Studying the Structure and Kinetics of Functional Nucleic Acids

Aptamers are valuable tools in a variety of biotechnological applications, ranging from biosensing to in vivo cell imaging. Studying these molecules can further our understanding of the structure and kinetics of nucleic acid-ligand binding. The sulforhodamine B binding aptamer (SRB-2) is a polyanionic molecule that was selected to bind a ligand with an overall negative charge. It is also a somewhat promiscuous aptamer that can bind ligands that vary markedly in shape, size and charge. These characteristics make its binding mechanisms of interest to characterize. Studying the binding kinetics of SRB-2 resulted in the identification of essential functional groups and led to the conclusion that a ligand that was not the selection target, rhodamine B, actually had the most ideal binding properties of those studied. It had a higher binding affinity than the original ligand, sulforhodamine B. In previous studies with SRB-2, several other ligands showed promise due to their high fluorescence enhancements, but we showed here that they were in fact binding to the aptamer non-specifically, questioning the usefulness of these ligands in potential applications. We used state of the art NMR methodologies to characterize the structure of SRB-2 bound to sulforhodamine B, but came short of elucidating a solution structure. We were, however, able to successfully calculate a structure for one of the stem-loop regions contained in the aptamer sequence. We also studied a DNAzyme containing a Na+ aptamer that clearly discriminates between Na+ and K+ without the use of a G-quadruplex, as would generally be expected. This existence of a selective Na+ aptamer is also encouraging with respect to biosensor development for metal ions. Distinguishing ions as similar in properties to Li+, Na+ and K+ is no small feat and indicates that DNAzymes and/or aptazymes may be a useful method for sensing unique metal ions

Modeling the replication of submicron-structured surfaces by micro injection molding

Abstract The replication of submicron surface structures by micro injection molding is a crucial factor in achieving advanced functionalities, such as antimicrobial resistance, in mass-produced plastic products. In this work, we investigate and model the replication quality of laser-induced periodic surface structures by micro injection molding of different bio-based polymers. A comprehensive multiscale model was developed to predict the submicron scale polymer flow, using a numerical model to analyze the polymer behavior in the mold macro cavity and determine the boundary conditions for the filling of the surface structures. The replication of the mold topography was modeled considering topographical parameters, polymer rheology and thermal behavior, and the mold surface energy, which was modified by depositing an atomic layer of alumina on the steel surface structures. The modeling approach was validated against injection molding experiments, in which the mold temperature was varied due to its well-known influence on replication. The sensitivity to polymer selection, mold surface properties, and mold temperature, was captured. A maximum error of 8% showed the accuracy of the multi-scale model

Ligand Specificity and Affinity in the Sulforhodamine B Binding RNA Aptamer

The final publication is available at Elsevier via http://dx.doi.org/10.1016/j.bbrc.2020.06.056. © 2020. This manuscript version is made available under the CC-BY-NC-ND 4.0 license http://creativecommons.org/licenses/by-nc-nd/4.0/Binding affinity and selectivity are critical properties of aptamers that must be optimized for any application. The sulforhodamine B binding RNA aptamer (SRB‐2) is a somewhat promiscuous aptamer that can bind ligands that vary markedly in shape, size and charge. Here we categorize potential ligands based on their binding mode and structural characteristics required for high affinity and selectivity. Several known and potential ligands of SRB‐2 were screened for binding affinity using LSPR, ITC and NMR spectroscopy. The study shows that rhodamine B has the ideal structural and electrostatic properties for selective and high‐affinity binding of the SRB‐2 aptamer.NSERC Discovery Grant 30345

Thioflavin T fluorescence and NMR spectroscopy 1 suggesting a non-G-quadruplex structure for a 2 sodium binding aptamer embedded in DNAzymes

Recently, a Na+-binding aptamer was reported to be embedded in a few RNA-cleaving DNAzymes including NaA43, Ce13d and NaH1. These DNAzymes require Na+ for activity but show no activity in the presence of K+ or other metal ions. Given that DNA can selectively bind K+ by forming a G-quadruplex structure, this work aims to answer whether this Na+ aptamer also uses a G-quadruplex to bind Na+. The Na+ aptamer embedded in Ce13d consists of multiple GG sequences, which is also a pre-requisite for the formation of G4 structures. To delineate the structural differences and similarities between Ce13d and G-quadruplex in terms of metal binding, thioflavin T (ThT) fluorescence spectroscopy, NMR spectroscopy and CD spectroscopy were used. Through comparative ThT fluorescence spectrometry studies, we deciphered that while a control G-quadruplex DNA exhibited notable fluorescence enhancement up to 5 mM K+ with a Kd of 0.52 mM, the Ce13d DNAzyme fluorescence was negligibly perturbed with similar concentrations of K+. Opposed to this, Ce13d displayed specific remarkable fluorescence decrease with low millimolar concentrations of Na+. NMR experiments at two different pH values suggest that Ce13d adopts a significantly different conformation or equilibrium of conformations in the presence of Na+ versus K+ and has a more stable structure in the presence of Na+. Additionally, absence of characteristic peaks expected for a G-quadruplex structure in 1D 1H NMR suggest that G4 is not responsible for the Na+ binding. This theory is confirmed by absence of characteristic peaks in the CD spectra of this sequence. Therefore, we concluded that the aptamer must be selective for Na+ and binds using a structural element that does not contain G4.Natural Sciences and Engineering Research Council of Canada, Discovery Grant 303454

Measurement of the top quark forward-backward production asymmetry and the anomalous chromoelectric and chromomagnetic moments in pp collisions at √s = 13 TeV

Abstract The parton-level top quark (t) forward-backward asymmetry and the anomalous chromoelectric (d̂ t) and chromomagnetic (μ̂ t) moments have been measured using LHC pp collisions at a center-of-mass energy of 13 TeV, collected in the CMS detector in a data sample corresponding to an integrated luminosity of 35.9 fb−1. The linearized variable AFB(1) is used to approximate the asymmetry. Candidate t t ¯ events decaying to a muon or electron and jets in final states with low and high Lorentz boosts are selected and reconstructed using a fit of the kinematic distributions of the decay products to those expected for t t ¯ final states. The values found for the parameters are AFB(1)=0.048−0.087+0.095(stat)−0.029+0.020(syst),μ̂t=−0.024−0.009+0.013(stat)−0.011+0.016(syst), and a limit is placed on the magnitude of | d̂ t| < 0.03 at 95% confidence level. [Figure not available: see fulltext.

Measurement of t(t)over-bar normalised multi-differential cross sections in pp collisions at root s=13 TeV, and simultaneous determination of the strong coupling strength, top quark pole mass, and parton distribution functions

Peer reviewe

An embedding technique to determine ττ backgrounds in proton-proton collision data

An embedding technique is presented to estimate standard model tau tau backgrounds from data with minimal simulation input. In the data, the muons are removed from reconstructed mu mu events and replaced with simulated tau leptons with the same kinematic properties. In this way, a set of hybrid events is obtained that does not rely on simulation except for the decay of the tau leptons. The challenges in describing the underlying event or the production of associated jets in the simulation are avoided. The technique described in this paper was developed for CMS. Its validation and the inherent uncertainties are also discussed. The demonstration of the performance of the technique is based on a sample of proton-proton collisions collected by CMS in 2017 at root s = 13 TeV corresponding to an integrated luminosity of 41.5 fb(-1).Peer reviewe

Measurement of the Splitting Function in &ITpp &ITand Pb-Pb Collisions at root&ITsNN&IT=5.02 TeV

Data from heavy ion collisions suggest that the evolution of a parton shower is modified by interactions with the color charges in the dense partonic medium created in these collisions, but it is not known where in the shower evolution the modifications occur. The momentum ratio of the two leading partons, resolved as subjets, provides information about the parton shower evolution. This substructure observable, known as the splitting function, reflects the process of a parton splitting into two other partons and has been measured for jets with transverse momentum between 140 and 500 GeV, in pp and PbPb collisions at a center-of-mass energy of 5.02 TeV per nucleon pair. In central PbPb collisions, the splitting function indicates a more unbalanced momentum ratio, compared to peripheral PbPb and pp collisions.. The measurements are compared to various predictions from event generators and analytical calculations.Peer reviewe

MUSiC : a model-unspecific search for new physics in proton-proton collisions at root s=13TeV

Results of the Model Unspecific Search in CMS (MUSiC), using proton-proton collision data recorded at the LHC at a centre-of-mass energy of 13 TeV, corresponding to an integrated luminosity of 35.9 fb(-1), are presented. The MUSiC analysis searches for anomalies that could be signatures of physics beyond the standard model. The analysis is based on the comparison of observed data with the standard model prediction, as determined from simulation, in several hundred final states and multiple kinematic distributions. Events containing at least one electron or muon are classified based on their final state topology, and an automated search algorithm surveys the observed data for deviations from the prediction. The sensitivity of the search is validated using multiple methods. No significant deviations from the predictions have been observed. For a wide range of final state topologies, agreement is found between the data and the standard model simulation. This analysis complements dedicated search analyses by significantly expanding the range of final states covered using a model independent approach with the largest data set to date to probe phase space regions beyond the reach of previous general searches.Peer reviewe

Search for new particles in events with energetic jets and large missing transverse momentum in proton-proton collisions at root s=13 TeV

A search is presented for new particles produced at the LHC in proton-proton collisions at root s = 13 TeV, using events with energetic jets and large missing transverse momentum. The analysis is based on a data sample corresponding to an integrated luminosity of 101 fb(-1), collected in 2017-2018 with the CMS detector. Machine learning techniques are used to define separate categories for events with narrow jets from initial-state radiation and events with large-radius jets consistent with a hadronic decay of a W or Z boson. A statistical combination is made with an earlier search based on a data sample of 36 fb(-1), collected in 2016. No significant excess of events is observed with respect to the standard model background expectation determined from control samples in data. The results are interpreted in terms of limits on the branching fraction of an invisible decay of the Higgs boson, as well as constraints on simplified models of dark matter, on first-generation scalar leptoquarks decaying to quarks and neutrinos, and on models with large extra dimensions. Several of the new limits, specifically for spin-1 dark matter mediators, pseudoscalar mediators, colored mediators, and leptoquarks, are the most restrictive to date.Peer reviewe