Biochemistry Commons^™

Erecta Family Genes Regulate The Shoot Apical Meristem And Organ Formation, Daniel A. Degennaro

Doctoral Dissertations

Plants are sessile and must adjust their organ growth to their environments. A reservoir of stem cells in the shoot apical meristem (SAM) supplies cells for differentiation into organs. The SAM must balance organ production with stem cell maintenance. The ERECTA family (ERfs) encodes the leucine-rich repeat receptor-like kinases ERECTA (ER), ERECTA-LIKE 1 (ERL1), and ERL2. ERf signaling regulates organ initiation and stem cell maintenance. Results presented in this work include the following:

1) WUSCHEL (WUS) and CLAVATA3 (CLV3) make up a negative feedback loop to maintain SAM size. WUS and CLV3 expression localization is critical for …

Understanding How Camkii Holoenzyme Dynamics Facilities Activation-Triggered Subunit Exchange, Ana P. Torres-Ocampo

Doctoral Dissertations

Long-term memory and learning are still poorly understood from a molecular and cellular standpoint. Ca²⁺/calmodulin-dependent protein kinase II (CaMKII) is an oligomeric kinase that is involved in this remarkable process. However, the molecular details of its specific roles in these processes remains elusive. CaMKII activation-triggered subunit exchange presents a novel possible mechanism involved in long-term memory and learning by exchanging active subunits with other CaMKIIs. CaMKII subunit exchange also shows that exchanged CaMKIIs spread their phosphorylation state to newly synthesized CaMKIIs. This provides a long-lasting signal that might possibly be involved in long-term memory by escaping a cell’s …

Mechanistic Insights Into Diverse Protease Adaptor Functions, Nathan J. Kuhlmann

Doctoral Dissertations

Protein degradation is an essential cellular process that helps maintain proper homeostasis. The ClpXP protease broadly regulates bacterial development and quality control during the cell cycle. The range and order of substrates that ClpXP degrades during the cell cycle is dictated by 3 accessory proteins, which are known as adaptors. This thesis will elaborate on how dimerization tightly regulates the stability and activity of the adaptor protein at the center of this hierarchy, RcdA, and show how this affects normal cellular processes in Caulobacter crescentus. I will discuss the mechanism by which dimerization limits RcdA activity and how the dimerization …

Amyloidogenesis Of Β-2-Microglobulin Studied By Mass Spectrometry And Covalent Labeling, Blaise G. Arden

Doctoral Dissertations

Amyloid-forming proteins are implicated in a number of debilitating diseases. While many amyloid-forming proteins are well studied, the early stages of amyloidosis are still not well understood on a molecular level. Covalent labeling, combined with mass spectrometry (CL-MS), is uniquely well suited to provide molecular-level insight into the factors governing the early stages of amyloidosis. This dissertation leverages CL-MS techniques to examine the early stages of β-2-microglobulin (β2m) amyloidosis. β2m is the protein that forms amyloids in the condition known as dialysis-related amyloidosis. An automated CL-MS technique that uses dimethyl(2-hydroxy-5-nitrobenzyl) sulfonium bromide as a labeling reagent was developed and used …

Primary Kinetic Isotope Effect For Substrate Hydroxylation And Iron Binding Regulation In Factor Inhibiting Hif-1, Michael A. Mingroni

Doctoral Dissertations

The hypoxic response is a vast and complex system, delicately designed through evolution to allow our tissues to rapidly adapt and survive fluctuations in pO2. HIF1α, the master regulator of oxygen, is tightly controlled through oxygen-dependent, post-translational hydroxylation via PHD2 and FIH1.The aberrant stabilization of HIF1α is an adverse consequence of disease states, but can also occur under normoxia in the presence of ROS or in iron depletion. The extensive transcriptional network regulated by HIF1α makes the HIF pathway an attractive therapeutic target, particularly through the inhibition of the hydroxylases, PHD2 and FIH, however the active site chemistry is mechanistically …

Novel Approaches Towards Improved Purity In High Yield Transcription Reactions, Elvan Cavac

Doctoral Dissertations

High yields of RNA (e.g., mRNA, gRNA, lncRNA) are routinely prepared following a two-step approach: high yield in vitro transcription using T7 RNA polymerase, followed by extensive purification using gel or chromatic methods. In high yield transcription reactions, as RNA accumulates in solution, T7 RNA polymerase rebinds and extends the encoded RNA (using the RNA as a template), resulting in a product pool contaminated with longer than desired, (partially) double stranded impurities. Current purification methods often fail to fully eliminate these impurities which, if present in therapeutics, can stimulate the innate immune response with potentially fatal consequences. This study establishes …

Pharmacological Chaperoning Of Human Lysosomal Neuraminidase 1, Di Chu

Doctoral Dissertations

Human lysosomal neuraminidase 1 (hNEU1) is an exo-a-sialidase which cleaves a(2-3) and a(2-6) linked sialic acids on glycoproteins in the lysosome. Deficiency of hNEU1 in the lysosome results in sialidosis, a lysosomal storage disease. Currently there is no effective treatment for sialidosis, which leads to a rising interest in discovering potential therapies. Here we presented a small molecule, α-D-N-acetylneuraminic acid (NANA), increases the protein amount and activity of both wild-type hNEU1 and three different hNEU1 mutations found in sialidosis patients in our mammalian cell system, suggesting that NANA works as a potential pharmacological chaperone for hNEU1 and provides …

The Hidden Life Of Tropical Roots: Functional Root Traits And Their Response To Climatic Disturbances, Daniela Yaffar

Doctoral Dissertations

Roots play a critical role in plant nutrition, and terrestrial carbon cycling. However, they are often understudied compared to their aboveground counterparts; especially in the tropics, where more carbon is cycled than in any other ecosystem. Some tropical forests, like in Puerto Rico, are more represented in scientific studies than others. However, this information is sparse, complicating the interpretation of root trait patterns. Trees in Puerto Rico have adapted mechanisms for withstanding hurricane disturbances, including in their roots. Additionally, as many tropical forests, some in Puerto Rico have low available phosphorus (P); thus, trees rely on root traits to enhance …

Engineering Modularity Of Ester Biosynthesis Across Biological Scales, Hyeongmin Seo

Doctoral Dissertations

Metabolic engineering and synthetic biology enable controlled manipulation of whole-cell biocatalysts to produce valuable chemicals from renewable feedstocks in a rapid and efficient manner, helping reduce our reliance on the conventional petroleum-based chemical synthesis. However, strain engineering process is costly and time-consuming that developing economically competitive bioprocess at industrial scale is still challenging. To accelerate the strain engineering process, modular cell engineering has been proposed as an innovative approach that harnesses modularity of metabolism for designing microbial cell factories. It is important to understand biological modularity and to develop design principles for effective implementation of modular cell engineering. In this …

An Investigation On The Regulation Of The Tweak–Fn14–Nf-Kb Pathway, Jawahar Khetan

Doctoral Dissertations

“Dysregulation of inflammatory pathways is strongly implicated in cancers and autoimmune diseases. The most consequential of these pathways involves the nuclear translocation of NF-kB, a transcription factor that induces the transcription of multiple proteins associated with cell survival, inflammation, proliferation and death. It is activated when the fibroblast growth factor-inducible 14 kDa protein (Fn14), a trimeric receptor recruits its ligand, TWEAK. Studies have shown that Fn14 is over-expressed in many tumors, the aggressiveness of which is often correlated with the degree of upregulation. Furthermore, TWEAK-Fn14 activation has been shown to result in persistent NF-kB activation. Using a mechanistic model of …

High Throughput Analysis To Study Emerging Pollutants And Nanoparticle Fate In Biological Systems, Xiaolong He

Doctoral Dissertations

”The increasing applications of emerging and fugitive contaminants (EFCs) and engineered nanoparticles (ENPs) attract significant research interest for their potential risks to human health and the environment. In order to assess the health risks of these emerging contaminants, rapid and reliable analytical methods to measure the concentrations and fates of these contaminants are imperative. This dissertation focuses on the developments of advanced analytical methods and their applications to study those emerging contaminants in crop plant and simulated gastric fluid (SGF). Three types of mass spectrometry based methodologies have been developed, one is freeze-thaw/centrifugation extraction followed by high performance liquid chromatography …