Biochemistry, Biophysics, and Structural Biology Commons^™

Structure Of Blood Coagulation Factor Viii In Complex With Anti-C2 Domain Inhibitory Antibody, Estelle K. Ronayne, Shaun C. Peters, Joseph Gish, Celena Wilson, H. Trent Spencer, Christopher B. Doering, Pete Lollar, P. Clint Spiegel Jr., Kenneth C. Childers

WWU Honors College Senior Projects

Factor VIII (fVIII) is a procoagulant protein that binds to activated factor IX (fIXa) on platelet surfaces to form the intrinsic tenase complex. Due to the high immunogenicity of fVIII, generation of antibody inhibitors is a common occurrence in patients during hemophilia A treatment and spontaneously occurs in acquired hemophilia A patients. Non-classical antibody inhibitors, which block fVIII activation by thrombin and formation of the tenase complex, are the most common anti-C2 domain pathogenic inhibitors in hemophilia A murine models and have been identified in patient plasmas. In this study, we report on the X-ray crystal structure of a B …

The Biosynthesis Of The Cannabinoids, M. Nazir Tahir, Fred Shahbazi, Simon Rondeau-Gagne, John F. Trant

Chemistry and Biochemistry Publications

Cannabis has been integral to Eurasian civilization for millennia, but a century of prohibition has limited investigation. With spreading legalization, science is pivoting to study the pharmacopeia of the cannabinoids, and a thorough understanding of their biosynthesis is required to engineer strains with specific cannabinoid profiles. This review surveys the biosynthesis and biochemistry of cannabinoids. The pathways and the enzymes’ mechanisms of action are discussed as is the non-enzymatic decarboxylation of the cannabinoic acids. There are still many gaps in our knowledge about the biosynthesis of the cannabinoids, especially for the minor components, and this review highlights the tools and …

Cown Sustains Nitrogenase Turnover In The Presence Of The Inhibitor Carbon Monoxide, Michael S. Medina, Kevin O. Bretzing, Richard A. Aviles, Kiersten M. Chong, Alejandro Espinoza, Chloe Nicole G. Garcia, Benjamin B. Katz, Ruchita N. Kharwa, Andrea Hernandez, Justin L. Lee, Terrence M. Lee, Christine Lo Verde, Max W. Strul, Emily Y. Wong, Cedric P. Owens

Biology, Chemistry, and Environmental Sciences Faculty Articles and Research

Nitrogenase is the only enzyme capable of catalyzing nitrogen fixation, the reduction of dinitrogen gas (N₂) to ammonia (NH₃). Nitrogenase is tightly inhibited by the environmental gas carbon monoxide (CO). Nitrogen-fixing bacteria rely on the protein CowN to grow in the presence of CO. However, the mechanism by which CowN operates is unknown. Here, we present the biochemical characterization of CowN and examine how CowN protects nitrogenase from CO. We determine that CowN interacts directly with nitrogenase and that CowN protection observes hyperbolic kinetics with respect to CowN concentration. At a CO concentration of 0.001 atm, …

Not-So-Innocent Anions Determine The Mechanism Of Cationic Alkylators, S. Maryamdokht Taimoory, Vincenzo Alessandro Cataldo, Andreas Schäfer, John F. Trant, Ryan Guterman

Chemistry and Biochemistry Publications

Alkylating reagents based on thioimidazolium ionic liquids were synthesized and the influence of the anion on the alkylation reaction mechanism explored in detail using both experimental and computational methods. Thioimidazolium cations transfer alkyl substituents to nucleophiles, however the reaction rate was highly dependent on anion identity, demonstrating that the anion is not innocent in the mechanism. Detailed analysis of the computationally-derived potential energy surfaces associated with possible mechanisms indicated that this dependence arises from a combination of anion induced electronic, steric and coordinating effects, with highly nucleophilic anions catalyzing a 2-step process while highly non-nucleophilic, delocalized anions favor a 1-step …

Methylation Of The D2 Dopamine Receptor Affects Binding With The Human Regulatory Proteins Par-4 And Calmodulin, Alexander Bowitch, Ansuman Sahoo, Andrea M. Clark, Christiana Ntangka, Krishna K. Raut, Paul Gollnick, Michael C. Yu, Steven M. Pascal, Sarah E. Walker, Denise M. Ferkey

Chemistry & Biochemistry Faculty Publications

No abstract provided.

Experimental And Computational Observations Of Immunogenic Cobalt Porphyrin Lipid Bilayers: Nanodomain-Enhanced Antigen Association., Jasmin Federizon, Conrard Giresse Tetsassi Feugmo, Wei-Chiao Huang, Xuedan He, Kazutoyo Miura, Aida Razi, Joaquin Ortega, Mikko Karttunen, Jonathan F Lovell

Chemistry Publications

Cobalt porphyrin phospholipid (CoPoP) can incorporate within bilayers to enable non-covalent surface-display of antigens on liposomes by mixing with proteins bearing a polyhistidine tag (his-tag); however, the mechanisms for how this occurs are poorly understood. These were investigated using the his-tagged model antigen Pfs25, a protein antigen candidate for malaria transmission-blocking vaccines. Pfs25 was found to associate with the small molecule aquocobalamin, a form of vitamin B12 and a cobalt-containing corrin macrocycle, but without particle formation, enabling comparative assessment. Relative to CoPoP liposomes, binding and serum stability studies indicated a weaker association of Pfs25 to aquocobalamin or cobalt nitrilotriacetic acid …

Modeling The Bidirectional Glutamine/ Ammonium Conversion Between Cancer Cells And Cancer-Associated Fibroblasts, Peter Hinow, Gabriella Pinter, Wei Yan, Shizhen Emily Wang

Mathematical Sciences Faculty Articles

Like in an ecosystem, cancer and other cells residing in the tumor microenvironment engage in various modes of interactions to buffer the negative effects of environmental changes. One such change is the consumption of common nutrients (such as glutamine/Gln) and the consequent accumulation of toxic metabolic byproducts (such as ammonium/NH⁴⁾. Ammonium is a waste product of cellular metabolism whose accumulation causes cell stress. In tumors, it is known that it can be recycled into nutrients by cancer associated fibroblasts (CAFs). Here we present monoculture and coculture growth of cancer cells and CAFs on different substrates: glutamine and ammonium. …

Time-Resolved Sans Reveals Pore-Forming Peptides Cause Rapid Lipid Reorganization, Michael H.L. Nguyen, Mitchell Dipasquale, Brett W. Rickeard, Caesar G. Yip, Kaity N. Greco, Elizabeth G. Kelley, Drew Marquardt

Chemistry and Biochemistry Publications

Cells depend on proper lipid transport and their precise distribution for vital cellular function. Disruption of such lipid organization can be initiated by external agents to cause cell death. Here, we investigate two antimicrobial pore-forming peptides, alamethicin and melittin, and their influence on lipid intervesicular exchange and transverse lipid diffusion (i.e. flip-flop) in model lipid vesicles. Small angle neutron scattering (SANS) and a strategic contrast matching scheme show the mixing of two isotopically distinct dimyristoylphosphocholine (DMPC) vesicle populations is promoted upon the addition of high (1/40) and low (1/150, 1/1000) peptide-to-lipid (P/L) molar ratios. Parsing out the individual exchange and …

Self-Assembled Thermoresponsive Nanogel From Grafted Hyaluronic Acid As A Biocompatible Delivery Platform For Curcumin With Enhanced Drug Loading And Biological Activities, Jittima Amie Luckanagul, Pahweenvaj Ratnatilaka Na Bhuket, Chawanphat Muangnoi, Pranee Rojsitthisak, Qian Wang, Pornchai Rojsitthisak

Faculty Publications

A hyaluronic acid-grafted poly(N-isopropylacrylamide) (HA-pNIPAM) was synthesized as a polymeric nanogel platform for encapsulation and delivery of hydrophobic bioactive compounds using curcumin as a model drug. As demonstrated by transmission electron microscopy and dynamic light scattering techniques, the HA-pNIPAM was simply assembled into spherical nano-sized particles with the thermoresponsive behavior. The success of curcumin aqueous solubilization was confirmed by fluorescent spectroscopy. The resulting nanogel formulation enhanced the aqueous solubility and uptake into NIH-3T3 cells of curcumin. This nanogel formulation also demonstrates cytocompatibility against NIH-3T3 cells, which deems it safe as a delivery vehicle. Moreover, the formulation has a slight skin-protection …

Evidence For The Role Of Cyp51a And Xenobiotic Detoxification In Differential Sensitivity To Azole Fungicides In Boxwood Blight Pathogens, Stefanos Stravoravdis, Robert E. Marra, Nicholas R. Leblanc, Joanne Crouch, Jonathan P. Hulvey

Microbiology Department Faculty Publication Series

Boxwood blight, a fungal disease of ornamental plants (Buxus spp.), is caused by two sister species, Calonectria pseudonaviculata (Cps) and C. henricotiae (Che). Compared to Cps, Che is documented to display reduced sensitivity to fungicides, including the azole class of antifungals, which block synthesis of a key fungal membrane component, ergosterol. A previous study reported an ergosterol biosynthesis gene in Cps, CYP51A, to be a pseudogene, and RNA-Seq data confirm that a functional CYP51A is expressed only in Che. The lack of additional ergosterol biosynthesis genes showing significant differential expression suggests that the functional CYP51A in Che could contribute to …

Polymeric Nanoparticles Active Against Dual-Species Bacterial Biofilms, Jessa Marie V. Makabenta, Jungmi Park, Cheng-Hsuan Li, Aritra Nath Chattopadhyay, Ahmed Nabawy, Ryan F. Landis, Akash Gupta, Suzannah Schmidt-Malan, Robin Patel, Vincent M. Rotello

Chemistry Department Faculty Publication Series

Biofilm infections are a global public health threat, necessitating new treatment strategies. Biofilm formation also contributes to the development and spread of multidrug-resistant (MDR) bacterial strains. Biofilm-associated chronic infections typically involve colonization by more than one bacterial species. The co-existence of multiple species of bacteria in biofilms exacerbates therapeutic challenges and can render traditional antibiotics ineffective. Polymeric nanoparticles offer alternative antimicrobial approaches to antibiotics, owing to their tunable physico-chemical properties. Here, we report the efficacy of poly(oxanorborneneimide) (PONI)-based antimicrobial polymeric nanoparticles (PNPs) against multi-species bacterial biofilms. PNPs showed good dual-species biofilm penetration profiles as confirmed by confocal laser scanning microscopy. …

A Prototype Compact Accelerator-Based Neutron Source (Cans) For Canada, Robert Laxdal, Dalini D. Maharaj, Mina Abbaslou, Zin Tun, Daniel Banks, Alexander Gottberg, Marco Marchetto, Eduardo Rodriguez, Zahra Yamani, Helmut Fritzsche, Ronald Rogge, Ming Pan, Oliver Kester, Drew Marquardt

Chemistry and Biochemistry Publications

Canada's access to neutron beams for neutron scattering was significantly curtailed in 2018 with the closure of the National Research Universal (NRU) reactor in Chalk River, Ontario, Canada. New sources are needed for the long-term; otherwise, access will only become harder as the global supply shrinks. Compact Accelerator-based Neutron Sources (CANS) offer the possibility of an intense source of neutrons with a capital cost significantly lower than spallation sources. In this paper, we propose a CANS for Canada. The proposal is staged with the first stage offering a medium neutron flux, linear accelerator-based approach for neutron scattering that is also …

Monitoring Stem Cell Differentiation Using Raman Microspectroscopy: Chondrogenic Differentiation, Towards Cartilage Formation, Francesca Ravera, Esen Efeoglu, Hugh Byrne

Articles

Mesenchymal Stem Cells (MSCs) have the ability to differentiate into chondrocytes, the only cellular components of cartilage and are therefore ideal candidates for cartilage and tissue repair technologies. Chondrocytes are surrounded by cartilage-like extracellular matrix (ECM), a complex network rich in glycosaminoglycans, proteoglycans, and collagen, which, together with a multitude of intracellular signalling molecules, trigger the chondrogenesis and allow the chondroprogenitor to acquire the spherical morphology of the chondrocytes. However, although the mechanisms of the differentiation of MSCs have been extensively explored, it has been difficult to provide a holistic picture of the process, in situ. Raman Micro Spectroscopy (RMS) …

Synthetic Pathways For Potential Platinum 1,10-Phenanthroline Compounds, Jacob Young

Mahurin Honors College Capstone Experience/Thesis Projects

There are extremely important uses for platinum when it comes to the treatment of cancers and dementias. One potential therapeutic combines platinum with a structure known as phenanthroline. This platinum phenanthroline structure is normally extremely water-insoluble, rendering it difficult to study and use in biological systems. The Williams’ Biochemistry Laboratory endeavored to find a simpler synthetic pathway for a water-soluble phenanthroline product. While unsuccessful in identifying a product as of yet, many synthetic pathways have been ruled out, and more hypothetical pathways are being formulated for testing.

Among the strategies analyzed are varying solvent, reagents, apparatus, methodology, and temperature in …

In Vitro Screening Of Tp5801 Against Kinases For Nature Communications Publication, Joshua Andersen

ScholarsArchive Data

Data from screen of TP-5801 against in vitro kinases

Improvement In 14-3-3 Binding Site Prediction, Katherine K. Mccormack

ScholarsArchive Data

The 14-3-3 family of phospho-binding proteins regulate a variety of major cellular processes through interaction with a network of dynamic proteins. Deregulation of the 14-3-3 interaction network contributes to a variety of degenerative disorders and cancers. Our lab focuses on identifying novel 14-3-3 interactions and understanding how 14-3-3 binding regulates protein function. A major gap in this process is that identifying the phospho-site where 14-3-3 docks on a given protein is time- and resource-consuming. Prediction algorithms have been developed to predict canonical 14-3-3 binding sites, however, there are many non-canonical sites that existing software is unable to predict. To fill …

Kinase Screen Data For Nature Communications Publication, Joshua Andersen

ScholarsArchive Data

A TNK1 peptide encompassing S502 was screened against 245 Ser/Thr kinases. This file contains kinase activity values (against S502) from an n=1 experiment. These data support a manuscript to be published in Nature Communications (Chan and Egbert et al.)

Scaling The Effects Of Ocean Acidification On Coral Growth And Coral-Coral Competition On Coral Community Recovery, Nicolas R. Evensen, Yves-Marie Bozec, Peter J. Edmunds, Peter J. Mumby

Biological Sciences Faculty Publications

Ocean acidification (OA) is negatively affecting calcification in a wide variety of marine organisms. These effects are acute for many tropical scleractinian corals under short-term experimental conditions, but it is unclear how these effects interact with ecological processes, such as competition for space, to impact coral communities over multiple years. This study sought to test the use of individual-based models (IBMs) as a tool to scale up the effects of OA recorded in short-term studies to community-scale impacts, combining data from field surveys and mesocosm experiments to parameterize an IBM of coral community recovery on the fore reef of Moorea, …

Detrital Neodymium And (Radio)Carbon As Complementary Sedimentary Bedfellows? The Western Arctic Ocean As A Testbed, Melissa S. Schwab, Jörg D. Rickli, Robie W. Macdonald, H. Rodger Harvey, Negar Haghipour, Timothy I. Eglinton

OES Faculty Publications

Interactions between organic and detrital mineral phases strongly influence both the dispersal and accumulation of terrestrial organic carbon (OC) in continental margin sediments. Yet the complex interplay among biological, chemical, and physical processes limits our understanding of how organo-mineral interactions evolve during sediment transfer and burial. In particular, diverse OC sources and complex hydrodynamic processes hinder the assessment of how the partnership of organic matter and its mineral host evolves during supply and dispersal over continental margins. In this study, we integrate new and compiled sedimentological (grain size, surface area), organic (%OC, OC-δ¹³C, OC-F¹⁴C), and inorganic …

Adenosine Triphosphate (Atp) As A Metric Of Microbial Biomass In Aquatic Systems: New Simplified Protocols, Laboratory Validation, And A Reflection On Data From The Literature, Alexander B. Bochdansky, Alison N. Stouffer, Nyjaee N. Washington

OES Faculty Publications

The use of adenosine triphosphate (ATP) as a universal biomass indicator is built on the premise that ATP concentration tracks biomass rather than the physiological condition of cells. However, reportedly high variability in ATP in response to environmental conditions is the main reason the method has not found widespread application. To test possible sources of this variability, we used the diatom Thalassiosira weissflogii as a model and manipulated its growth rate through nutrient limitation and through exposure to three different temperatures (15°C, 20°C, and 25°C). We simplified the ATP protocol with hot‐water or chemical extraction methods, modified a commercially available …

Marine Phytoplankton Functional Types Exhibit Diverse Responses To Thermal Change, S. I. Anderson, A. D. Barton, Sophie Clayton, S. Dutkiewicz, T. A. Rynearson

OES Faculty Publications

Marine phytoplankton generate half of global primary production, making them essential to ecosystem functioning and biogeochemical cycling. Though phytoplankton are phylogenetically diverse, studies rarely designate unique thermal traits to different taxa, resulting in coarse representations of phytoplankton thermal responses. Here we assessed phytoplankton functional responses to temperature using empirically derived thermal growth rates from four principal contributors to marine productivity: diatoms, dinoflagellates, cyanobacteria, and coccolithophores. Using modeled sea surface temperatures for 1950-1970 and 2080-2100, we explored potential alterations to each group's growth rates and geographical distribution under a future climate change scenario. Contrary to the commonly applied Eppley formulation, our …

Structual Analysis Of The Cl-Par-4 Tumor Suppressor As A Function Of Ionic Environment, Krishna K. Raut, Komala Ponniah, Steven M. Pascal

Chemistry & Biochemistry Faculty Publications

Prostate apoptosis response-4 (Par-4) is a proapoptotic tumor suppressor protein that has been linked to a large number of cancers. This 38 kilodalton (kDa) protein has been shown to be predominantly intrinsically disordered in vitro. In vivo, Par-4 is cleaved by caspase-3 at Asp-131 to generate the 25 kDa functionally active cleaved Par-4 protein (cl-Par-4) that inhibits NF-κB-mediated cell survival pathways and causes selective apoptosis in tumor cells. Here, we have employed circular dichroism (CD) spectroscopy and dynamic light scattering (DLS) to assess the effects of various monovalent and divalent salts upon the conformation of cl-Par-4 in vitro. We have …