Biochemistry, Biophysics, and Structural Biology Commons^™

Chemical Synthesis Of Sensitive Dna, Komal Chillar

Dissertations, Master's Theses and Master's Reports

Over the past decades, researchers have tried various chemical methods to synthesize modified oligodeoxynucleotides (ODNs, i.e. short segments of DNAs). Traditional ODN synthesis methods require strong basic, and nucleophilic conditions for the deprotection and cleavage of the ODN from the solid support. However, the sensitive ODNs containing labile functionalities are vulnerable to such harsh conditions. Sensitive ODNs have a wide range of applications in research and pharmaceuticals. To synthesize sensitive ODNs, researchers devised different strategies but no practical methods have been developed. To overcome these challenges, we developed alkyl Dim alkyl Dmoc technology. This innovative technology uses weakly basic and …

Impact Of Sample Conditions On Dna Phosphodiester Backbone Bi/Bii Conformational Equilibrium Dynamics, Autumn C. Pilarski

MSU Graduate Theses

DNA damage, such as single base lesions and mismatches, is highly prevalent within cells. If these DNA damage events are not repaired, they could lead to mutations and thus disease and cancer. Intricate repair mechanisms are in place to fix these damage events, one such being Base Excision Repair (BER) and associated enzyme: Thymine DNA Glycosylase (TDG). The first step of this repair process, recognition of the lesion by TDG, is not well understood. The following thesis presents results to better understand the fundamental biophysical question of how a DNA lesion within a mismatch context is recognized in a million …

Functional Characterization Of The Newly Discovered Type V Crispr-Cas Protein Cas12a2, Dylan J. Keiser

All Graduate Theses and Dissertations, Spring 1920 to Summer 2023

Similarly to people, bacteria are under the treat of infection by viruses. To circumvent these threats, bacteria evolve complex immune systems. Our understanding of some of these immune systems has led to many advancements in the field of Biotechnology including tools that made expressing proteins for study in a lab easier, tools that revolutionized the feasibility of gene editing, and tools that could change the way we think about viral diagnostics and cancer therapeutics. A certain type of immune system that bacteria use to fight virus is called a CRISPR system. Presented here is work to understand the function of …

Effects Of Crowding Agents On I-Motif Dna, Hayden Brines

Honors Theses

Deoxyribonucleic acid (DNA) is a well-known double stranded, helical, biological molecule. In addition to its more commonly known structure, DNA can also form more complicated structures like G-quadruplexes and i-motifs (iM). The iMs are formed by cytosine rich DNA and are a four stranded structure that is typically looped around itself. The iM formation is typically pH-dependent and is favored in more acidic conditions; the pKa value is approximately 6.5. This pKa value allows for potential in vivo formation, since the cells have a pH of approximately 7.3. Due to this, iMs are thought to be powerful, innovative molecules for …

Development Of Dual Functional Dna/Rna Nanostructures For Drug Delivery, Vibhav Amit Valsangkar

Legacy Theses & Dissertations (2009 - 2024)

In addition to the traditional biochemical functions, DNA and RNA have been increasingly studied as building blocks for the formation of various 2D and 3D nanostructures. DNA has emerged as a versatile building block for programmable self-assembly. DNA-based nanostructures have been widely applied in biosensing, bioimaging, drug delivery, molecular computation and macromolecular scaffolding. A variety of strategies have been developed to functionalize these nanostructures. The major advantage is that DNA is a very stable molecule and its base-pairing properties can be easily utilized to control and program the formation of desired nanostructures. In addition, some of these DNA/RNA nanostructures have …

Effects Of The Dihydrouracil Lesion On Dna Using 1h/31p 1d And 2d Solution Nmr, Benjamin M. Boyd

MSU Graduate Theses

The effects of the dihydrouracil lesion in DNA were studied using two dimensional NMR spectroscopy. The sequence used was based off of the Drew-Dickerson Dodecamer, with the cytosine in the three position replaced by a dihydrouracil. All of the nonexchangeable proton chemical shifts, with the exception of the H2, H5’, and H5’’, of the lesioned DNA were identified using NOESY spectra and then compared to the chemical shift values of the Drew Dickerson Dodecamer. The largest differences in chemical shifts were observed in the nucleotides neighboring the lesion, both within the strand and on the opposite strand. The imino exchangeable …

Dna Functionalized Nanoparticles In Nanobiosensor And Sensor Array Development For Molecular Diagnostics And In Vitro Identification Of Biomolecules, Mustafa Salih Hizir

Legacy Theses & Dissertations (2009 - 2024)

Nucleic acid technology along with vast variety of nanomaterials has demonstrated a great potential in many applications from biosensing studies to molecular diagnostics, from biomedical and bioanalytical research to environmental analysis. Especially short single stranded (ss) DNA molecules, called oligonucleotides, are extraordinary biopolymers featuring diverse functionality on the nanoparticles thanks to their high degree of programmability, target-specific binding or cleavage, molecular recognition ability, structure-switching capability, and unique interactions at the bio-nano interfaces. Among those, there have been many biosensing applications utilizing ss DNAs and numerous nanomaterials through various detection techniques such as fluorometric, colorimetric or electrochemical methods. Although many groundbreaking …

Mutagenic And Spectroscopic Investigation Of Ph Dependent Cooa Dna Binding, Brian R. Weaver

Chemistry Honors Papers

The carbon monoxide (CO) sensing heme protein, CooA, is a transcription factor which exists in several bacteria that utilize CO as an energy source. CooA positively regulates the expression of coo genes in the presence of CO such that the corresponding proteins may metabolize CO. The present studies have yielded the unexpected result that Fe(III) CooA binds DNA tightly at pH < 7, deviating from all previously reported work which indicate that CooA DNA binding is initiated only when the exogenous CO effector reacts with the Fe(II) CooA heme. This observation suggests that the disruption of one or more salt bridges upon effector binding may be a critical feature of the normal CooA activation mechanism. To test this possibility, several protein variants that eliminated a selected salt bridge for the CooA homolog from Rhodospirillum rubrum were prepared via site-directed mutagenesis. Samples of these variant proteins, which were overexpressed in Escherichia coli, were then characterized by spectroscopic methods and functional assays to investigate the impact these mutations had on CooA heme coordination …

Silica Nanoparticles For The Delivery Of Dna And Rnai In Cancer Treatment, Michael Aaron Vrolijk

Graduate College Dissertations and Theses

DNA and interfering RNA (RNAi) – short interfering RNA (siRNA) and micro RNA (miRNA) – are promising new cancer therapies, especially for drug resistant lines. However, they require a delivery system in vivo to prevent degradation and off target effects. Silica based nanoparticles, both solid and mesoporous, are a promising option due to their biocompatibility, ease of preparation and morphology control, reproducibility, and facile addition of functional groups including targeting ligands.

After a brief introduction to cancer treatment and review of the current nanoparticle treatments undergoing clinical trials, this thesis details the many methods explored over the past ten years …

X-Ray Characterization Of Mesophases And Phase Transitions Of Dna Analogues In Solutions, Mustafa Selcuk Yasar

Doctoral Dissertations

We think of DNA as double-stranded helices (duplex), but the polymer exists in many conformations. Several triplex and quadruplex DNA structures can be formed in laboratory settings and exist in nature. This thesis first provides a brief description of the nature of the order in arrays of duplex DNA under biologically relevant molecular crowding conditions. Then we compare the duplex DNA mesophases with the corresponding liquid crystalline phase behavior of the triplex and quadruplex DNA analogues. In particular, we focus on G-quadruplexes. Observed in the folds of guanine-rich oligonucleotides, G-quadruplex structures are based on G-quartets formed by hydrogen bonding and …

Specific Binding Affinity Of The Non-Catalytic Domain Of Eukaryotic Like Type Ib Topoisomerase Of Vaccinia Virus, Benjamin R. Reed

Dissertations, Theses, and Capstone Projects

Topoisomerases are ubiquitous proteins that alter supercoiling in double stranded DNA (dsDNA) during transcription and replication and. vaccinia and the closely related poxvirus variola virus, at 314 amino acids in length, encode the smallest of the type I topoisomerases(TopIB). TopIB is a two domain protein that recognizes the sequence 5’-T/CCCTT, cleaves at the 3’-end and relaxes supercoiling through rotation. The C-terminal domain (CTD) alone contains the catalytic activity and specificity. Deletion of the N-terminal domain results in a greatly reduced rate of relaxation and rapid dissociation. Biochemical data suggests that the N-terminal domain (NTD) is important for pre-cleavage binding and …

Dna Aptamers Selected Against Wild-Type Helix 69 Ribosomal Rna And Their Implications In Combating Antibiotic Resistance, Sakina Miriam Hill

Wayne State University Dissertations

Outbreaks of advanced antibiotic-resistant strains of microbes have hastened the need to identify new viable molecular targets for the development of novel anti-infectives. For this purpose, helix 69 (H69, or m3a 19-nucleotide (nt) hairpin motif that is highly conserved throughout phylogeny and rich in modified nucleotides, including pseudouridine () and 3-methylpseudouridine (m3) was chosen as a potential target. Helix 69, which is located in domain IV of Escherichia coli 23S ribosomal RNA (rRNA), undergoes conformational changes when in close proximity to the decoding region of 16S rRNA and transfer RNAs (tRNAs) in the peptidyl-transferase center (PTC). Functionally, the exact biological …

Artificial And Natural Nucleic Acid Self Assembling Systems, Marcus Wood

Wayne State University Dissertations

Nucleic acids are good candidates for nanomachine construction. They participate in all the processes of life, and so can function as structural building blocks and dynamic catalysts. However, to use nucleic acids as nanomachines, a better understanding of their material properties, how to design structures using them, and their dynamics is needed. We have tried to address these issues, in a small way, with nucleic acid force field development, an attempt at nanostructural design and synthesis using DNA, and a study of the RNA/protein regulatory dynamics of the tryptophan regulatory attenuation protein.

Dna-Templated Nanomaterials, Hector Alejandro Becerril-Garcia

Theses and Dissertations

Nanomaterials display interesting physical and chemical properties depending on their shape, size and composition. Self assembly is an intriguing route to producing nanomaterials with controllable compositions and morphologies. DNA has been used to guide the self assembly of materials, resulting in: (1) metal nanowires; (2) metal or semiconductor nanorods; (3) carbon nanotubes; and (4) semiconductor, metal or biological nanoparticles. My work expands the range of DNA templated nanomaterials and develops novel ways of using DNA to pattern nanostructures on surfaces. I have performed the first synthesis of silver nanorods on single stranded DNA, an attractive material for localizing DNA coupled …

The Role Of The Voltage Gradient In The Agarose Gel Electrophoresis Of Dna, David Wheeler

Theses and Dissertations in Biomedical Sciences

In Part I of this dissertation, empirical equations for predicting DNA mobility during agarose gel electrophoresis (AGE) from voltage gradient are developed from the data of McDonnel (36) for electrophoresis in a 1.6% agarose gel. These equations represented the data well for DNA between 2 and 10 kilobase pairs (KBp) in length. A computer program, called GELSIM, which incorporates these equations is described in Part II. GELSIM was designed to allow researchers to analyze electrophoresis data by predicting the effect on DNA migration of altering the voltage of electrophoresis. In this way, electrophoretic banding patterns produced using different voltages could …

Transforming Ability Of Bacillus Subtilis Dna Taken Up By Barley Embryos, Christine Tolman Ence

Theses and Dissertations

Purified Bacillus subtilis DNA taken up by one-day-old barley embryos and permanently fixed within the barley cell is shown to have maintained its biological activity in B. subtilis transformation. Cesuim chloride density gradient centrifugation of DNA isolated from the roots of barley embryos after the uptake and incorporation of radioactive, bromouracil-labelled B. subtilis DNA indicates the presence of the bacterial DNA in several discreet bands of well-defined density, whose density is changed by sonication. These preliminary findings concerning the fate of bacterial DNA taken up by barley roots are discussed in terms of the recombination model of Ledoux and Huart …