Chemistry & Biochemistry Research Works

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3D DNA Crystals and Nanotechnology
(MDPI, 2016-08-18) Paukstelis, Paul J.; Seeman, Nadrian C.
DNA’s molecular recognition properties have made it one of the most widely used biomacromolecular construction materials. The programmed assembly of DNA oligonucleotides has been used to create complex 2D and 3D self-assembled architectures and to guide the assembly of other molecules. The origins of DNA nanotechnology are rooted in the goal of assembling DNA molecules into designed periodic arrays, i.e., crystals. Here, we highlight several DNA crystal structures, the progress made in designing DNA crystals, and look at the current prospects and future directions of DNA crystals in nanotechnology.
A Pro-Drug Approach for Selective Modulation of AI-2-Mediated Bacterial Cell-to-Cell Communication
(MDPI, 2012-03-21) Guo, Min; Gamby, Sonja; Nakayama, Shizuka; Smith, Jacqueline; Sintim, Herman O.
The universal quorum sensing autoinducer, AI-2, is utilized by several bacteria. Analogs of AI-2 have the potential to modulate bacterial behavior. Selectively quenching the communication of a few bacteria, in the presence of several others in an ecosystem, using analogs of AI-2 is non-trivial due to the ubiquity of AI-2 processing receptors in many bacteria that co-exist. Herein, we demonstrate that when an AI-2 analog, isobutyl DPD (which has been previously shown to be a quorum sensing, QS, quencher in both Escherichia coli and Salmonella typhimurium) is modified with ester groups, which get hydrolyzed once inside the bacterial cells, only QS in E. coli, but not in S. typhimurium, is inhibited. The origin of this differential QS inhibition could be due to differences in analog permeation of the bacterial membranes or ester hydrolysis rates. Such differences could be utilized to selectively target QS in specific bacteria amongst a consortium of other species that also use AI-2 signaling.
Acyclic Cucurbit[n]uril Bearing Alkyl Sulfate Ionic Groups - Electronic Supporting Data
(Beilstein Journal of Organic Chemistry, 2025-01-09) Akakpo, Christian; Zavalij, Peter Y.; Isaacs, Lyle
This dataset contains the electronic data files that support the publication.
Anthracene-Walled Acyclic CB[n] Receptors: in vitro and in vivo Binding Properties toward Drugs of Abuse
(Wiley, 2022-03-03) DiMaggio, Delaney; Brockett, Adam T.; Shuster, Michael; Murkli, Steven; Zhai, Canjia; King, David; O'Dowd, Brona; Cheng, Ming; Brady, Kimberly; Briken, Volker; Roesch, Matthew R.; Isaacs, Lyle
We report studies of the interaction of six acyclic CB[n]-type receptors toward a panel of drugs of abuse by a combination of isothermal titration calorimetry and 1H NMR spectroscopy. Anthracene walled acyclic CB[n] host (M3) displays highest binding affinity toward methamphetamine (Kd=15 nM) and fentanyl (Kd=4 nM). Host M3 is well tolerated by Hep G2 and HEK 293 cells up to 100 μM according to MTS metabolic and adenylate kinase release assays. An in vivo maximum tolerated dose study with Swiss Webster mice showed no adverse effects at the highest dose studied (44.7 mg kg−1). Host M3 is not mutagenic based on the Ames fluctuation test and does not inhibit the hERG ion channel. In vivo efficacy studies showed that pretreatment of mice with M3 significantly reduces the hyperlocomotion after treatment with methamphetamine, but M3 does not function similarly when administered 30 seconds after methamphetamine.
Book of Abstracts: 8th International Symposium on Macrocyclic and Supramolecular Chemistry
(2013-07-07) Davis, Jeffery; Flood, Amar; Isaacs, Lyle
This pdf file is the finalized version of the book of abstracts for the 8th International Symposium on Macrocyclic Chemistry held in Arlington, Virginia, USA from July 7-11, 2013
Branching activity switches actin network between connected and fragmented states in a myosin-dependent manner
(2021) Chandrasekaran, Aravind; Giniger, Edward; Papoian, Garegin
Actin networks rely on nucleation mechanisms to generate new filaments because de-novo nucleation is kinetically disfavored. Branching nucleation of actin filaments by Arp2/3, in particular, is critical for actin self-organization. In this study, we use the simulation platform for active matter, MEDYAN, to generate 2000s long stochastic trajectories of actin networks, under varying Arp2/3 concentrations, in reaction volumes of biologically meaningful size (> 20m3). We find that mechanosensitive dynamics of Arp2/3 increases the abundance of short filaments and increases network treadmilling rate. By analyzing the density-fields of F-actin, we find that at low Arp2/3 concentration, F-actin is organized into a single, connected and contractile domain, while at elevated Arp2/3 levels (10nM and above), such contractile actin domains fragment into smaller domains spanning a wide range of volumes. These fragmented domains are extremely dynamic, continuously merging and splitting, owing to the high treadmilling rate of the underlying actin network. Treating the domain dynamics as a drift-diffusion process, we find that the fragmented state is stochastically favored, and the network state slowly drifts towards the fragmented state with considerable diffusion (variability) in the number of domains. We suggest that tuning the Arp2/3 concentration enables cells to transition from a globally coherent cytoskeleton, whose response involves the entire cytoplasmic network, to a fragmented cytoskeleton where domains can respond independently to local varying signals.
Chemoenzymatic Synthesis and Antibody Binding of HIV-1 V1/V2 Glycopeptide-Bacteriophage Qβ Conjugates as a Vaccine Candidate
(MDPI, 2021-11-21) Zong, Guanghui; Toonstra, Christian; Yang, Qiang; Zhang, Roushu; Wang, Lai-Xi
The broadly neutralizing antibody PG9 recognizes a unique glycopeptide epitope in the V1V2 domain of HIV-1 gp120 envelope glycoprotein. The present study describes the design, synthesis, and antibody-binding analysis of HIV-1 V1V2 glycopeptide-Qβ conjugates as a mimic of the proposed neutralizing epitope of PG9. The glycopeptides were synthesized using a highly efficient chemoenzymatic method. The alkyne-tagged glycopeptides were then conjugated to the recombinant bacteriophage (Qβ), a virus-like nanoparticle, through a click reaction. Antibody-binding analysis indicated that the synthetic glycoconjugates showed significantly enhanced affinity for antibody PG9 compared with the monomeric glycopeptides. It was also shown that the affinity of the Qβ-conjugates for antibody PG9 was dependent on the density of the glycopeptide antigen display. The glycopeptide-Qβ conjugates synthesized represent a promising candidate of HIV-1 vaccine.
Comments regarding “Seismic damage analysis due to near-fault multipulse ground motion” by Guan Chen, Jiashu Yang, Ruohan Wang, Kaiqi Li, Yong Liu, Michael Beer; Earthquake Engineering & Structural Dynamics, 2023
(Wiley, 2023-11-27) Hariri-Ardebili, Mohammad Amin
This discussion is based on the paper by Chen et al. in 2023 (hereafter referred to as “the original paper/authors”). In their study, the original authors conducted a series of analyses using nonpulse, single-pulse, and multipulse ground motion records, evaluating their impact on a frame structure, a slope, and a gravity dam. Their key finding suggests that multipulse ground motion leads to more severe structural damage compared to nonpulse and single-pulse ground motions. However, it is important to note that the seismic damage analysis of the gravity dam in this paper does not adhere to state-of-the-practice recommendations. Consequently, drawing a definitive conclusion regarding the influence and importance of multipulse ground motion records on the seismic response of concrete dams requires further justification. This necessitates the incorporation of high-fidelity numerical models and probabilistic performance evaluation. We will discuss the significance of modeling assumptions, specifically addressing the dam–rock dynamic interaction in crack propagation and failure in dams.
Construction and Characterization of a Torsional Pendulum that Detects a Novel Form of Cranial Energy
(2009) Hansen, John Norman; Lieberman, Joshua A.
A torsional pendulum consisting of a dome-shaped energy collector and a nylon monofilament support fiber was suspended above the cranium of a seated human subject and the effects of the subject on the oscillations of the pendulum were measured. There were dramatic effects, with FFT analysis of the oscillation signal showing many new frequencies in addition to the natural frequency of 0.034 Hz. The lowest new frequencies (0.0-0.002 Hz) were accompanied by a shift in the Center of Oscillation (COO) of the pendulum, and the higher frequencies were associated with changes in the amplitude of oscillation. The Delta COO (7.3 deg) and the amplitude (12 deg) effects were substantial, and would require forces equivalent to 34 and 56 mg, respectively. Residual effects on the Delta COO and amplitudes persisted for at least 30 min after the subject departed, and the rate at which they subsided conformed to the kinetics of a chemical relaxation process with a relaxation time of 600 sec. Shifts in the magnitude of the Delta COO with the subject present also conformed to chemical relaxations processes, with relaxation times of 35 and 200 sec. It is proposed that the energy that drives the anomalous oscillations when the subject is present is the result of enzyme-mediated energy transductions that convert metabolic energy into a form of energy that can affect the pendulum. Although highly speculative, it is suggested that aspects of quantum entanglement are involved in the energy transduction process.
Cryptococcal Immune Reconstitution Inflammatory Syndrome: From Clinical Studies to Animal Experiments
(MDPI, 2022-12-07) Shi, Zoe W.; Chen, Yanli; Ogoke, Krystal M.; Strickland, Ashley B.; Shi, Meiqing
Cryptococcus neoformans is an encapsulated pathogenic fungus that initially infects the lung but can migrate to the central nervous system (CNS), resulting in meningoencephalitis. The organism causes the CNS infection primarily in immunocompromised individuals including HIV/AIDS patients, but also, rarely, in immunocompetent individuals. In HIV/AIDS patients, limited inflammation in the CNS, due to impaired cellular immunity, cannot efficiently clear a C. neoformans infection. Antiretroviral therapy (ART) can rapidly restore cellular immunity in HIV/AIDS patients. Paradoxically, ART induces an exaggerated inflammatory response, termed immune reconstitution inflammatory syndrome (IRIS), in some HIV/AIDS patients co-infected with C. neoformans. A similar excessive inflammation, referred to as post-infectious inflammatory response syndrome (PIIRS), is also frequently seen in previously healthy individuals suffering from cryptococcal meningoencephalitis. Cryptococcal IRIS and PIIRS are life-threatening complications that kill up to one-third of affected people. In this review, we summarize the inflammatory responses in the CNS during HIV-associated cryptococcal meningoencephalitis. We overview the current understanding of cryptococcal IRIS developed in HIV/AIDS patients and cryptococcal PIIRS occurring in HIV-uninfected individuals. We also describe currently available animal models that closely mimic aspects of cryptococcal IRIS observed in HIV/AIDS patients.
Cryptococcus neoformans Infection in the Central Nervous System: The Battle between Host and Pathogen
(MDPI, 2022-10-12) Chen, Yanli; Shi, Zoe W.; Strickland, Ashley B.; Shi, Meiqing
Cryptococcus neoformans (C. neoformans) is a pathogenic fungus with a global distribution. Humans become infected by inhaling the fungus from the environment, and the fungus initially colonizes the lungs. If the immune system fails to contain C. neoformans in the lungs, the fungus can disseminate to the blood and invade the central nervous system, resulting in fatal meningoencephalitis particularly in immunocompromised individuals including HIV/AIDS patients. Following brain invasion, C. neoformans will encounter host defenses involving resident as well as recruited immune cells in the brain. To overcome host defenses, C. neoformans possesses multiple virulence factors capable of modulating immune responses. The outcome of the interactions between the host and C. neoformans will determine the disease progression. In this review, we describe the current understanding of how C. neoformans migrates to the brain across the blood–brain barrier, and how the host immune system responds to the invading organism in the brain. We will also discuss the virulence factors that C. neoformans uses to modulate host immune responses.
A DNA G-quadruplex/i-motif hybrid
(Oxford U Press, 2019-12-16) Chu, Betty; Zhang, Daoning; Paukstelis, Paul J.
DNA can form many structures beyond the canonical Watson–Crick double helix. It is now clear that noncanonical structures are present in genomic DNA and have biological functions. G-rich G-quadruplexes and C-rich i-motifs are the most well-characterized noncanonical DNA motifs that have been detected in vivo with either proscribed or postulated biological roles. Because of their independent sequence requirements, these structures have largely been considered distinct types of quadruplexes. Here, we describe the crystal structure of the DNA oligonucleotide, d(CCAGGCTGCAA), that self-associates to form a quadruplex structure containing two central antiparallel G-tetrads and six i-motif C–C+ base pairs. Solution studies suggest a robust structural motif capable of assembling as a tetramer of individual strands or as a dimer when composed of tandem repeats. This hybrid structure highlights the growing structural diversity of DNA and suggests that biological systems may harbor many functionally important non-duplex structures.
Endo-S-c-di-GMP Analogues-Polymorphism and Binding Studies with Class I Riboswitch
(MDPI, 2012-11-09) Zhou, Jie; Sayre, David A.; Wang, Jingxin; Pahadi, Nirmal; Sintim, Herman O.
C-di-GMP, a cyclic guanine dinucleotide, has been shown to regulate biofilm formation as well as virulence gene expression in a variety of bacteria. Analogues of c-di-GMP have the potential to be used as chemical probes to study c-di-GMP signaling and could even become drug leads for the development of anti-biofilm compounds. Herein we report the synthesis and biophysical studies of a series of c-di-GMP analogues, which have both phosphate and sugar moieties simultaneously modified (called endo-S-c-di-GMP analogues). We used computational methods to predict the relative orientation of the guanine nucleobases in c-di-GMP and analogues. DOSY NMR of the endo-S-c-di-GMP series showed that the polymorphism of c-di-GMP can be tuned with conservative modifications to the phosphate and sugar moieties (conformational steering). Binding studies with Vc2 RNA (a class I c-di-GMP riboswitch) revealed that conservative modifications to the phosphate and 2'-positions of c-di-GMP dramatically affected binding to class I riboswitch.
Exploring the pH dependence of the SARS-CoV-2 complete fusion domain and the role of its unique structural features
(Wiley, 2022-08-11) Birtles, Daniel; Oh, Anna E.; Lee, Jinwoo
SARS-CoV-2 may enter target cells through the process of membrane fusion at either the plasma (~pH 7.4–7.0) or endosomal (~pH 6.5–5.0) membrane in order to deliver its genetic information. The fusion domain (FD) of the spike glycoprotein is responsible for initiating fusion and is thus integral to the viral life cycle. The FD of SARS-CoV-2 is unique in that it consists of two structurally distinctive regions referred to as the fusion peptide (FP) and the fusion loop (FL); yet the molecular mechanisms behind how this FD perturbs the membrane to initiate fusion remains unclear. In this study via solution NMR, we witnessed only a slight conformational change in the FD between pH 7.4 and pH 5.0, resulting in a minor elongation of helix 1. However, we found that the FD's ability to mediate membrane fusion has a large and significant pH dependence, with fusion events being more readily induced at low pH. Interestingly, a biphasic relationship between the environmental pH and fusogenicity was discovered, suggesting a preference for the FD to initiate fusion at the late endosomal membrane. Furthermore, the conserved disulfide bond and hydrophobic motif “LLF” were found to be critical for the function of the complete FD, with minimal activity witnessed when either was perturbed. In conclusion, these findings indicate that the SARS-CoV-2 FD preferably initiates fusion at a pH similar to the late endosome through a mechanism that heavily relies on the internal disulfide bond of the FL and hydrophobic LLF motif within the FP.
Extracellular vesicle proteomes reflect developmental phases of Bacillus subtilis
(Springer Nature, 2016-03-09) Kim, Yeji; Edwards, Nathan; Fenselau, Catherine
Extracellular vesicles (EV) are spherical membrane-bound vesicles with nano-scale diameters, which are shed to the extracellular region by most eukaryotic and prokaryotic cells. Bacterial EV are proposed to contribute to intercellular communication, bacterial survival and human pathogenesis as a novel secretion system. EV have been characterized from many Gram-negative species and, more recently, from several vegetative Gram-positive bacteria. Further characterization of EV and their molecular cargos will contribute to understanding bacterial physiology and to developing therapeutic approaches. Bacillus subtilis were observed to release EV to a similar extent during sporulation as during the vegetative growth phase. However, the two vesicular cargos show qualitatively and quantitatively different proteomes. Among 193 total proteins identified across both samples, 61 were shown to be significantly more abundant in EV shed by sporulating cells, with (log) ratio of spectral counts RSC > 1 and Fisher-exact test FDR < 5 %. Sixty-two proteins were found to be significantly more abundant in EV shed by vegetative cells. Membrane fusion was shown to take place between these EVs and Gram-positive cells. Biogenesis of EV is a continuous process over the entire life cycle of this sporulating bacterium. The formation of EV during sporulation is strongly supported by the delineation of protein content that differs from the proteome of EV formed by vegetative spores.
Flexibility and Control of Protein-DNA Loops
(World Scientific Publishing Company, 2006-10) Kahn, Jason D.; Cheong, Raymond; Edelman, Laurence M.; Mehta, Ruchi A.; Morgan, Michael A.
Protein-DNA loops are essential for efficient transcriptional repression and activation. The geometry and stability of the archetypal Lac repressor tetramer (LacI)-DNA loop were investigated using designed hyperstable loops containing lac operators bracketing a sequence-directed bend. Electrophoretic mobility shift assays, DNA cyclization, and bulk and single-molecule fluorescence resonance energy transfer (FRET) demonstrate that the DNA sequence controls whether the LacI-DNA loop forms a compact loop with positive writhe or an open loop with little writhe. Monte Carlo methods for simulation of DNA ring closure were extended to DNA loops, including treatment of variable protein hinge angles. The observed distribution of topoisomer products upon cyclization provides a strong constraint on possible models. The experiments and modeling imply that LacI-DNA can adopt a wide range of geometries but has a strong intrinsic preference for an open form. The flexibility of LacI helps explain in vivo observations that DNA looping is less sensitive to DNA length and shape than would be expected from the physical properties of DNA. While DNA cyclization suggests two pools of precursor loops for the 9C14 construct, single-molecule FRET demonstrates a single population. This discrepancy suggests that the LacI-DNA structure is strongly influenced by flanking DNA.
Griffithsin tandemers: flexible and potent lectin inhibitors of the human immunodeficiency virus
(Springer Nature, 2015-01-23) Moulaei, Tinoush; Alexandre, Kabamba B; Shenoy, Shilpa R; Meyerson, Joel R; Krumpe, Lauren RH; Constantine, Brian; Wilson, Jennifer; Buckheit, Robert W Jr.; McMahon, James B; Subramaniam, Sriram; Wlodawer, Alexander; O’Keefe, Barry R
The lectin griffithsin (GRFT) is a potent antiviral agent capable of prevention and treatment of infections caused by a number of enveloped viruses and is currently under development as an anti-HIV microbicide. In addition to its broad antiviral activity, GRFT is stable at high temperature and at a broad pH range, displays little toxicity and immunogenicity, and is amenable to large-scale manufacturing. Native GRFT is a domain-swapped homodimer that binds to viral envelope glycoproteins and has displayed mid-picomolar activity in cell-based anti-HIV assays. Previously, we have engineered and analyzed several monomeric forms of this lectin (mGRFT) with anti-HIV EC50 values ranging up to 323 nM. Based on our previous analysis of mGRFT, we hypothesized that the orientation and spacing of the carbohydrate binding domains GRFT were key to its antiviral activity. Here we present data on engineered tandem repeats of mGRFT (mGRFT tandemers) with antiviral activity at concentrations as low as one picomolar in whole-cell anti-HIV assays. mGRFT tandemers were analyzed thermodynamically, both individually and in complex with HIV-1 gp120. We also demonstrate by dynamic light scattering and cryo-electron microscopy that mGRFT tandemers do not aggregate HIV virions. This establishes that, although the intra-virion crosslinking of HIV envelope glycoproteins is likely integral to their activity, the antiviral activity of these lectins is not due to virus aggregation caused by inter-virion crosslinking. The engineered tandemer constructs of mGRFT may provide novel and powerful agents for prevention of infection by HIV and other enveloped viruses.
Histone post-translational modifications in frontal cortex from human donors with Alzheimer’s disease
(Springer Nature, 2015-10-01) Anderson, Kyle W.; Turko, Illarion V.
Alzheimer’s disease (AD) is the sixth leading cause of death and the most costly disease in the US. Despite the enormous impact of AD, there are no treatments that delay onset or stop disease progression currently on the market. This is partly due to the complexity of the disease and the largely unknown pathogenesis of sporadic AD, which accounts for the vast majority of cases. Epigenetics has been implicated as a critical component to AD pathology and a potential “hot spot” for treatments. Histone post-translational modifications (PTMs) are a key element in epigenetic regulation of gene expression and are known to be associated with the pathology of numerous diseases. Investigation of histone PTMs can help elucidate AD pathology and identify targets for therapies. A multiple reaction monitoring mass spectrometry assay was used to measure changes in abundance of several histone PTMs in frontal cortex from human donors affected with AD (n = 6) and age-matched, normal donors (n = 6). Of the changes observed, notable decreases in methylation of H2B residue K108 by 25 % and H4 residue R55 by 35 % were measured and are likely associated with hydrogen bonding networks important for nucleosome stability. Additionally, a 91 % increase in ubiquitination of K120 on H2B was measured as well as an apparent loss in acetylation of the region near the N-terminus of H4. Our method of quantification was also determined to be precise and robust, signifying measured changes were representative of true biological differences between donors and sample groups. We are the first to report changes in methylation of H2B K108, methylation of H4 R55, and ubiquitination of H2B K120 in frontal cortex from human donors with AD. These notable PTM changes may be of great importance in elucidating the epigenetic mechanism of AD as it relates to disease pathology. Beyond the structural and functional impacts of the changes we have measured, the sites of altered PTMs may be used to identify enzymes responsible for their modulation, which could be used as prospective drug targets for highly specific AD therapies.
HUMAN SUBJECT EFFECTS ON TORSION PENDULUM OSCILLATIONS: IMPORTANCE OF ESTABLISHING THE CONTRIBUTION OF THERMAL CONVECTION AIR CURRENTS
(Elsevier, 2017-02) Hansen, John Norman
Studies of thermal effects on torsion pendulum oscillations.
Implementation of Glycan Remodeling to Plant-Made Therapeutic Antibodies
(MDPI, 2018-01-31) Bennett, Lindsay D.; Yang, Qiang; Berquist, Brian R.; Giddens, John P.; Ren, Zhongjie; Kommineni, Vally; Murray, Ryan P.; White, Earl L.; Holtz, Barry R.; Wang, Lai-Xi; Marcel, Sylvain
N-glycosylation profoundly affects the biological stability and function of therapeutic proteins, which explains the recent interest in glycoengineering technologies as methods to develop biobetter therapeutics. In current manufacturing processes, N-glycosylation is host-specific and remains difficult to control in a production environment that changes with scale and production batches leading to glycosylation heterogeneity and inconsistency. On the other hand, in vitro chemoenzymatic glycan remodeling has been successful in producing homogeneous pre-defined protein glycoforms, but needs to be combined with a cost-effective and scalable production method. An efficient chemoenzymatic glycan remodeling technology using a plant expression system that combines in vivo deglycosylation with an in vitro chemoenzymatic glycosylation is described. Using the monoclonal antibody rituximab as a model therapeutic protein, a uniform Gal2GlcNAc2Man3GlcNAc2 (A2G2) glycoform without α-1,6-fucose, plant-specific α-1,3-fucose or β-1,2-xylose residues was produced. When compared with the innovator product Rituxan®, the plant-made remodeled afucosylated antibody showed similar binding affinity to the CD20 antigen but significantly enhanced cell cytotoxicity in vitro. Using a scalable plant expression system and reducing the in vitro deglycosylation burden creates the potential to eliminate glycan heterogeneity and provide affordable customization of therapeutics’ glycosylation for maximal and targeted biological activity. This feature can reduce cost and provide an affordable platform to manufacture biobetter antibodies.