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Antibiotic resistance is a global threat beside the ongoing pandemic by SARS-CoV-2.The number of deaths due to antibiotic-resistant infections is increasing at an alarmingrate. The COVID-19 pandemic has already claimed millions of deaths worldwide.Fighting against antibiotic-resistant superbugs and the SARS-CoV-2 has become achallenge. A significant amount of research is going on to develop the vaccine and smallmolecule antiviral and antibacterial therapeutics targeting proteins. Fortunately, novelnon-coding regulatory RNA targets have been identified for developing new antibacterialand antiviral drugs such as bacterial T-box riboswitch, RNA thermometers, and viralstem-loop II motif. T-box riboswitch can control the transcription or translation of aminoacid-related genes in bacteria by forming unique interactions between tRNA and mRNA.RNA thermometers (RNATs) are temperature-responsive riboswitches that control thetranslation based on temperature sensing thus controlling the interaction with the mRNAand 16S rRNA. In Shigella dysenteriae, three RNATs, i.e., ompA, shuT, and shuA, havebeen discovered. ompA RNAT controls the translation of outer membrane protein A.shuT, and shuA RNAT controls the translation of two proteins that are crucial to thebacterial heme utilization system. The Stem-loop II motif (S2M) is a highly conservedRNA element found in most coronaviruses, astroviruses, and picornaviruses that plays apotential role in viral replication and invasion. The RNA structure plays a significant rolein its regulatory function for all of these potential therapeutic targets. Consequently, it isessential to examine the factors that affect the RNA structure and RNA-RNA interaction.Despite having limited building blocks, RNA has diverse functions in the cells. Baseprotonation and protonated base pairs often occur in RNA when interacting with otherbiomolecules, thus could play a critical role in vital biological processes. Differentbiophysical assays, including UV use in monitoring thermal denaturation, FRET,fluorescence polarization, chemical and enzymatic probing, EMSA, and In vitrotranscription assay were used to probe the RNA structures, stability, conformation, andpotential function. These assays were also used to investigate the effect of potentialligands targeting these non-coding regulatory RNAs. Our study showed that pH affectsthe stability and conformations, including antiterminator model RNAs, RNATs, stemloopII motif, and the RNA-RNA interaction between the tRNA and antiterminator andRNAT and 16S rRNA mimic. Using the drug discovery screening assays, we identifiedseveral quinoline compounds that affected the riboswitch function and alteredconformation of the stem-loop II motif of SARS-CoV-2 and antiterminator element of Tboxriboswitch. Several 4,5-disubstituted oxazolidinones were also identified as weakinhibitors of T-box riboswitch function.

Abstract: This thesis studies the intrinsic disorder effects and the electro- and magnetotransport properties of some perovskite manganites. Recently reported colossal electro-resistance (ER) (a relative change in resistance with electric field /current ) in manganites, analogous to their respective colossal magnetoresistance (CMR) behavior, demonstrates another way of controlling the transport properties of manganites directly by using the electric current. In the last few years, there is considerable debate regarding the origin of this ER. In order to better understand this effect in some manganites, a technique for precisely measuring the intrinsic ER of a material has been developed and the technique has been utilized to measure the ER of polycrystalline Sm0.60Sr0.40MnO3 and Sm0.55Sr0.45MnO3 manganites. In this technique, the contribution of Joule heating to the ER has been successfully eliminated. It is found that the changes of the intrinsic ER with current density are much more significant in Sm0.60Sr0.40MnO3 compared to Sm0.55Sr0.45MnO3. We have also tuned the existing disorders/inhomogeneity in Sm0.60Sr0.40MnO3 (SSMO) by Fe doping in Mn site to probe the possible mechanisms responsible for ER. It is found that ER increases with Fe doping and it is strongly suppressed by an applied dc magnetic field. A discussion of the mechanism behind these phenomena is presented. We have investigated the time dependence of the electrical resistivity in the Sm0.60Sr0.40Mn1-xFexO3, (1-y) Sm0.60Sr0.40 MnO3 +yBaTiO3 and (Sm0.5Gd0.5)0.55 Sr0.45Mn1-zRuzO3 manganites by doping with Fe, BaTiO3 and Ru to understand better the origin of relaxation effects and to learn how to control it. It is found that the relaxation effects strongly depend on the relative fraction of coexisting phases (for instance, ferromagnetic metallic (FMM), antiferromagnetic (AF) insulating with charge-orbital ordered (CO/OO) state, paramagnetic, etc.). We have shown that the relaxation effect in manganites can be controlled by using the doping element. The relaxation effects are enhanced due to both the Fe and BaTiO3 doping in SSMO; however, they are suppressed by Ru substitution in (Sm0.5Gd0.5)0.55Sr0.45MnO3. The mechanism behind these changes in the relaxation effects is discussed. We have also investigated the effect of strain induced disorder on the physical properties of BaTiO3/La0.66Ca0.34MnO3 heterostructure grown on a SrTiO3 substrate where very thin (16 nm) La0.66Ca0.34MnO3 films grown on SrTiO3 are normally subjected to tensile strain. We found that this strain sensitive heterostructure has significant magnetocapacitance effect which is important for technological applications. A discussion of the mechanism behind this magnetocapacitance effect is introduced

Antifreeze proteins (AFPs) evolved in cold-adapted organisms and serve to protect them against freezing cold conditions by arresting ice crystal growth and inhibiting ice recrystallization. The freezing point depression by AFPs is defined as thermal hysteresis (TH) and AFPs are classified as hyperactive (hypAFPs) and moderate according to their TH activities. The mechanism of action of AFPs is not well understood. In particular, it is not clear what determines the concentration dependence of TH and whether the binding of AFP to ice is irreversible. Additionally, it is not known why some types of AFP are hyperactive compared to others and it was suggested that hyperactivity might be related to basal plane affinity of hypAFP to ice.The present study utilizes the techniques of microfluidic devices and fluorescence microscopy to study the interaction of AFPs with ice crystals. With novel temperature controlled microfluidic devices, we showed the accumulation and affinity of hypAFPs on the basal plane of ice. This supports the view that hypAFPs adhere to the basal plane. Additionally, for the first time in literature, small ice crystals of 30-50 µm sizes covered with adsorbed GFP tagged hypAFPs were stabilized in supercooled non-AFP solutions for hours with no observed ice growth in temperature controlled microfluidic devices. Repeated TH experiments of ice crystals incubated in AFP solutions before and after the exchange of liquids in microfluidic devices gave the same TH activity. This finding clarifies our understanding of concentration dependence of TH. Furthermore, we found that hypAFPs protect ice against melting as well as freezing, resulting in superheated ice. Ice crystals were superheated up to 0.5 °C above their equilibrium melting temperatures and remained stable in this superheated state for hours. Measurements of fast melting velocities added additional evidence to the observed superheating of ice in AFP solutions. The experimental results of the current study provide strong evidence that AFPs bind to ice surfaces via irreversible binding. We have demonstrated that the use of microfluidics in combination with fluorescence microscopy is a valuable technique to study the binding mechanisms of AFPs and the concentration dependence of AFP activity.

Many organisms are exposed to subzero temperatures in nature and can survive these temperatures by the effect of antifreeze proteins (AFPs), which inhibit ice crystal growth and change the morphology of ice crystals. Although the effects of these proteins, such as recrystallization inhibition, ice growth inhibition, and crystal habit changes, are known, a conclusive description of the protein-ice crystal interaction including interaction energy, surface coverage, and lifetime of adsorbate has been elusive.In this study, different antifreeze protein constructs are designed and expressed such that they can be conjugated to polymers to increase the thermal hysteresis activity especially at low concentrations. Trimers of these proteins are also constructed using a foldon domain attached to their C-terminus. New constructs of type I and type III antifreeze proteins yield significantly higher thermal hysteresis activities than the native protein. Furthermore, we determine the binding equilibrium constant for a type III fish antifreeze protein and the relationship between thermal hysteresis and surface coverage for this protein. This is possible using experimental data from a two-domain antifreeze protein and its related single domain protein. The classical Langmuir isotherm is used to describe the equilibrium exchange of the single domain type III AFP molecules at the ice crystal surface, while a modification of the Langmuir isotherm is derived to describe the adsorption of the two-domain AFP. Because the protein adsorption is governed by different isotherm relationships, there are two independent data sets allowing the determination of the two unknowns of surface coverage and binding energy. The data yield a binding equilibrium constant of 1.9 mM-1 for the type III AFP-ice interaction. The analysis results in a relationship between surface coverage and thermal hysteresis, as well as kinetic equations of the adsorption of the proteins onto the ice surface.

Antifreeze proteins (AFPs) are class of proteins that protect organisms from damages caused by freezing, either by preventing freezing or minimizing frost damages. AFPs effectively lower the temperature at which water freezes. They are classified by the depression of the freezing temperature compared to the melting temperature, i.e. Thermal Hysteresis activity (TH): moderately active AFPs and hyperactive AFPs. It is still unknown what makes some AFPs hyperactive compared to the much less active classes of AFPs. Previous studies showed that fusion proteins of fish type III AFP bind independently to ice. This conclusion was derived from experiments with bulky proteins that were fused to this moderately AFP. One possible explanation for the increased activity of the hyperactive AFPs is that they might function cooperatively. To investigate this, the hyperactive AFP from the mealworm, Tenebrio molitor (TmAFP), was linked to bulky proteins. In this thesis, these fusion proteins were assayed by a nanoliter osmometer, a device that has been designed to measure TH of AFPs. The results indicate that the addition of large molecules to the TmAFP does not induce any loss of thermal hysteresis activity; these fusion proteins were rather more active than free TmAFP at almost all concentrations. Further, ice crystal morphologies obtained by the fusion proteins were the same as the ones of free TmAFP. Therefore, it is concluded that TmAFPs independently bind to ice and their enhanced thermal hysteresis activity does not result from cooperativity.