MS Theses - DO NOT EDIThttp://hdl.handle.net/10106/117002024-03-28T16:07:20Z2024-03-28T16:07:20ZIdentification of acidic residues in proteins by selective chemical labeling and tandem mass spectrometryhttp://hdl.handle.net/10106/317892023-11-09T22:37:12Z2023-08-15T00:00:00ZIdentification of acidic residues in proteins by selective chemical labeling and tandem mass spectrometry
**Please note that the full text is embargoed until 08/01/2025** Proteins play important role in carrying out a wide range of cellular activities. They are subjected to various post-translational modifications (PTMs) by the addition of functional groups such as acetyl, phosphoryl, and methyl which leads to changes in their biological functions, localization, activity, and structure. The growth of MS technology has immensely facilitated the identification of PTMs comprehensively. This dissertation focuses on mapping these changes using covalent labeling techniques by tagging carboxylic acid residues, which undergo various kinds of PTMs. The second chapter focuses on the methods designed for Affinity Purification Mass Spectrometry (AP-MS) chemical cross-linking (CXL) of protein complexes.
There are numerous methods available for bioconjugation which target cysteine, lysine, and arginine residues. Although carboxyl residues, which are prevalent in proteins, are essential for preserving the protein's functionality, there are currently few accessible selective labeling procedures. We have described a novel reactive probe that allows for the chemoselective modification of acidic residues in peptides and proteins. We evaluated the reactivity of diphenyldiazomethane (DPDAM) in peptides and proteins in the third chapter. The fragmentation patterns of these labeled peptides have been investigated using tandem mass spectrometry.
2023-08-15T00:00:00ZReactive Atmosphere Pyrolysis(RAP) of Silicon Carbo Nitride (SiCN)http://hdl.handle.net/10106/317352023-11-09T22:48:53Z2023-08-17T00:00:00ZReactive Atmosphere Pyrolysis(RAP) of Silicon Carbo Nitride (SiCN)
Polymer Derived Ceramics (PDCs) are synthesized by converting liquid polymer precursors into ceramics through controlled pyrolysis. This study focuses on silicon carbonitride (SiCN) ceramics obtained from Ceraset Polysilazane (Durazane1800) precursor. The influence of pyrolysis atmospheres, including inert (Ar, N2) and reactive (H2) environments, on the thermal conversion of Durazane1800 into SiCN ceramics is investigated. The resulting ceramics' chemical variations are explored through phase analysis using X-ray diffraction (XRD), Raman spectroscopy, and thermogravimetric analysis (TGA). Notably, the impact of hydrogen versus nitrogen atmospheres on the conversion process is analyzed, shedding light on their distinct contributions to the composition and properties of SiCN ceramics. The findings contribute to a comprehensive understanding of PDC synthesis, offering insights into tailoring ceramics for diverse applications.
2023-08-17T00:00:00ZInvestigation of F420-dependent glucose-6-phosphate dehydrogenase variants using pH profiles and stopped-flow spectrometric ligand-binding methodshttp://hdl.handle.net/10106/314322023-06-30T08:26:24Z2022-05-18T00:00:00ZInvestigation of F420-dependent glucose-6-phosphate dehydrogenase variants using pH profiles and stopped-flow spectrometric ligand-binding methods
**Please note that the full text is embargoed until 5/17/2024** ABSTRACT: F420-dependent glucose-6-phosphate dehydrogenase (FGD) from Mycobacteria tuberculosis, catalyzes a reaction the conversion of glucose-6-phosphate (G6P) to 6-phosphogluconolactone using the oxidized F420 cofactor, which is reduced to F420H2. The reaction is important within M. tuberculosis, the cause of tuberculosis disease (TB) due to its health relevance for treatment of multiple drug resistant and extreme drug resistant strains of M. tuberculosis. TB affects millions of people worldwide, and although treatable, multiple drug resistant and extreme drug resistant forms of TB adds strain on the current generation of drugs. Since this enzyme is not found in humans, is an ideal drug target.
A past crystal structure of wild-type FGD from M. tuberculosis proposed that among the conserved residues, H40 and E109 function as the active site base and active site acid, respectively. In contrast, later pH profile studies on Glu109 and His 40 revealed that the while Glu109 acts as the acid, His 40 does not act as an active site base. Hence, our current focus is to determine which active site residue may act as active site base using the FGD variants of E13, H260 and H40. pH dependence studies were conducted to elucidate their roles in catalysis. The time-dependent binding experiments for wtFGD, E13, H40 were performed to get more mechanistic information such as koff and kon to incorporate into our global analyses. This work was conducted in collaboration with other group members and will be discussed here.
2022-05-18T00:00:00ZAn Investigation of Peak Shape Models in Chiral Separationshttp://hdl.handle.net/10106/312302023-06-15T08:27:04Z2023-05-04T00:00:00ZAn Investigation of Peak Shape Models in Chiral Separations
The use of superficially porous particles in chromatography has led to significant improvements in separation efficiency. However, peak asymmetry in enantiomeric separations causes performance comparisons across particle types a challenge. In this study, we screened 28 pharmaceutically relevant compounds and developed practical methods to reduce peak asymmetry in normal phase chiral chromatography. The use of additives was found to be effective in reducing peak tailing for all compounds, including neutrals. Additionally, we observed that solvent mismatch with the eluent can cause system peak interference, which can be managed by reducing injection volumes. To more accurately assess the performance of the separations across the two column types, mathematical models that accounted for peak shape distortions were applied. After minimizing these distortions, we found that the SPP column had more efficient kinetics than the FPP column, despite the latter performing better in some separations. Our findings highlight the importance of optimizing peak shape in enantiomeric separations to achieve accurate and reliable results.
2023-05-04T00:00:00Z