Multi-Step Reactions: The Methods for Analytical Solving the Direct Problem

Korobov, Viktor I.; Ochkov, Valery

doi:10.1007/978-3-7091-0531-3_2

Cited by 6 publications

(3 citation statements)

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“…The rate of pNPhOH formation in the three sequential irreversible reactions depicted in Scheme 3 is expressed by eq 7. The differential equation was derived by reported approaches 18 (see SI) and simulated at different pH values using MS Excel. Here, the F terms represent amplitude coefficients for each of three exponentials representing each rate constant (see SI).…”

Section: ■ Resultsmentioning

confidence: 99%

β-Eliminative Releasable Linkers Adapted for Bioconjugation of Macromolecules to Phenols

et al. 2013

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We recently reported a chemical approach for half-life extension that utilizes sets of releasable linkers to attach drugs to macromolecules via a cleavable carbamate group (Santi et al., Proc. Nat. Acad. Sci. U.S.A. 2012, 109, 6211-6216). The linkers undergo a β-elimination cleavage to release the free, native amine-containing drug. A limitation of the technology is the requirement for an amino group on the drug in order to form the carbamate bond, since most small molecules do not have an amine functional group. Here, we describe an approach to adapt these same β-elimination carbamate linkers so they can be used to connect other acidic heteroatoms, in particular, phenolic hydroxyl groups. The approach utilizes a methylene adaptor to connect the drug to the carbamate nitrogen, and an electron-withdrawing group attached to carbamate nitrogen to stabilize the system against a pH-independent spontaneous cleavage. Carbamate cleavage is driven by β-elimination to give a carboxylated aryl amino Mannich base which rapidly collapses to give the free drug, an aryl amine, and formaldehyde.

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Section: ■ Resultsmentioning

confidence: 99%

β-Eliminative Releasable Linkers Adapted for Bioconjugation of Macromolecules to Phenols

et al. 2013

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“…Relaxation kinetics of binding were collected and plotted as the FRET efficiency versus time. The resulting transients were fit to the integrated rate law for the bimolecular reaction (assuming equimolar total protein and RNA concentrations): with the solution: where [ P ] T is the total protein concentration used for the measurement, [ PR ] t is the complex concentration at time t , k on is the association rate coefficient, and k off is the dissociation rate coefficient. The observed signal is proportional to the change in [PR] t : where K d is the equilibrium dissociation constant for the binding reaction.…”

Section: Methodsmentioning

confidence: 99%

Cellular Sticking Can Strongly Reduce Complex Binding by Speeding Dissociation

Davis

Gruebele

2021

J. Phys. Chem. B

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While extensive studies have been carried out to determine protein-RNA binding affinities, mechanisms, and dynamics in vitro, such studies do not take into consideration the effect of the many weak nonspecific interactions in a cell filled with potential binding partners. Here we experimentally tested the role of the cellular environment on affinity and binding dynamics between a protein and RNA in living U-2 OS cells. Our model system is the spliceosomal protein U1A and its binding partner SL2 of the U1 snRNA. The binding equilibrium was perturbed by a laser-induced temperature jump and monitored by Förster resonance energy transfer. The apparent binding affinity in live cells was reduced by up to 2 orders of magnitude compared to in vitro. The measured in-cell dissociation rate coefficients were up to 2 orders of magnitude larger, whereas no change in the measured association rate coefficient was observed. The latter is not what would be anticipated due to macromolecular crowding or nonspecific sticking of the uncomplexed U1A and SL2 in the cell. A quantitative model fits our experimental results, with the major cellular effect being that U1A and SL2 sticking to cellular components are capable of binding, just not as strongly as the free complex. This observation suggests that high binding affinities measured or designed in vitro are necessary for proper binding in vivo, where competition with many nonspecific interactions exists, especially for strongly interacting species with high charge or large hydrophobic surface areas.

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“…For all chemical reactions, pre-exponential factors in the transition state theory equation k on = ν exp[(−Δ G ⧧ )/( k B T )], which can be thought of as vibrational frequencies in the reactant state, were calculated from the MD trajectories as an inverse of the autocorrelation times calculated for the reacting partners’ distance. , From the obtained free energy profiles and autocorrelation MD data, rate constants were calculated and used to solve a set of coupled ordinary differential equations describing the kinetics of the studied reactions (see Figure S9 and eq S1). Rate constants for the reverse reactions were calculated on the basis of equilibrium constants, K = exp[(−Δ G °)/( k B T )], as k off = k on / K .…”

Section: Methodsmentioning

confidence: 99%

Dominant Pathways of Adenosyl Radical-Induced DNA Damage Revealed by QM/MM Metadynamics

Wityk

Wieczór

Makurat

et al. 2017

J. Chem. Theory Comput.

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Brominated nucleobases sensitize double stranded DNA to hydrated electrons, one of the dominant genotoxic species produced in hypoxic cancer cells during radiotherapy. Such radiosensitizers can therefore be administered locally to enhance treatment efficiency within the solid tumor while protecting the neighboring tissue. When a solvated electron attaches to 8-bromoadenosine, a potential sensitizer, the dissociation of bromide leads to a reactive C8 adenosyl radical known to generate a range of DNA lesions. In the current work, we propose a multiscale computational approach to elucidate the mechanism by which this unstable radical causes further damage in genomic DNA. We employed a combination of classical molecular dynamics conformational sampling and QM/MM metadynamics to study the thermodynamics and kinetics of plausible reaction pathways in a realistic model, bridging between different time scales of the key processes and accounting for the spatial constraints in DNA. The obtained data allowed us to build a kinetic model that correctly predicts the products predominantly observed in experimental settings-cyclopurine and β-elimination (single strand break) lesions-with their ratio and yield dependent on the effective lifetime of the radical species. To date, our study provides the most complete description of purine radical reactivity in double stranded DNA, explaining the radiosensitizing action of electrophilic purines in molecular detail as well as providing a conceptual framework for the computational modeling of competing reaction pathways in biomolecules.

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Multi-Step Reactions: The Methods for Analytical Solving the Direct Problem

Cited by 6 publications

References 0 publications

β-Eliminative Releasable Linkers Adapted for Bioconjugation of Macromolecules to Phenols

β-Eliminative Releasable Linkers Adapted for Bioconjugation of Macromolecules to Phenols

Cellular Sticking Can Strongly Reduce Complex Binding by Speeding Dissociation

Dominant Pathways of Adenosyl Radical-Induced DNA Damage Revealed by QM/MM Metadynamics

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