Vinyldeoxyadenosine in a Sarcin−Ricin RNA Loop and Its Binding to Ricin Toxin A-Chain

Roday, Setu; Saen‐oon, Suwipa; Schramm, Vern L.

doi:10.1021/bi0621821

Cited by 5 publications

(4 citation statements)

References 45 publications

(72 reference statements)

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“…Inhibitors were designed and synthesized to mimic the stem–loop substrate. Alterations toward the transition state included replacing the substrate adenylate with a hydroxypyrrolidine mimic of the ribocation in an N1′-methylene bridge to 9-deazaadenine placed at the depurination site in tetraloops of varied length (Figures and ). , An abasic, 14mer construct inhibited ricin A-chain with a K d of 480 nM (1N-14, Figure ). Filling the catalytic site with adenine improved binding of IN-14 to 12 nM, demonstrating the benefit of the adenine in combination with the transition state ribocation mimic .…”

Section: Ribosome-inactivating Proteinsmentioning

confidence: 99%

Enzymatic Transition States and Drug Design

Schramm

2018

Chem. Rev.

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Transition state theory teaches that chemically stable mimics of enzymatic transition states will bind tightly to their cognate enzymes. Kinetic isotope effects combined with computational quantum chemistry provides enzymatic transition state information with sufficient fidelity to design transition state analogues. Examples are selected from various stages of drug development to demonstrate the application of transition state theory, inhibitor design, physicochemical characterization of transition state analogues, and their progress in drug development.

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Section: Ribosome-inactivating Proteinsmentioning

confidence: 99%

Enzymatic Transition States and Drug Design

Schramm

2018

Chem. Rev.

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“…19 These characteristics have been exploited to study changes in RNA structure and binding in ricin toxin protein. 21 When 8VA is base-stacked in DNA its fluorescence is quenched significantly. 18 This kind of behavior is observed in other fluorescent adenine analogues like 2AP [22][23][24][25] and the pteridines 6MAP and DMAP.…”

Section: Introductionmentioning

confidence: 99%

“…Recent work by Kenfack et al showed that 8VA is better stabilized by base stacking than 2-aminopurine (2AP) in dsDNA, and thus they suggest that 8VA is a better probe than 2AP . These characteristics have been exploited to study changes in RNA structure and binding in ricin toxin protein …”

Section: Introductionmentioning

confidence: 99%

Change in Electronic Structure upon Optical Excitation of 8-Vinyladenosine: An Experimental and Theoretical Study

et al. 2009

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8-Vinyladenosine (8VA) is an adenosine analog, like 2-aminopurine (2AP), that has a red-shifted absorption and high fluorescence quantum yield. When introduced into double-stranded DNA (dsDNA), its base-pairing and base-stacking properties are similar to those of adenine. Of particular interest, the fluorescence quantum yield of 8VA is sensitive to base stacking, making it a very useful real-time probe of DNA structure. The fundamental photophysics underlying this fluorescence quenching by base stacking is not well understood, and thus exploring the excited state electronic structure of the analog is warranted. In this study, we report on changes in the electronic structure of 8VA upon optical excitation. Stark spectroscopy was performed on 8VA monomer in frozen ethanol glass at 77 K to obtain the direction and degree of charge redistribution in the form of the difference dipole moment, Deltamu(01) = 4.7 +/- 0.3 D, and difference static polarizability, tr(Delta(alpha)01) = 21 +/- 11 A(3), for the S(0)-->S(1) transition. In addition, solvatochromism experiments were performed on 8VA in various solvents and analyzed using Bakhshiev's model. High level ab initio methods were employed to calculate transition energies, oscillator strengths, and dipole moments of the ground and excited states of 8VA. The direction of Deltamu(01) was assigned in the molecular frame for the lowest optically accessible state. Our study shows that the angle between ground and excited state dipole moment plays a critical role in understanding the change in electronic structure upon optical excitation. Compared to 2AP, 8VA has a larger difference dipole moment which, with twice the extinction coefficient, suggests that 8VA is superior as a two-photon probe for microscopy studies. To this end, we have measured the ratio of the two-photon fluorescence yields of the two analogs by excitation at the respective monomer absorption maxima. We show that 8VA is indeed a significantly brighter two-photon fluorophore, based on our experimental and computational results.

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“…Early studies identified substrate analogs that target the active site and inhibit RTA with modest inhibitory activity. , However, these compounds had poor solubility, and none of them exhibited potent inhibitory activity against ricin due to the large and highly polar active site . Transition state analogs of the RTA catalytic reaction were powerful inhibitors of the depurination of small stem-loop RNA substrates at low pH but did not protect eukaryotic ribosomes at physiological pH. ,− High-throughput screening (HTS) of compounds against protein synthesis inhibitory activity or cytotoxicity yielded compounds that target toxin trafficking or cell death pathways but not the toxin itself. − …”

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confidence: 99%

Small Molecule Inhibitors Targeting the Interaction of Ricin Toxin A Subunit with Ribosomes

Harijan

Kahn

et al. 2020

ACS Infect. Dis.

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Ricin toxin A subunit (RTA) removes an adenine from the universally conserved sarcin/ricin loop (SRL) on eukaryotic ribosomes, thereby inhibiting protein synthesis. No high affinity and selective small molecule therapeutic antidotes have been reported against ricin toxicity. RTA binds to the ribosomal P stalk to access the SRL. The interaction anchors RTA to the P protein C-termini at a well-defined hydrophobic pocket, which is on the opposite face relative to the active site. The RTA ribosome binding site has not been previously targeted by small molecule inhibitors. We used fragment screening with surface plasmon resonance to identify small molecular weight lead compounds that bind RTA and defined their interactions by crystallography. We identified five fragments, which bound RTA with mid-micromolar affinity. Three chemically distinct binding fragments were cocrystallized with RTA, and crystal structures were solved. Two fragments bound at the P stalk binding site, and the third bound to helix D, a motif distinct from the P stalk binding site. All fragments bound RTA remote from the catalytic site and caused little change in catalytic site geometry. Two fragments uniquely bound at the hydrophobic pocket with affinity sufficient to inhibit the catalytic activity on eukaryotic ribosomes in the low micromolar range. The binding mode of these inhibitors mimicked the interaction of the P stalk peptide, establishing that small molecule inhibitors can inhibit RTA binding to the ribosome with the potential for therapeutic intervention.

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Vinyldeoxyadenosine in a Sarcin−Ricin RNA Loop and Its Binding to Ricin Toxin A-Chain

Cited by 5 publications

References 45 publications

Enzymatic Transition States and Drug Design

Enzymatic Transition States and Drug Design

Change in Electronic Structure upon Optical Excitation of 8-Vinyladenosine: An Experimental and Theoretical Study

Small Molecule Inhibitors Targeting the Interaction of Ricin Toxin A Subunit with Ribosomes

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