Wendi David | Texas State University (original) (raw)

Papers by Wendi David

The primary goal of the funded research involved using surface plasmon resonance (SPR) for determ... more The primary goal of the funded research involved using surface plasmon resonance (SPR) for determining real-time interactions between viral SV40 large T-antigen (T-ag), DNA, and a stream of quadruplex DNA-interactive small molecules. We successfully developed an SPR-based assay for monitoring the ability of T-ag to unwind both normal duplex DNA and quadruplex DNA and characterized the ability of several small molecules to inhibit the helicase activity of T-ag. The ability to monitor unwinding of quadruplex DNA in real-time is an advance for investigating the inhibitory effects of quadruplex-interactive agents and this work was published recently in Cell Biochemistry and Biophysics (Feb. 2009). Our results will aid in future characterization of potential therapeutic approaches involving G-quadruplex interactive small molecules and in understanding the structural basis for selective targeting of different families of G-quadruplex DNA helicases. These results form the basis for a recen...

Biochemistry, 2002

The (Z)-hex-1,5-diyne-3-ene reactive core common to the enediyne antitumor antibiotics undergoes ... more The (Z)-hex-1,5-diyne-3-ene reactive core common to the enediyne antitumor antibiotics undergoes a Bergman cyclization after proper activation to afford reactive diradical intermediates that are responsible for initiating DNA cleavage. Direct modification of the enediyne core has been proposed as a method to permit cancer cell-specific triggering of the diradical-generating cyclization. For example, 3-aza-3-ene-1,5-diynes undergo an aza-Bergman cyclization to afford the fleeting 2,5-didehydropyridine diradicals. While protonation of these aza-enediynes can afford products of diradical trapping, the hydrolytic instability of the 3-aza-3-ene-1,5-diyne moiety prevents its use in pH-triggered DNA cleaving anticancer agents. Recently, more hydrolytically stable systems incorporating the 4-aza-3-ene-1,6-diyne moiety were developed. We report here studies of the 4-aza-3-ene-1,6-diyne-containing benzimidazolium salt AZB002 [1-methyl-2-(phenylethynyl)-3-(3-phenylprop-2-ynyl)-3H-benzimidazolium tetrafluoroborate] and two structurally related heterocycles that lack the aza-enediyne functionality, AZB016 [1,3-dimethyl-2-(phenylethynyl)-3H-benzimidazolium triflate] and AZB004 [3-methyl-2-(phenylethynyl)benzothiazolium triflate]. The interaction of these compounds with supercoiled DNA, a double-stranded DNA fragment, and a short DNA duplex oligonucleotide was investigated. There are three distinct DNA interactions exhibited by AZB002: a frank strand scission leading to the relaxation of supercoiled DNA and formation of at least two different DNA adducts, one of which leads to cytosine-specific cleavage after piperidine/heat treatment. In contrast, analogues lacking the aza-enediyne functionality either fail to interact with DNA (AZB016) or cleave DNA at guanine residues, presumably through alkylation of the N-7 position (AZB004). We also investigated the cytotoxicity of AZB002 and the related heterocyclic compounds AZB004 and AZB016 and find that only the DNA interactive compounds AZB002 and AZB004 display significant cytotoxicity. In particular, AZB002 is cytotoxic against a wide range of cancer cell lines.

Comp Biochem Physiol Pt C, 2004

Cloning of the Xiphophorus maculatus Polbeta gene and overexpression of the recombinant Polbeta p... more Cloning of the Xiphophorus maculatus Polbeta gene and overexpression of the recombinant Polbeta protein has been performed. The organization of the XiphPolbeta introns and exons, including intron-exon boundaries, have been assigned and were found to be similar to that for human Polbeta with identical exon sizes except for exon XII coding for an additional two amino acid residues in Xiphophorus. The cDNA sequence encoding the 337-amino acid X. maculatus DNA polymerase beta (Polbeta) protein was subcloned into the Escherichia coli expression plasmid pET. Induction of transformed E. coli cells resulted in the high-level expression of soluble recombinant Polbeta, which catalyzed DNA synthesis on template-primer substrates. The steady-state Michaelis constants (Km) and catalytic efficiencies (kcat/Km) of the recombinant XiphPolbeta for nucleotide insertion opposite single-nucleotide gap DNA substrates were measured and compared with previously published values for recombinant human Polbeta. Steady-state in vitro Km and kcat/Km values for correct nucleotide insertion by XiphPolbeta and human Polbeta were similar, although the recombinant Xiphophorus protein exhibited 2.5-7-fold higher catalytic efficiencies for dGTP and dCTP insertion versus human Polbeta. In contrast, the recombinant XiphPolbeta displayed significantly lower fidelities than human Polbeta for dNTP insertion opposite a single-nucleotide gap at 37 degrees C.

Analytical Chemistry, 2002

Selectivity, binding stoichiometry, and mode of binding of Tel01, distamycin A, and diethylthioca... more Selectivity, binding stoichiometry, and mode of binding of Tel01, distamycin A, and diethylthiocarbocyanine iodide (DTC) to the parallel stranded G4-quadruplex [d(T 2 G 5 T)] 4 were investigated by ESI-MS. The first drug/ quadruplex complexes observed by ESI-MS are described. Tel01, distamycin A, and DTC all form complexes with quadruplex DNA, but only Tel01 is completely selective for quadruplex versus duplex oligonucleotide under the conditions employed. Previous solution determinations of the binding mode of Tel01 and distamycin A to quadruplex oligonucleotides indicate that Tel01 interacts through end-stacking with guanine tetrads of quadruplex DNA, while distamycin A interacts by binding to quadruplex grooves. When these two different drug/quadruplex complexes are subjected to collisionally activated dissociation in a mass spectrometer, the observed fragmentation patterns are distinct. Tel01/quadruplex complexes undergo facile loss of drug and dissociation to single-strand oligonucleotide ions, while distamycin/quadruplex complexes fragment into single-strand oligonucleotide ions in which the drug molecule is retained. Dissociation patterns for DTC/quadruplex complexes are similar to those of distamycin; therefore, it is concluded that DTC interacts with [d(T 2 G 5 T)] 4 through groove-binding. These ESI-MS results are applicable to both the identification and characterization of G-quadruplex interactive agents and may also be useful in probing unusual DNA structures.

Org Lett, 2002

[reaction: see text] Simple, acyclic 3-aza-3-ene-1,5-diynes undergo an aza-Bergman rearrangement ... more [reaction: see text] Simple, acyclic 3-aza-3-ene-1,5-diynes undergo an aza-Bergman rearrangement to a fleeting 2,5-didehydropyridine (2,5-ddp) intermediate that rapidly ring-opens to beta-alkynylacrylonitrile products. In an effort to access longer-lived 2,5-ddp intermediates, we have prepared heterocyclic 3-aza-3-ene-1,5-diynes. The thermolysis of one such heterocyclic aza-enediyne does not afford products derived from trapping a 2,5-ddp intermediate but rather cyclopropanes that appear to arise from a carbene intermediate and a product that appears to be a trapping product from a 2,3-ddp intermediate.

Bioorganic Medicinal Chemistry Letters, Nov 19, 2001

The 2-alkynylbenzothiazolium salts 3a-d incorporating an N-propargyl moiety have been prepared as... more The 2-alkynylbenzothiazolium salts 3a-d incorporating an N-propargyl moiety have been prepared as aza-enediyne analogues. While these aza-enediynes are shown to be modest DNA cleavage agents, DNA cleavage was also observed with the Nmethyl-2-alkynylbenzothiazolium salt 4, which lacks the aza-enediyne moiety. The structural requirements for DNA cleavage, and the correlation of DNA cleavage efficiency with the propensity of these compounds to undergo nucleophilic addition by methanol support a proposed DNA cleavage mechanism involving DNA alkylation by appropriate 2-alkynyl-substituted benzothiazolium salts. #

The Faseb Journal, Mar 1, 2008

The Faseb Journal, Apr 1, 2014

Journal of the American Society For Mass Spectrometry, Apr 30, 2003

Electrospray ionization mass spectrometry (ESI-MS) is increasingly used to probe the nature of no... more Electrospray ionization mass spectrometry (ESI-MS) is increasingly used to probe the nature of noncovalent complexes; however, assessing the relevance of gas-phase results to structures of complexes in solution requires knowledge of the types of interactions that are maintained in a solventless environment and how these might compare to key interactions in solution. This study addresses the factors impacting the strength of hydrogen bonding noncovalent interactions in the gas phase. Hydrogen bonded complexes consisting of ammonium ions bound to polydentate ethers are transported to the gas phase with ESI, and energy-variable collisional activated dissociation (CAD) is used to map the relative dissociation energies. The measured relative dissociation energies are correlated with the gas-phase basicities and steric factors of the amine and polyether constituents. To develop correlations between hydrogen bonding strength and structural features of the donor and acceptor molecules, a variety of amines with different gas-phase basicities and structures were selected, including primary, secondary, and tertiary amines, as well as those that are bidentate to promote intramolecular hydrogen bonding. The acceptor molecules are polydentate ethers, such as 18-crown-6. Four primary factors influence the observed dissociation energies of the polyether/ammonium ion complexes: the gas-phase basicities of the polyether and amine, steric effects of the amines, conformational flexibility of the polyethers, and the inhibition of intramolecular hydrogen bonds of the guest ammonium ions in the resulting ammonium/polyether noncovalent complexes. (J Am Soc Mass Spectrom 2003, 14, 383-392) E lectrospray ionization mass spectrometry (ESI-MS) is being increasingly used for the analysis of noncovalent complexes, especially those involving biomolecules, because it is a "soft" method for the transfer of solution species into the gas phase . A key question revolves around the nature of the noncovalent complexes once they enter the gas phase. The specificity and the type of interactions that are retained in a solventless environment are important considerations when assessing the relevance of gas-phase results to the structures of solution-phase complexes . Information on the relative stabilities of noncovalent complexes is often obtained from their gas-phase dissociation behavior , and many recent examples of biologically interesting macromolecular complexes demonstrate the importance of electrostatic interactions and hydrogen bonds in maintaining these noncovalent associations in the gas phase . In some cases, the stabilities of complexes obtained based on gas-phase measurements do not correlate well with those obtained in solution , whereas other reports have shown remarkable agreement between gas-phase and solution results .

The Faseb Journal, Apr 1, 2009

Http Dx Doi Org 10 1080 10610270412331295995, Aug 6, 2006

Self-assembly processes based on shape complementarity and noncovalent binding interactions are w... more Self-assembly processes based on shape complementarity and noncovalent binding interactions are widely recognized as a fundamental principle in nature. Besides charge pairing and hydrogen bonding, hydrophobic interactions play a crucial role in water. Here we report the self-assembly of structurally simple monomers to yield defined dimeric and trimeric aggregates in highly polar media, based on ionic and solvophobic interactions. NMR, mass spectrometry and curve fitting were used to characterize these supramolecular assemblies in water-methanol solutions.

ChemInform, 1997

ABSTRACT ChemInform is a weekly Abstracting Service, delivering concise information at a glance t... more ABSTRACT ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Due to the increasing number of helicases shown to unwind quadruplex DNA structures in addition t... more Due to the increasing number of helicases shown to unwind quadruplex DNA structures in addition to duplex DNA, the biological significance of this activity is currently under investigation. One limitation of traditional gel analysis of helicase activity is the inability to effectively monitor unwinding of intramolecular G-quadruplex DNA substrates. Optimization of our novel SPR-based assay for monitoring the helicase activity of simian virus 40 (SV40) large T-antigen (T-ag) was undertaken to explore limitations and improvements in the ability to investigate G-quadruplex helicase activity. Although T-ag helicase was used, the assay is general in nature. An improved method for assessing unwinding of intramolecular G-quadruplex DNA substrates was developed.

ACS Symposium Series, 2011

The ability of G-quadruplex interactive small molecules to inhibit SV40 Large T-Antigen G-quadrup... more The ability of G-quadruplex interactive small molecules to inhibit SV40 Large T-Antigen G-quadruplex helicase activity was determined using a surface plasmon resonance based assay. Unwinding of a physiologically relevant intramolecular G-quadruplex DNA substrate was completely inhibited by one novel porphyrin derivative under these conditions. This SPR-based method represents an important strategy for comparing helicase inhibition effectiveness.

ChemInform, 2002