hosts for this class of compounds. PAMAM dendrimers are water-soluble globular hyperbranched polymers that, at neutral pH, display a homogeneous array of primary ammonium groups on their surface. Such hosts can establish electrostatic and H-bonding interactions with a variety of anionic species, among which organic carboxylates. Such interactions, however, are typically hard to use analytically because neither the carboxylate analytes nor the PAMAM receptors have detectable signals in optical spectroscopy (e.g. absorbance, fluorescence techniques).

In this study, we circumvented this problem by utilizing a dye-displacement assay based on PAMAM-dye complexes to sense these biologically relevant carboxylate anions. Analytical data obtained from an array of such sensors was then interpreted using pattern recognition algorithms (e.g. principal component analysis) to recover discriminatory power. In neutral HEPES buffer, PAMAM dendrimer and calcein blue dye were first bound. Then, a carboxylate was introduced, which caused dye displacement from its complex with PAMAM. The displacement was monitored through absorbance, fluorescence intensity and anisotropy. Principal component analysis (PCA) was then used for data interpretation, to discriminate among these carboxylate analytes. We are currently working to expand the sensing array to improve selectivity, by the addition of a second fluorescent probe.

Poster Presentation Abstracts#

Synthesis of Novel and Sustainable (Bio-Based) Poly(Ether Amides)

C. Scott*, G. Saenz
*Department of Chemistry, Mississippi State University, Mississippi State, United States*

Petroleum-based products have dominated the commercial market for many decades; however, in recent years, there have been a push to replace petroleum-based products with those from renewable sources due to the unsustainability of petroleum source and the continued rising cost of petroleum. Novel and sustainable poly(ether amide)s were synthesized by melt polycondensation of two lignin-derived monoaromatic compounds: methyl 3-methoxy-4-hydroxybenzoate and methyl 4-hydroxybenzoate with 2-chloroethylamine. This presentation describes the design, synthesis and properties (thermal and physical) of our new poly(ether-amide)s as alternative thermoplastics from lignin-based raw materials. Our results show that our polymers’ glass transition temperatures (Tg) range from 96 °C – 100 °C, which is similar to hard plastic polymers such as polystyrene and poly(methyl methacrylate) with Tg ~ 100 °C. The melting points (mp) (245 °C – 255 °C) were more similar to polystyrene ~ 240 °C and higher than poly(methyl methacrylate) at 160 °C. We explored the structure/property relationship with respect to hydrogen bonding and its effects on the thermal and physical properties of these polymers. It was shown that hydrogen bond interactions appear to play an important role to define the thermal properties of the two poly(ether amide)s synthesized. In particular, the poly(ether amide) having methyl 3,4-dihydroxybenzoate as monomer exhibits the best thermal behavior.

How an Arginine Switch Preserves the Catalase Activity of KatG: Strategic Use of an Active Site Tryptophan for off-pathway Electron Transfer

Hui Xu, Jessica R. Krewall, Olive Njuma and Douglas C. Goodwin
Departments of Chemistry and Biochemistry, Auburn University, Alabama, AL 36830

KatG catalase activity is prone to inactivation due to off-pathway intramolecular electron transfer, beginning with radical migration from the enzyme’s heme to its proximal tryptophan (W321). Inclusion of a peroxidatic electron donor (PxED) mitigates against the accumulation of inactive states. All KatGs have a conformationally dynamic arginine (R418) whose position affects the electronic structure of the KatG active site. To investigate the impact of this arginine on off-pathway radical transfer and enzyme inactivation, we compared the properties of R418N and W321F/R418N variants to wild-type KatG (wtKatG). Like wtKatG, R418N underwent inactivation prior to H₂O₂ depletion, and catalase activity could be retained by inclusion of a PxED; however, depending on H₂O₂ concentration, R418N produced two- to 75-fold more oxidized PxED during O₂ production, suggesting a greater frequency of PxED-dependent rescue events. W321F/R418N exhibited much greater vulnerability to inactivation than R418N, even with a PxED included. W321F/R418N peroxidase activity far exceeded wtKatG, suggesting an uncoupling of KatG’s two major activities. For all variants, the k_obs values describing the decrease in O₂ production over time corresponded closely to the loss of PxED-rescue-able activity, indicating that initial formation of reversibly inactive states is rate-determining in irreversible inactivation. Our data suggest that R418 limits off-catalase electron transfer in wtKatG. However, even without R418, radical transfer via W321 produces intermediates rescue-able by PxEDs. When both R418 and W321 are absent, more frequent off-catalase electron transfer is combined with KatG oxidation that cannot be resolved by inclusion of exogenous electron donors.

Silyloxyallyl Cation: Synthesis of Functionalized 1,4-Diketones and Substituted Pyrroles

Joshua A. Malone, Joshua P. Van Houten, Moshood Ganiu, Binod Nepal, Rendy Kartika
Louisiana State University

The relevance of 1,4-diketones in various natural products and pharmaceutical relevant motifs, including heterocyclic compounds that make diketones an important synthetic target in organic synthesis. In this presentation we showcase a methodology for nucleophilic capture of unsymmetrical silyloxyallyl cations with various silylenol ethers to afford differentiated 1,4-diketones in a regioselective manner. The furnished 1,4-diketones allows for the subsequent three component, one pot synthesis of substituted pyrroles.

Fluorescence quenching of aromatic polymers with small PAHs

Nicholas J. White¹; Joshua Tropp²; Jason Azoulay²; Marco Bonizzoni¹*
¹*Department of Chemistry, The University of Alabama*
²*School of Polymers and High Performance Materials, The University of Southern Mississippi*

Polycyclic aromatic hydrocarbons (PAH) are an environmentally important class of molecules, which have been repeatedly introduced into waterways via e.g. petroleum spills and combustion byproducts. These molecules often have characteristic absorption and fluorescence spectra, however low concentrations can make differentiation and quantitation challenging. The conjugated core of these hydrocarbons can also interact with fluorescent moieties containing complementary aromatic systems to afford a method of measurement and detection through modulation of fluorescence emission.

We developed a small series of aromatic polymers that can bind small PAH molecules. These polymers contain a fluorene backbone, with variations occurring in the spacing in the aromatic core, as well as different substituents branching off the backbone, including a phenylbenzimidazole group and polyethylene glycol chains of varying length. Introducing aliquots of PAH into polymer solutions in DMF gradually quenches the fluorescence of these polymers, to different extents and at different molar ratios. The ability of small PAH molecules to quench the fluorescence of these polymers is not only a function of the orbital energy level matching, but it also depends on the relative sizes of the binding site and of the hydrocarbons.

In addition to benchtop titrations, these systems will be placed on a microwell plate to observe binding, and Principal Component Analysis (PCA) will be used to differentiate small PAH in solution. The data from these binding studies is being shared with computational collaborators as part of a larger effort to enhance binding models for improved sensor development.

Structural Study of Abruptex Region in Mouse Notch1 by Nuclear Magnetic Resonance Spectroscopy (NMR)

Huimin Zhong, Megan A. Macnaughtan
Department of Chemistry, Louisiana State University, Baton Rouge, LA 70803

Notch signaling is associated with various developmental disorders in humans, including neurodegenerative diseases and cancer. The Notch proteins of Notch signaling are intracellular proteins. In mouse Notch1, the Abruptex region containing epidermal growth factor-like (EGF) repeats 24-29 in the extracellular part, is a highly glycosylated and flexible region, which regulates ligand binding specificity. The long-term objective of this research is to determine the high-resolution 3D structure of mouse Notch1 Abruptex EGF26-28, and investigate the effect of glycosylation on the structure and dynamics using nuclear magnetic resonance (NMR) spectroscopy.

EGF27 was selected as our first domain of interest because of its uniqueness in the Abruptex region that, it contains both O-glucosylation and O-fucosylation sites. Different DNA sequences of EGF27 and E. coli cell lines were tested for a high expression of the target protein. The 7aa-EGF27-His was selected for the expression in either Origami B or Rosetta-gami B E. coli cells. Since a large part of the proteins stayed in the inclusion bodies, three detergents (sarkosyl, Triton X-100, and CHAPS) were applied to extract the proteins. The soluble protein and re-solubilized protein samples were characterized by ¹⁵N-HSQC NMR and mass spectrometry, which showed that the re-solubilized protein likely contained non-native disulfide bonds, and the proteins may be degraded. Refolding with glutathione (GSH) produced disulfide bond linkages similar to the soluble protein sample.

Continuing efforts on the structural study of EGF27 and other EGF repeats in the Abruptex region, with and without glycosylation, will be the future work.
Surface-initiated short chain ATRP of substituted styrenes on SiO₂ – A step towards Engineered Soil Surrogates (ESS)

Arjun Pandey, Stephen Smith, Benjamin Haywood, Balamurugan Subramanian, Robert Cook, David Spivak
Department of Chemistry, Louisiana State University

Immobilization of polymers onto surfaces can be accomplished by either growing a polymer chain from initiator functionalized surface known as “grafting from” or growing a polymer chain with a surface-attachable end functional group and then attaching it to the surface known as “grafting onto”. Growing a short oligomer with a narrow distribution requires a very specific set of conditions because the controlled living radical polymerization behaves as an uncontrolled polymerization at the initial stage of the reaction. Here, we explore the “grafting from” approach to polymerize short chains of 2-10 monomers of styrene and substituted styrenes on SiO₂ gel using Atom Transfer Radical Polymerization (ATRP) in an attempt to mimic the structure of natural soil with complex interaction among hydrophilic, hydrophobic groups and external components. The primary objective of this study is to determi ne how the molecular level chemical composition of the domains within the organic matter influences the macroscopic sorption, desorption, and competition of model probe compounds of varying polarity. A series ESSs of increasing complexity will be used in concert with sorption isotherm data obtained by batch mode experiments.

Nanopatterns of Phthalocyanines Prepared Using Particle Lithography Characterized with Atomic Force Microscopy

Kuruppu Arachchige, Neepa M K; Taylor, Ashley M; Okoth, Elizabeth A; Vicente, Graca; Garno, Jayne C.
Department of Chemistry, Louisiana State University, Baton Rouge, LA

Phthalocyanines (PCs) are a group of synthetic tetrapyrrolic compounds that have promising properties of photosensitivity due to intense absorption in the red region of visible light. Incorporation of diamagnetic metal ions such as zinc into PCs enhances the photosensitive properties by providing high triplet state quantum yield and longer lifetime. Thus, metal-PCs have been used in technologically advanced applications such as electrophotography, photoelectronic and photodynamic therapy. Researchers have begun to develop methods to form thin films of PCs on substrates as a key component of nanoscale devices. Efficient patterning methods at the nanoscale are needed for advancing applications of PCs. In this presentation, self-assembled monolayers of organosilanes were used to prepare nanoscale surface platforms using immersion particle lithography to prepare nanostructures. Holes within the organosilane films were used to direct the attachment of Zn-PC molecules to Si(111). The surface chemistry and morphology of surface platforms of Zn-PCs was investigated at the molecular-level using atomic force microscopy (AFM). An array of nanopores within a matrix of octadecyltrichlorosilane (OTS) was generated as a platform to confine the surface assembly of 3-aminopropyltriethoxysilane (APTES). A protocol was then developed to direct the addition of Zn-PC compounds onto active sites of the APTES nanopatterns. High resolution AFM were used for surface characterizations at each of the key steps of the experiment to characterize samples.

Development of an Expression System for the Investigation of Cobalamin-dependent Radical S-Adenosyl-L-methionine Enzymes Involved in Bacteriochlorophyll c Biosynthesis

Xingchen Huang and Steven O. Mansoorabadi
Department of Chemistry and Biochemistry, Auburn University, Auburn, AL 36849-5312

Green sulfur bacteria (GSB) produce a specialized photopigment, bacteriochlorophyll (Bchl) c, which enables them to grow phototrophically under extreme low-light intensity conditions. Bchl c has several unique structural features that allow it to self-assemble into the large nanostructures that comprise the highly efficient light-harvesting complex of GSB, the chlorosome. The biosynthetic pathway of Bchl c has yet to be fully characterized. Based on genetic information, three key reactions in Bchl c biosynthesis are thought to be catalyzed by homologous enzymes (BchE, BchQ, and BchR), which are annotated as cobalamin (B12)-dependent members of the radical S-adenosyl-L-methionine (SAM) superfamily. BchE catalyzes anaerobic O atom insertion and formation of the isocyclic ring of Bchl c. In contrast, BchQ and BchR are unusual methyltransferases capable of methylating unactivated C-H bonds. Specifically, BchQ transfers up to 3 methyl groups to the C8 ethyl group of Bchl c, while BchR catalyzes monomethylation of the C-12 methyl group. BchQ and BchR are expressed in a soluble form in the heterologous host Escherichia coli BL21(DE3), but the proteins are not purified in the holo form, even when grown in the presence of exogenous B12 and additional iron/sulfide. An expression system was therefore developed that simultaneously enhances the intracellular concentration of B12 and aids in the reconstitution of iron-sulfur clusters that can be used for the production of any cobalamin- and/or iron-sulfur cluster-dependent enzyme.

Predicting the Properties of High Oxidation State AnF_x Complexes

Zachary Lee, Monica Vasiliu, Emma Clements, Julia Kessel, and David A. Dixon
Department of Chemistry, The University of Alabama, Shelby Hall, Tuscaloosa, Alabama 35487-0336, USA

There is intense interest in compounds with metals in formal high oxidation states, especially the properties of fluorides for molten salt reactors. These AnF_x complexes are being studied to provide further insights into periodic trends as well as for the design of new separation systems for next generation nuclear fuels. Computational chemistry electronic structure approaches are being used to predict the structures and the vibrational frequencies of neutral, anionic, and cationic actinide fluorides AnF_x where An = Th - Cf to determine the maximum formal oxidation state of the actinide. The different oxidation states of these actinide fluorides together with varying high spin states of each complex were studied using DFT (density functional theory) with the B3LYP exchange-correlation functional. The TZVP basis set was used for F and cc-pvdz-pp basis sets were used for Th, Pa and U and the Stuttgart basis sets with effective core potentials were used for the actinides of Np – Cf. An NBO analysis was performed on each of the complexes and various energetics were calculated for each complex including the electron affinity, ionization energy, bond dissociation energy, fluoride affinity, and fluorocation affinity to provide information on redox properties and Lewis acidities. This work was supported by the US Department of Energy, Office of Science, Basic Energy Sciences.

Enhanced Ionization of Peptides using Cr(III): Developing the Basis for Proteomics Studies

Rudradatt (Randy) Persaud, Christena Rogers, Tyler Crawford, Carolyn J. Cassady, and David A. Dixon
Department of Chemistry, The University of Alabama Tuscaloosa, Alabama 35487-0336

The goal of the Human Proteome Project is to elucidate all of the proteins in the human body. One common, cost effective and reliable approach is the use of tandem mass spectrometry as it is a highly reproducible analytical technique. It has been shown that the addition of Cr(III) to peptide solutions being electrosprayed has great promise for expanding the range of acidic and basic peptides that can be studied by positive ion mass spectrometry and also for increasing the sensitivity and accuracy of the analysis. Computational chemistry is being used to study the interactions of Cr(III) with neutral and anionic (parent –H anions). By calculating the enthalpy of the reaction of the peptide anions and the neutral species, we are improving our understanding of how chromium cations interact with these species, providing structural and energetic information not available from mass spectrometry experiments. The calculations are done using density functional theory with the B3LYP exchange-correlation functional. We are also studying the diprotonated species as part of our study to see how Cr(III) in solution can enhance such protonation states. We have already discovered differences that are dependent on the structure of the side chain. This work is supported by the National Science Foundation.

Enhanced Calculated bond dissociation energies of simple peptides for use in analyzing collision induced dissociation processes

Ashley S. McNeill,* Can Cui,a Sean Miller, Michele Stover, Carolyn J. Cassady, and David A. Dixon
*The University of Alabama Tuscaloosa, AL*

Collision induced dissociation (CID) is often used for primary ion identification in the mass spectrometric analysis of proteins. Computational electronic structure methods have been employed to predict the energetics of possible fragmentation pathways and product identification. The gas phase acidities of amino acids and peptides at the correlated G3(MP2) molecular orbital theory level agree with the experimental values to within about ±2 kcal/mol and with Feller-Peterson-Dixon calculations. The calculations enable the prediction of the structures of the anions and neutrals including rearrangement processes. Specific results include: (1) The y1- anion NH₂CHRCO₂^– is the most intense product ion for dipeptides and their amides; (2) Loss of H₂O is observed for alanine, diglycine, and dialanine amide precursor anions; (3) Loss of CO₂ is only observed for dimers; (4) Among amide species, N≡C–O– was universally observed and the enthalpies for generation of this anion are less than ±5 kcal/mol; (5) The hydrogen at the C-terminus of the mono- and dipeptide amide molecules facilitated the observable loss of H₂O in the experimental spectra as loss of H₂O was not observed for non-amide species; and (6) Many predicted low energy pathways are not observed in the CID mass spectra. This work was supported by the National Science Foundation.

Directing off-pathway protein oxidation to preserve enzyme activity: At last, a role for the proximal tryptophan of KatG

Jessica R. Krewall, Hui Xu, Olive J. Njuma, and Douglas C. Goodwin
Department of Chemistry and Biochemistry, Auburn University, Auburn, AL, 36849

Catalase-peroxidase (KatG) strongly resembles cytochrome c peroxidase (CcP), and contains many of the same active site residues, including a proximal tryptophan necessary CcP activity. These two enzymes, though both capable of peroxidase activity, exhibit three orders of magnitude difference in catalase activity1. This striking difference in function given similar protein structure begs investigation, particularly of the proximal tryptophan (W321 in KatG). When wtKatG is reacted with H₂O₂, a catalytically essential radical centered on the KatG-unique Met-Tyr-Trp covalent adduct gives way to an exchange-coupled signal attributed to W321 at the time H₂O₂ consumption ceases. This suggests that W321 is part of a route for off-pathway electron transfer and KatG catalase inactivation. Consistent with this, the W321F KatG variant revealed higher initial rates of activity, seeming to indicate that the proximal Trp detracts from KatG catalase turnover. However, on further investigation we have observed that the increased initial rate of W321F catalase activity coincides with more rapid enzyme inactivation. In addition, W321F catalase inactivation could not be recovered by the inclusion of a peroxidatic electron donor (PxED) to the same extent as wtKatG, indicating that electron transfer through a route other than W321 disables the enzyme from accessing mechanisms to recover catalase activity. Turnover-dependent inactivation in conjunction with a time-dependent decay of PxED-recoverable activity suggests that KatG utilizes a hole-hopping mechanism very similar to CcP². However, in KatG it serves to produce a synergistic peroxidase-based mechanism to maintain KatG’s unique and robust catalase activity.

Characterization of a Protein-Protein Interaction Involving the Ras-Related Protein Rheb and a Tuberous Sclerosis Complex 2 Variant

Nosaiba Shokr, Dr. Paul Adams
Cell and Molecular Biology program, Department of Chemistry and Biochemistry, University of Arkansas at Fayetteville

In order to understand the biological interactions, it is essential to relate the structures with the functions of any complex. Tumor suppressor complex 2, TSC2, plays a significant role in many pathways such as proteins synthesis, cell cycle, and signaling. Importantly, it acts as a GTPase activating partner for Ras homology enriched in brain, Rheb. It partially controls converting Rheb-GDP inactive form to Rheb-GTP active form through mammalian Target of Rapamycin, mTOR, pathway. Mutation in TSC2 causes abnormal activation of Rheb, but the molecular details of this interaction are still mysterious. We will study the effect of mutated TSC2 that may alter the catalytic activity of GTP hydrolysis or may weaken the protein complex formation. If the mutation in the adjacent binding domain to the GAP binding activity, it may stabilize the interaction of TSC2-stimulated GAP activity, which will support if there is any difference in the orientation of the mutation.

Effects Of 5-Hydroxymethylcytosine Epigenetic Modifications Within The Vegf Promoter Region On G-Quadraplex And I-Motif Dna Structure And Stability

Michael M. Molnar¹, R.K. Morgan², B. Summerford¹, Tracy A. Brooks², Randy M. Wadkins^1
1Dept. Chemistry Biochemistry, University of Mississippi, University, MS, United States
²Pharmacology, 315 Fraser Hall, University of Mississippi, University, MS, United States

Formaldimines are a class of structurally simple and highly reactive imine compounds, and it resembled the structure of formaldehyde. Synthetic application of formaldimine has been limited due to their instability and high reactivity that is difficult to control. Formaldimine is a useful chemical reagent with nitrogen hetero-atom. In addition to that formaldimine can be considered as a suitable precursor for synthesis of useful pharmaceuticals. In this study 1,2-ditosyl-1,2-diazetidine, a highly stable four-membered cyclic structure with two adjacent nitrogen atoms, has been identified as a potential precursor for N-tosyl formaldimine. The 1,2-diazetidine undergoes Fe(II)-catalyzed retro-[2+2] ring opening and slowly releases formaldimine to the reaction system. Comparing to the direct usage of the labile and highly reactive formaldimine, this Fe(II)-catalyzed process is able to cyclize unactivated olefins via [2+2+2] cyclization and produced piperidine derivatives in one step with up to 99 % yield.

Synthesis of vicinal dichlorides via established Triphosgene-Pyridine protocol

Alex Cleveland
Louisiana State University

Over 5000 halogenated natural products exist in nature and the need for the selective and mild construction of unactivated carbon-chlorine bonds is unfounded in the field of synthetic organic chemistry. Recently our group illustrated a mild and selective method utilizing a triphosgene-amine base mixture for the conversion of aliphatic alcohols or ketones to their corresponding alkyl chloride and vinyl chloride analogs, respectively.

In an effort to further expand and probe the limitation of this protocol, we began subjecting unactivated epoxides to our chlorination protocol. We discovered that vicinal dichlorides could be synthesized under mild conditions forming very clean products prior to chromatographic separation as well as being very compatible with many functionalities.

An appropriate scope of substrates will be presented as well as a mechanistic hypothesis. A proposed model to account for the stereochemistry of the transformation will also be discussed.

Analyzing Flavin-Based Electron Bifurcation in Heterodisulfide Reductase

Carly Engel
Auburn University

Daily human activities (i.e. rice production, husbandry of ruminants) has significantly increased the amount of methane, into the atmosphere (1, 2). Key enzymes in the methane synthesis pathway are studied as possible targets for novel inhibitors, including the enzyme heterodisulfide reductase (HDR; E.C. 1.8.98.1) (1). This enzyme reduces mixed disulfide of coenzyme M and coenzyme B (CoM-S-S-CoB) through flavin-based electron bifurcation, a process that creates electrons with low redox potentials without coupling the electron transfer step to ATP hydrolysis (2). Electrons are transported through numerous iron-sulfur clusters while flavin adenine dinucleotide (FAD) bifurcates these electrons into two different directions: to the heterodisulfide reduction site and 2) to ferredoxin. Due to the unusually high number of iron-sulfur clusters, electron paramagnetic resonance (EPR) studies are necessary to understand how FAD performs this bifurcation process. Redox titration and rapid freeze quench experiments were performed on HDR in Methanothermobacter marburgensis to determine the potentials of the iron-sulfur clusters and the time at which each cluster receives the electron from FAD. Midpoint potentials were determined for several of the paramagnetic species present in HDR and their position in the electron paths will be discussed.

Evaluation of extraction methods for use with NMR-based metabolomics in Yersinia enterocolitica and Y. pseudotuberculosis

Brandon R. Gines¹*, Willard E. Collier2, Mohammed A. Abdalla2, Teshome Yehualaeshet^1
1Department of Pathobiology, College of Veterinary Medicine, Tuskegee University, Tuskegee, AL 36088, USA
²Department of Chemistry, Tuskegee University, Tuskegee, AL 36088, USA

Sample preparation is one of the limiting factors in microbial metabolome analysis. Several methods exist for metabolite extraction of microbes, but the literature is contradictory regarding the sample model, adequacy and performance of each method. In this study, an optimal extraction protocol for Yersinae intracellular metabolites was investigated. Five extraction protocols consisting of different extraction solvent systems (60% methanol:water, pure methanol, acetonitrile:methanol:water [2:2:1], chloroform:methanol:water [2:1:1], and 60% ethanol:water) on the metabolic profile were compared. The number of detected peaks, sample-to-sample variation, and overall metabolite yield were taken into account as criteria. Extracted metabolites were analyzed by ¹H-NMR and partial least squares – discriminant analysis (PLS-DA). The extraction protocol using pure methanol as the extraction solvent provided the highest number of detected peaks for both Yersinia species analyzed. Together with the reproducibility and spectrum quality, pure methanol extraction was suitable for intracellular metabolite extraction from both species. However, depending on the metabolites of interest, other solvents might be more suitable as distinct profiles can be observed amongst the extraction methods.