Enzymes have recently been incorporated into multiple high-value industrial syntheses, demonstrating the utility of enzymes as highly selective catalysts for practical industrial processes. However, the current scope of non-biological enzymatic reactions is narrow and new reactions and reaction pathways need to be engineered. The goal of our work in the Zalatan lab is engineering enzymes as catalysts in carbon-hydrogen bond functionalization reactions, a transformation critical for practical industrial synthesis where selective catalysis is still a major challenge. Importantly, we are interested in exploring more efficient and informed engineering approaches by establishing structure-function relationships with the enzymes that we work with. Our model system for this work is the non-heme iron(II) 2-oxoglutarate dependent oxygenase superfamily (Fe(II)-2OGs). We are using a high-throughput microfluidics based kinetic assay to determine key sites that we can target for mutagenesis and directed evolution in a candidate Fe(II)-2OG found to catalyze a new reaction. Overall, we expect that this work will enable new directions and principles for engineering Fe(II)-2OGs, and that lessons learned here can then be extended to additional industrially-relevant enzyme families.

Irritable bowel syndrome (IBS) is a disease that affects many people worldwide. Clinical trials using probiotic bacteria to treat IBS show promising outcomes, although the molecular mechanisms of probiotics and their effect on gut health is not completely understood. Multiple studies comparing different strains of probiotics show Lactobacillus plantarum and Bifidobacterium longum as the most effective strains in inhibiting the release of proinflammatory factors in the gut and promoting proliferation of healthy gut bacteria. The purpose of this study is to evaluate the protein profile of probiotic bacteria Lactobacillus plantarum and Bifidobacterium longum in both healthy gut and IBS gut conditions. To observe how Lactobacillus plantarum and Bifidobacterium longum respond to different simulated gut conditions, samples of the bacteria will be isolated from probiotic supplements. These samples will be grown in aliquots representing different simulated anaerobic gut conditions for 12-16 hours at 37°C. Lactobacillus plantarum and Bifidobacterium longum will be grown separately and grown combined within these conditions. To enrich secreted proteins, the supernatant from each aliquot sample will be clarified using centrifugation. Secreted proteins will be solubilized and denatured with sodium deoxycholate. The denatured proteins will be reduced with dithiothreitol and alkylated with iodoacetamide. The sample will be diluted using ammonium bicarbonate and digested by trypsin. Tryptic peptides will be analyzed using liquid chromatography and mass spectrometry. Tryptic peptides will be identified using the SEQUEST search engine with a reference database. I anticipate the probiotic strains will demonstrate differential proteomics when isolated and combined in the healthy gut and IBS gut conditions. The secreted protein analysis completed in this study will further the understanding of the probiotic molecular mechanism of promoting gut health.

Metal–organic frameworks (MOFs) are crystalline, porous extended solids that are formed through coordination between metal cations and bridging organic ligands. Since their discovery in the late 1990s, MOFs have been a topic of acute interest in the scientific community due to their intrinsic porosity, high surface area, and precise tunability. However, MOFs are typically insulating, which limits the scope of their applications. The recent development of electrically conductive MOFs has opened the door to exciting multifunctional applications in electrocatalysis, advanced electrochemical energy storage, chemical sensing, and much more. However, a molecular-level understanding of charge transport in MOFs remains lacking. My research aims to address this knowledge gap through the investigation of one-dimensional (1D) metal organic chains. These 1D chains can be thought of as the primary subunit of higher-dimensional MOFs; they allow for high synthetic and electronic tunability, making them ideal model materials for studying the genesis and tuning of electronic properties in conductive MOFs. Here, I will present the synthesis of a series of highly-tunable 1D metal–organic chains that exhibit delocalized π systems and high electrical conductivity along with our studies of how structural parameters such as metal identity, chain structure (linear vs. zig-zag), and metal/ligand oxidation state can influence the overall electrical and magnetic properties of the resulting chain.

The Wnt signaling pathway plays a critical role in mammalian cell development and regulates cell growth and differentiation. Two central proteins in this pathway are glycogen synthase kinase 3β (GSK3β) and β-catenin. Another protein, Axin, is responsible for holding these two proteins in close proximity in order to promote the reaction between them. The kinetic mechanism for the Axin-mediated reaction is well understood, but the relationship between this function and Axin’s structure is poorly characterized. I am currently generating a cryogenic electron microscopy (Cryo-EM) structure of Axin bound to GSK3β and β-catenin in order to investigate this mechanism. Cryo-EM is a technique for determining the structures of proteins that is especially suitable for large protein complexes, such as the one I have produced. Inital results indicate that the stoichiometry of the complex is much more complicated than initially assumed and does not follow the predicted 1:1:1 complex. Elucidating this structure will provide insight into how the structure of Axin promotes the reaction between GSK3β and β-catenin as well as how it provides Wnt signaling specificity. This structure will also be important for understanding and intervening in diseases such as cancer where Wnt signaling is dysregulated.

Test questions are commonly written with the intent to assess a student's higher level thinking skills, but the phrasing of a question may not align with this intention. One way to remedy this is to assess the cognitive complexity of existing test questions, and use these findings to reform instructional materials and subsequent examinations. We used Marzano’s Taxonomy to characterize a bank of multiple-choice exam questions from general chemistry by the level of cognitive complexity required of students' thinking. Marzano’s Taxonomy has been applied to introductory physics questions to characterize quantitative thinking and problem-solving skills (Teodorescu et al., 2013), and to general chemistry curricula to guide course and assessment development (Toledo & Dubas, 2016). Here we will describe our implementation of Marzano's Taxonomy in the context of large-lecture general chemistry, and our development of a rubric that chemistry instructors can utilize to evaluate their own exam questions. The development team included an undergraduate biology major and former general chemistry peer mentor; a chemistry graduate student and experienced general chemistry teaching assistant; and a chemistry faculty member. When applied to multiple choice questions, we found that the cognitive level of most questions was lower than expected, even for lengthy questions that are normally considered challenging by students and instructors alike. We will discuss ways to increase the cognitive complexity of problems to allow instructors to elicit the intended level of thinking from the student, and to align the cognitive levels of assessment and instruction. This can also improve the clarity of expectations of the cognitive complexity that is required of the student and promote higher-order thinking.

Standard diagnostic tests for common bacterial infections such as strep throat are invasive and uncomfortable, especially for children. Difficulty performing these tests may delay or prevent diagnosis, which leads to more severe consequences, such as secondary conditions or organ damage. To address these barriers to care, we developed the CandyCollect, a novel saliva collection platform that is child-friendly, effective at pathogen collection, and suitable in at-home and clinical settings.The purpose of this study is to test the feasibility of our device’s bacteria capture for home use in healthy people. Staphylococcus aureus and Streptococcus mutans are common oral bacteria that are targeted in this study to assess our device’s ability to capture and detect bacteria. The findings in this study will inform our ability to use the CandyCollect for the bacterial pathogen responsible for strep throat, Streptococcus pyogenes, in future studies. We initally recruited 5 participants (>18 years) nationwide to test our CandyCollect device against oral swabs and spit tubes to compare the efficacy and user feedback of CandyCollect with traditional methods of saliva collection. The study is continuing with additional participants. qPCR analysis and fluorescent imaging were used to detect the target oral bacteria in collected samples. Responses from the user feedback survey indicated that the majority of the participants were satisfied with the design and were optimistic about implementing this device for children. Future applications include quantitative determination of bacteria concentration as well as targeting S. pyogenes. We aim to expand this human subjects study to recruit younger participants, especially children (> 3 years), which will help us achieve the ultimate goal of delivering a comfortable saliva collection platform to pediatric patients.

Microfluidics is the technology of systems in which microscale channels are used to manipulate small quantities of fluids (microliter to picoliter or less). Open microfluidics provides a platform to control the movement of microscale volumes of liquid in open space, making every position more accessible than conventional closed microfluidics. In many applications–including cell culture, chemical synthesis, and high throughput screening–merging droplets is essential for the experimental workflow. For example, merging droplets containing reagents can initiate a chemical reaction, or adding a drug to a cell culture can stimulate cells. In conventional closed droplet-based microfluidics, merging techniques often rely on external components such as merging by electrofusion using electrodes. In other cases, like merging using hydrophilic strips in a channel, the merging method is built into the device and the location of merging must be determined in advance. Therefore, the ability to easily initiate droplet merging in an open microfluidic system would be beneficial to researchers and offer flexibility in experimental design and applications. In this study, I developed an easily accessible droplet fusion technique with the prick of a needle at the liquid interface; my method fuses multiple droplets simultaneously (2-15 droplets). Importantly, this fusion method can be used on-demand at any point in the open microfluidic device and does not require external equipment nor features built into the device. This presentation characterizes the experimental parameters required for successful and controlled droplet fusion and explores the physics behind this phenomenon.

The construction sector has always had a significant influence on CO₂ footprints in a given area (Pomponi, 2021). However, there are no recent studies quantifying the effect of construction activities on localized CO₂ concentrations. Previous research conducted in the early 1990s has shown that construction increases CO₂ concentrations by over 11% (Mazria, 2018). However, this research was done over three decades ago, and since then, CO₂ concentrations have increased globally over 40%, necessitating updated measurements. My research group and I will investigate construction sites of various degrees in the Puget Sound region to determine the effect of construction activities on local CO₂ concentrations. Preliminary data has shown an increase of 140 ppm (±8 ppm) in CO₂ concentration when implementing a method of measuring CO₂ concentrations in regions of high and low construction activity within urban, suburban, and rural areas. This research is key to understanding the health implications of the increased construction within the Puget Sound region and its effects on our ecosystems.

Since the beginning of the COVID-19 outbreak in 2020, many students have participated in online learning. Online students spend many hours every day studying at home in indoor spaces that lack ventilation and may have hazardous air quality. Prior research conducted by Tyler Jacobsen et al, (2019) reveals that CO₂ concentrations over 1,000 parts per million (ppm) can increase physiological stress and lower cognitive abilities. The goal of this project was to determine if the CO₂ levels in the at-home study environment of a student conducting online learning are higher than those of a student conducting in-person learning during study times. I predicted that if a student participates in online learning, then the CO₂ levels in their study environment will be higher than those in the study environment of a student conducting in-person learning during school hours (8 am-3 pm), but the levels will be similar during homework hours (4 pm-5 pm). A CO₂ air monitor collected data in the study rooms of an online learning student and an in-person learning student. Over several days, I recorded data every hour from 8 am to 5 pm, resulting in a time series showing CO₂ concentrations throughout a school day. Preliminary data reveals that the online student’s room consistently exceeds 1,000 ppm and varies greatly throughout the day, while the in-person student’s room remains below 1,000 ppm until they arrive home at 4 pm, at which point levels increase significantly. I only observed two locations during this quarter-long research study in General Chemistry and I need to collect data at more locations to determine if this study’s findings are representative of online learning versus in-person learning as a whole. Findings may be useful to educators when they are deciding if online education is a viable option for students in the future.

A critical strategy for achieving carbon neutrality is carbon dioxide (CO2) storage and sequestration, wherein CO2 is removed from emissions and stored underground. Current state of the art technologies, such as aqueous amine solutions, suffer from poor stability and recyclability primarily due to their homogenous nature. Metal–organic frameworks (MOFs), extended porous crystalline networks of metal nodes connected by organic linkers, have emerged as an exciting class of potential solid state CO2 storage materials owing to their high internal surface area and facile tunability. Here, we outline the synthesis of an expanded framework analogue of Matériaux de l′Institut Lavoisier-53 (MIL-53) framework functionalized with either ester based crosslinked ligands or bulky tert-butoxycarbonyl (BOC) protected carboxylates or amines, both of which prop open the pores of the framework. Expanded MIL-53 was characterized by powder x-ray diffraction, gas sorption analysis, thermogravimetric analysis, and proton nuclear magnetic resonance (NMR) digestion experiments. These myriad techniques reveal that the expanded MIL-53 framework’s surface area increases with higher incorporation of either ester based crosslinkers or bulky BOC groups. Subsequent thermal removal of the crosslinker or BOC groups to reveal the respective free functional group proceeds without compromising the structural integrity of the framework. Notably, the resultant ‘open’ pore structure is inaccessible without first propping the pores open. Future work will focus on investigating the carboxylate functionalized and amine functionalized frameworks for gas separations and storage.

Chronic inflammation in the lung often leads to airway remodeling, which can worsen symptoms in inflammatory diseases such as asthma. Airway remodeling is attributed to the excessive deposition of the extracellular matrix (ECM) by myofibroblasts, which are a differentiated form of fibroblasts. Eosinophils, when activated by interleukin-3 (IL-3), release certain soluble factors that were found to be associated with inflammation in asthmatic tissues. Hence, it is crucial to study cellular communication in airway remodeling to facilitate the development of treatments. The aim of this project is to establish an in vitro model of asthma by coculturing primary human lung fibroblasts and eosinophils to study the soluble factors that trigger airway remodeling. We hypothesize that IL-3 activated eosinophils, when immunoglobulin (IgG) is added, release soluble factors that trigger the gene expression and phenotypic changes in fibroblasts. The coculture device has two chambers, in which two types of cells can be cocultured in the shared media while being physically separated by a half wall. Eosinophils are seeded in the outer chamber of the devices and degranulated. The differentiation of fibroblasts would then be quantified by utilizing immunocytochemistry to see the differences in expression levels of alpha smooth muscle actin (ɑSMA) in fibroblasts, in addition to quantitative polymerase chain reaction (qPCR) to detect messenger RNA (mRNA) level associated with inflammation and tissue remodeling. The initial experiments were focused on the monoculture of fibroblasts to ensure that reliable readouts can be obtained from fibroblasts before initiating the coculture. The preliminary data suggest that the fibroblasts treated with transforming growth factor beta 1 (TGF-β1), which promote differentiation, result in significantly higher expression of ɑSMA. Our future experiments include initiating the coculture of eosinophils and fibroblasts to fully illustrate this crucial cellular communication in airway remodeling.

Solar panels are a compelling renewable energy alternative to fossil fuels for reducing greenhouse gas emissions and combating global climate change. Mainstream photovoltaic technologies do not effectively utilize photon energies in the near-infrared, a large portion of the solar spectrum. Triplet-triplet annihilation upconversion (TTA UC) provides a method to convert low energy, red and near-infrared photons to higher energy photons to increase the efficiency of solar cells. Previous projects in our lab have shown that a thiosquaraine molecular sensitizer combined with a rubrene emitter produce successful red to yellow upconversion. In this project, we use a combination of theoretical computational chemistry and experimental photoinduced absorption spectroscopy to study the molecular properties of oxo- and thiosquaraines with various arylamine donor moieties. Our goal is to identify properties that lead to near-infrared absorbance, high triplet yields, and efficient intersystem crossing from the first singlet to first triplet excited state. We report the absorbance energy for squaraines with donor moieties in perpendicular and parallel configurations relative to the molecular axis, as we hypothesize that the dihedral angle of the donor relative to the molecule’s core affects the effective conjugation length of the molecule and its absorbance energy. We also compare the absorbance energy for donor moieties containing different numbers of amine groups, which are correlated with the strength of the donor in the excitation. Using computational chemistry to understand these trends is more efficient than synthesizing each squaraine and will serve as a guide for which donor groups to test experimentally. For example, increasing the number of aromatic rings on the donor redshifts the absorbance energy, indicating that such squaraines may exhibit the properties we want in experiment. We will synthesize and perform photoinduced spectroscopy on the most promising candidate(s) to determine whether they exhibit the desired near-infrared absorbance and high quantum yield.

Exhibiting narrowband photoluminescence and tunable surface functionalization, colloidal semiconductor nanostructures are promising candidates for wide-ranging applications from light-emitting technologies to quantum information processing. In recent decades, the syntheses of II-VI and III-V quantum dots, nanorods, and nanoplatelets have been in the spotlight of nanotechnology, whereas the potential of perovskite quantum dots, a zero-dimensional variant, remains to be fully tapped. Boasting fast radiative lifetimes and long optical coherence times, perovskite quantum dots are ideal candidates for coherent single-photon emitters in integrated photonic circuits. Integrated quantum nanophotonics utilize light-matter coupling with embedded light-emitting media to achieve state-of-art engineering processes such as ultralow threshold lasing. However, the scalable integration of single photon emitters in nanophotonic platforms remains a challenge. The subject of this research presents a solution to this problem: giant lead halide perovskite quantum dots for deterministic positioning into optical nanocavities. Owing to the exceptional tunability of their surface chemistry, perovskite quantum dots can be efficiently solution processed with clear pathways for deterministic positioning. Size and compositional modulation of these nanomaterials enable optimization of single-photon behaviors, especially high quantum yields and narrow emission linewidths. This investigation enlists a solution-based colloidal synthesis combined with silica encapsulation techniques to afford giant lead halide perovskite quantum dots with desirable sizes, quantum yields, absorption and emission signatures, and narrow photoluminescence linewidths well-suited for single-photon applications. Experiments are designed to 1) reproducibly synthesize photostable and highly luminescent silica-coated perovskite quantum dots, and 2) deterministically position the quantum dots on silicon nitride nanobeam cavities. All products are analyzed using UV-vis and fluorescence spectroscopies, along with transmission electron microscopy. The outcome of this study also paves the way for understanding the robust single-photon properties of perovskite quantum dots in optical cavities, thereby providing insights into the design of colloidal single-photon emitters in other morphologies.

Given the harsh impact of climate change and industrial pollution on the planet, it is imperative that we lower the energy consumption of chemical synthesis and reduce hazardous byproducts. As many organic reactions involve oxidation and reduction, electrocatalysis has been solidified as a promising route toward cleaner, more energy-efficient redox reactions. However, high energy barriers for charge transfer between electrodes and dissolved reactants can serve as a critical obstacle. We propose the use of supramolecular cages with redox-active ligands as electrocatalysts to address this challenge. Supramolecular cages are compounds formed from the self-assembly of organic ligands and metal ions, with large, open cavities that could act as hosts for reactive species. Electrocatalysts are compounds that reversibly transfer charge from electrodes to dissolved reactants, lowering the overall energy barrier. Specifically, we are targeting a giant electroactive Fe2+ cage with perylene-bisimide ligands as an electrocatalyst for the reduction of vicinal dihalides, perfluoroalkyl iodides, and aldehydes. We have made significant progress in the five-step synthesis of the perylene-bisimide ligand required for cage assembly and have conducted preliminary cyclic voltammetry experiments for the vicinal dihalide reduction. Ultimately, our goal is to use this supramolecular cage to establish new electrocatalytic pathways for the synthesis of industrially relevant compounds and chemical feedstocks.

Proton-coupled electron transfer (PCET) has been studied extensively, with the anticipation that it could enable applications in energy storage and conversion. We have previously studied 2,5,8-tris(4-methoxyphenyl)-1,3,4,6,7,9,9b-heptaazaphenalene (TAHz), a derivative of carbon nitride, as a promising organic photocatalyst. This is because the heptazine core of TAHz is composed entirely of carbon and nitrogen, while most photocatalysts comprise rare transition metal centers. Additionally, TAHz has been spectroscopically shown to form excited-state complexes with hydroxylic species. However, practical organic synthesis using TAHz as a photocatalyst has proven difficult due to its low solubility. Thus, we synthesized heptazine-based molecules with better solubility and more electron-withdrawing functionalities so future photochemical reactions can be more practically carried out. As a future direction, we plan to explore these new derivatives in the direct oxidative C-H amidation of the model substrates N-methylindole and N-methyl-para-toluenesulfonamide by PCET. This model reaction may improve upon previous catalytic methods in that the photocatalytic mechanism with heptazines replaces the need for heavy-metal photocatalysts and stoichiometric oxidants. Demonstrating the efficacy of this reaction may also present new opportunities for larger-scale application of heptazine derivatives to energy storage devices.

Polymers are important in all areas of life from commodity plastics to vaccine delivery. Currently, plasticizers are small molecules used to make rigid polymers into the flexible plastics we need. However, the problem with these plasticizers is that by disrupting intermolecular polymer chain interactions, they can degrade the polymer over time and leach out into the environment. When plasticizers escape, they can be toxic to human health and harmful to the environment. By inserting a plasticizer into the backbone of the polymer rather than into the polymer solution, we reduce the risk of toxic molecules escaping the polymer. To solve this problem, a fluxional molecule such as bullvalene can be inserted into the backbone of the polymer. Bullvalene is a small molecule and due to its fluxional property, it will induce flexibility into the polymer as an internal plasticizer. Bullvalene is considered fluxional because it can undergo Cope rearrangements at room temperature. The purpose of this research is to develop the methods for synthesizing a novel polymer and to analyze its chemical and mechanical properties such as glass transition temperature and tensile strength. My project focuses on polycarbonates, a specific class of polymers. Polycarbonates are in many plastic materials such as water bottles and are increasingly important for consumer electronics. The results will have positive implications for human and environmental health by providing methods to develop safer plastic material.

With the shift towards renewable energy and the promising performance that has been shown by organic-inorganic halide perovskite thin films to harvest solar energy, extensive research efforts have been devoted to further enhancing perovskite solar cell performance. To bring the power conversion efficiency of perovskite solar cells closer to the Shockley-Queisser limit, a high-quality perovskite film with fewer defects is desired. Antisolvent-assisted crystallization is a good way to get compact and homogeneous perovskite films with less defect density. Past studies have demonstrated that additive may be used in both the antisolvent and as a post-treatment to fine-tune perovskite crystallization and passivate defects at grain boundaries or at the surface. Here, we study the influence of multiple guanidinium molecules on the optoelectronic property of wide bandgap perovskite films by using a combination of steady-state and time-resolved photoluminescence (PL). In our testing, we have seen a significant increase in PL emission and PL lifetime of guanidinium molecule treated perovskites, qualitatively indicating that perovskite films with lower defect density is obtained. X-ray diffraction (XRD) and atomic force microscopy (AFM) are used to investigate the effect of these molecules on the crystallinity of wide bandgap perovskite films. From these we confirmed the ability of the tested guanidinium molecules in reducing the defect density of halide perovskite films via antisolvent-assisted crystallization method. This understanding would help us to further push the efficiency of perovskite solar cells and ultimately reach a full carbon-neutral environment.

 Quantum dots are semiconducting nanomaterials that are useful in converting solar energy because of their high absorption and tunable photophysical and chemical properties. They can have various organic ligands bound to their inorganic surface, creating quantum dot-ligand systems useful for charge and energy transfers. When a quantum do is excited by external radiation, an electron inside the dot is promoted to a higher energy level and can be extracted form the quantum dot through the ligands surrounding the dot. We hypothesize that by binding the charge accepting ligand directly to the quantum dot, charge extraction from the quantum dot will be promoted. In my experiments, I am trying to determine the extent of ligand binding to the quantum dot and how this binding is affected by the identity of the binding moiety. To do this, I am synthesizing cadmium sulfiide quantum dots and attaching various ferrocene derivatives through a titration experiment. I analyze each step of the titration experiment for each quantum dot-ligand system with cyclic voltammetry (CV), an electrochemical technique that is sensitive to the diffusivity of the charge acceptor. The resulting CV curve can then be modeled with DigiElch to extract the favorability of the binding of the ligand to the quantum dot. Based on the magnitude of the equilibrium coefficients, we can determine which ligand binds most favorable to the quantum dot and compare this to the efficiency of charge extraction. This research has many applications, from solar energy to medical imaging, and the ability to extract charge outt of the quantum dot in an efficient and quick manner is one worth researching. This research is supported and overseen by Prof. Brandi Cossairt, and graduate students, Micaela Homer and Florence Dou. 

T cell activation is an essential element of the adaptive immune response. When the T-cell antigen receptor (TCR) interacts with a peptide antigen bound to a major histocompatibility complex (MHC), lymphocyte-specific protein tyrosine kinase (Lck) phosphorylates the CD3ζ chains of the receptor complex, initiating an intracellular signaling cascade. Therefore, Lck plays a crucial role in TCR signal transduction and thus Lck’s kinase activity and scaffolding function are tightly regulated by intramolecular protein-protein interactions (PPIs). Regulation is conferred by phosphorylation of two tyrosine residues on Lck, leading to conformational changes. When the C-terminal tail tyrosine is phosphorylated, the alignment of catalytically necessary residues is perturbed, rendering an inactive closed conformation. However, when the C-terminal tail is dephosphorylated, and the activation loop tyrosine is phosphorylated, Lck adopts a catalytically active open conformation. Using its regulatory domains, Lck also functions as a scaffold. First, Lck uses its tyrosine kinase domain to phosphorylate the kinase ZAP-70. Then, Lck bridges ZAP-70 to LAT by binding both proteins using its regulatory SH2 and SH3 domains. To study the intramolecular regulation of Lck’s kinase activity, we performed a Deep Mutational Scan (DMS) in yeast to measure the activity of ~5,000 single amino acid variants of Lck. Because Lck’s kinase activity is toxic to yeast, we can infer information about Lck’s kinase activity from changes in abundance of yeast expressing individual Lck variants, relative to wild type, over time. Specifically, we analyze the mutations along the SH2-SH3/kinase domain PPI which we hypothesize perturb the accessibility of Lck’s SH2/SH3 domains. In Lck variant expressing T cells, we expect to see differences in Lck’s binding partners due to differences in Lck’s SH2 and SH3 domain accessibility. This research will help us better understand Lck’s role in T-cell activation, with the long-term vision of leveraging Lck variants in next-generation T-cell therapies.

Proteins serve a pivotal role as building blocks for all living species in the biochemical world. An understanding of the function of a protein often requires first obtaining knowledge on the protein’s structure. Our research utilizes an innovative spectroscopic approach, double electron-electron resonance (DEER), to analyze protein structure. In the DEER experiment, we tag the analyte protein with magnetically active labels, the magnitude of magnetic interaction reflects the distance between the labeled sites. This technique has been utilized in determining the structure of many proteins that play important roles in society and public such as cell-membrane ion channels. A key component of using this method is rigorous data analysis and uncertainty quantification to achieve higher resolution and confidence in the structural conclusions made from the data. To conquer the constraints of traditional data fitting schemes that can usually succumb to bias and overconfidence, we apply the principles of Bayesian statistics through a Python based program, Dive, that utilizes Markov Chain Monte Carlo simulation to sample the data and analytical model. The program generates a fitted distribution of parameters to best characterize the data such that a good balance between uncertainty and bias can be achieved, which will help DEER practitioners in processing the data and making conclusions with respect to the critical functions of proteins.

The rapid rise of bioanalytical testing has renewed discussions in improving the collection of bacteria through oral sampling. Current diagnostic tests for common respiratory illnesses such as strep throat are invasive and uncomfortable, especially for children. Undiagnosed, these treatable diseases can cause serious damage. The Theberge lab has created the CandyCollect, a saliva sampling device intended to be a child-friendly alternative to current sampling techniques for home and clinical settings. The CandyCollect utilizes candy in the design to appeal to children, a unique component that requires additional testing and standardization to be appropriately implemented. This study explores the intersectionality of safety, design, effectiveness, and ease of application by using candy to increase children's compliance and act as a built-in timer. Our goal is to modify the existing device with these objectives in mind. Several types of candy were made using different ingredients, flavors, and textures, and the interaction of these candies with oral bacteria and saliva was investigated. We engineered the device using rapid prototyping, computer-numerical-control (CNC) milling, and silicone mold development. These methods allow for the flexibility to modify the design of the candy and sampling device. We then designed a survey for a human subjects study for adults (>18 years) to receive useability feedback and adapt the device components. Our analysis of these results determines the variety of candy that is most suitable for large scale use. The findings of this study will offer an improved method for child diagnostics and encourage other industries to redesign traditional sampling procedures.