126 results on '"Nancy Kelley-Loughnane"'
Search Results
2. The microbiota drives diurnal rhythms in tryptophan metabolism in the stressed gut
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Cassandra E. Gheorghe, Sarah-Jane Leigh, Gabriel S.S. Tofani, Thomaz F.S. Bastiaanssen, Joshua M. Lyte, Elisa Gardellin, Ashokkumar Govindan, Conall Strain, Sonia Martinez-Herrero, Michael S. Goodson, Nancy Kelley-Loughnane, John F. Cryan, and Gerard Clarke
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CP: Microbiology ,CP: Metabolism ,Biology (General) ,QH301-705.5 - Abstract
Summary: Chronic stress disrupts microbiota-gut-brain axis function and is associated with altered tryptophan metabolism, impaired gut barrier function, and disrupted diurnal rhythms. However, little is known about the effects of acute stress on the gut and how it is influenced by diurnal physiology. Here, we used germ-free and antibiotic-depleted mice to understand how microbiota-dependent oscillations in tryptophan metabolism would alter gut barrier function at baseline and in response to an acute stressor. Cecal metabolomics identified tryptophan metabolism as most responsive to a 15-min acute stressor, while shotgun metagenomics revealed that most bacterial species exhibiting rhythmicity metabolize tryptophan. Our findings highlight that the gastrointestinal response to acute stress is dependent on the time of day and the microbiome, with a signature of stress-induced functional alterations in the ileum and altered tryptophan metabolism in the colon.
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- 2024
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3. Iridescent biofilms of Cellulophaga lytica are tunable platforms for scalable, ordered materials
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Claretta J. Sullivan, Kennedy Brown, Chia-Suei Hung, Joseph Kuo-Hsiang Tang, Mark DeSimone, Vincent Chen, Pamela F. Lloyd, Maneesh Gupta, Abby Juhl, Wendy Crookes-Goodson, Milana Vasudev, Patrick B. Dennis, and Nancy Kelley-Loughnane
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Medicine ,Science - Abstract
Abstract Nature offers many examples of materials which exhibit exceptional properties due to hierarchical assembly of their constituents. In well-studied multi-cellular systems, such as the morpho butterfly, a visible indication of having ordered submicron features is given by the display of structural color. Detailed investigations of nature’s designs have yielded mechanistic insights and led to the development of biomimetic materials at laboratory scales. However, the manufacturing of hierarchical assemblies at industrial scales remains difficult. Biomanufacturing aims to leverage the autonomy of biological systems to produce materials at lower cost and with fewer carbon emissions. Earlier reports documented that some bacteria, particularly those with gliding motility, self-assemble into biofilms with polycrystalline structures and exhibit glittery, iridescent colors. The current study demonstrates the potential of using one of these bacteria, Cellulophaga lytica, as a platform for the large scale biomanufacturing of ordered materials. Specific approaches for controlling C. lytica biofilm optical, spatial and temporal properties are reported. Complementary microscopy-based studies reveal that biofilm color variations are attributed to changes in morphology induced by cellular responses to the local environment. Incorporation of C. lytica biofilms into materials is also demonstrated, thereby facilitating their handling and downstream processing, as would be needed during manufacturing processes. Finally, the utility of C. lytica as a self-printing, photonic ink is established by this study. In summary, autonomous surface assembly of C. lytica under ambient conditions and across multiple length scales circumvent challenges that currently hinder production of ordered materials in industrial settings.
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- 2023
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4. AlphaFold2 modeling and molecular dynamics simulations of an intrinsically disordered protein.
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Hao-Bo Guo, Baxter Huntington, Alexander Perminov, Kenya Smith, Nicholas Hastings, Patrick Dennis, Nancy Kelley-Loughnane, and Rajiv Berry
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Medicine ,Science - Abstract
We use AlphaFold2 (AF2) to model the monomer and dimer structures of an intrinsically disordered protein (IDP), Nvjp-1, assisted by molecular dynamics (MD) simulations. We observe relatively rigid dimeric structures of Nvjp-1 when compared with the monomer structures. We suggest that protein conformations from multiple AF2 models and those from MD trajectories exhibit a coherent trend: the conformations of an IDP are deviated from each other and the conformations of a well-folded protein are consistent with each other. We use a residue-residue interaction network (RIN) derived from the contact map which show that the residue-residue interactions in Nvjp-1 are mainly transient; however, those in a well-folded protein are mainly persistent. Despite the variation in 3D shapes, we show that the AF2 models of both disordered and ordered proteins exhibit highly consistent profiles of the pLDDT (predicted local distance difference test) scores. These results indicate a potential protocol to justify the IDPs based on multiple AF2 models and MD simulations.
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- 2024
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5. Accurate prediction by AlphaFold2 for ligand binding in a reductive dehalogenase and implications for PFAS (per- and polyfluoroalkyl substance) biodegradation
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Hao-Bo Guo, Vanessa A. Varaljay, Gary Kedziora, Kimberly Taylor, Sanaz Farajollahi, Nina Lombardo, Eric Harper, Chia Hung, Marie Gross, Alexander Perminov, Patrick Dennis, Nancy Kelley-Loughnane, and Rajiv Berry
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Medicine ,Science - Abstract
Abstract Despite the success of AlphaFold2 (AF2), it is unclear how AF2 models accommodate for ligand binding. Here, we start with a protein sequence from Acidimicrobiaceae TMED77 (T7RdhA) with potential for catalyzing the degradation of per- and polyfluoroalkyl substances (PFASs). AF2 models and experiments identified T7RdhA as a corrinoid iron-sulfur protein (CoFeSP) which uses a norpseudo-cobalamin (BVQ) cofactor and two Fe4S4 iron-sulfur clusters for catalysis. Docking and molecular dynamics simulations suggest that T7RdhA uses perfluorooctanoic acetate (PFOA) as a substrate, supporting the reported defluorination activity of its homolog, A6RdhA. We showed that AF2 provides processual (dynamic) predictions for the binding pockets of ligands (cofactors and/or substrates). Because the pLDDT scores provided by AF2 reflect the protein native states in complex with ligands as the evolutionary constraints, the Evoformer network of AF2 predicts protein structures and residue flexibility in complex with the ligands, i.e., in their native states. Therefore, an apo-protein predicted by AF2 is actually a holo-protein awaiting ligands.
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- 2023
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6. Molecular dynamics simulations explore effects of electric field orientations on spike proteins of SARS-CoV-2 virions
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Zhifeng Kuang, John Luginsland, Robert J. Thomas, Patrick B. Dennis, Nancy Kelley-Loughnane, William P. Roach, and Rajesh R. Naik
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Medicine ,Science - Abstract
Abstract Emergence of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and its current worldwide spread have caused a pandemic of acute respiratory disease COVID-19. The virus can result in mild to severe, and even to fatal respiratory illness in humans, threatening human health and public safety. The spike (S) protein on the surface of viral membrane is responsible for viral entry into host cells. The discovery of methods to inactivate the entry of SARS-CoV-2 through disruption of the S protein binding to its cognate receptor on the host cell is an active research area. To explore other prevention strategies against the quick spread of the virus and its mutants, non-equilibrium molecular dynamics simulations have been employed to explore the possibility of manipulating the structure–activity of the SARS-CoV-2 spike glycoprotein by applying electric fields (EFs) in both the protein axial directions and in the direction perpendicular to the protein axis. We have found out the application of EFs perpendicular to the protein axis is most effective in denaturing the HR2 domain which plays critical role in viral-host membrane fusion. This finding suggests that varying irradiation angles may be an important consideration in developing EF based non-invasive technologies to inactivate the virus.
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- 2022
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7. AlphaFold2 models indicate that protein sequence determines both structure and dynamics
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Hao-Bo Guo, Alexander Perminov, Selemon Bekele, Gary Kedziora, Sanaz Farajollahi, Vanessa Varaljay, Kevin Hinkle, Valeria Molinero, Konrad Meister, Chia Hung, Patrick Dennis, Nancy Kelley-Loughnane, and Rajiv Berry
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Medicine ,Science - Abstract
Abstract AlphaFold 2 (AF2) has placed Molecular Biology in a new era where we can visualize, analyze and interpret the structures and functions of all proteins solely from their primary sequences. We performed AF2 structure predictions for various protein systems, including globular proteins, a multi-domain protein, an intrinsically disordered protein (IDP), a randomized protein, two larger proteins (> 1000 AA), a heterodimer and a homodimer protein complex. Our results show that along with the three dimensional (3D) structures, AF2 also decodes protein sequences into residue flexibilities via both the predicted local distance difference test (pLDDT) scores of the models, and the predicted aligned error (PAE) maps. We show that PAE maps from AF2 are correlated with the distance variation (DV) matrices from molecular dynamics (MD) simulations, which reveals that the PAE maps can predict the dynamical nature of protein residues. Here, we introduce the AF2-scores, which are simply derived from pLDDT scores and are in the range of [0, 1]. We found that for most protein models, including large proteins and protein complexes, the AF2-scores are highly correlated with the root mean square fluctuations (RMSF) calculated from MD simulations. However, for an IDP and a randomized protein, the AF2-scores do not correlate with the RMSF from MD, especially for the IDP. Our results indicate that the protein structures predicted by AF2 also convey information of the residue flexibility, i.e., protein dynamics.
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- 2022
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8. Meeting report of the fourth annual Tri-Service Microbiome Consortium symposium
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Michael S. Goodson, Robyn A. Barbato, J. Philip Karl, Karl Indest, Nancy Kelley-Loughnane, Robert Kokoska, Camilla Mauzy, Kenneth Racicot, Vanessa Varaljay, and Jason Soares
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Microbiota ,Environmental microbiome ,Military ,Human performance ,Microbiome engineering ,Polymicrobial communities ,Environmental sciences ,GE1-350 ,Microbiology ,QR1-502 - Abstract
Abstract The Tri-Service Microbiome Consortium (TSMC) was founded to enhance collaboration, coordination, and communication of microbiome research among U.S. Department of Defense (DoD) organizations. The annual TSMC symposium is designed to enable information sharing between DoD scientists and leaders in the field of microbiome science, thereby keeping DoD consortium members informed of the latest advances within the microbiome community and facilitating the development of new collaborative research opportunities. The 2020 annual symposium was held virtually on 24–25 September 2020. Presentations and discussions centered on microbiome-related topics within four broad thematic areas: (1) Enabling Technologies; (2) Microbiome for Health and Performance; (3) Environmental Microbiome; and (4) Microbiome Analysis and Discovery. This report summarizes the presentations and outcomes of the 4th annual TSMC symposium.
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- 2021
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9. Meeting report of the third annual Tri-Service Microbiome Consortium symposium
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J. Philip Karl, Robyn A. Barbato, Laurel A. Doherty, Aarti Gautam, Sarah M. Glaven, Robert J. Kokoska, Dagmar Leary, Rebecca L. Mickol, Matthew A. Perisin, Andrew J. Hoisington, Edward J. Van Opstal, Vanessa Varaljay, Nancy Kelley-Loughnane, Camilla A. Mauzy, Michael S. Goodson, and Jason W. Soares
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Microbiota ,Environmental microbiome ,Military ,Human performance ,Microbiome engineering ,Polymicrobial communities ,Environmental sciences ,GE1-350 ,Microbiology ,QR1-502 - Abstract
Abstract The Tri-Service Microbiome Consortium (TSMC) was founded to enhance collaboration, coordination, and communication of microbiome research among U.S. Department of Defense (DoD) organizations and to facilitate resource, material and information sharing among consortium members. The 2019 annual symposium was held 22–24 October 2019 at Wright-Patterson Air Force Base in Dayton, OH. Presentations and discussions centered on microbiome-related topics within five broad thematic areas: 1) human microbiomes; 2) transitioning products into Warfighter solutions; 3) environmental microbiomes; 4) engineering microbiomes; and 5) microbiome simulation and characterization. Collectively, the symposium provided an update on the scope of current DoD microbiome research efforts, highlighted innovative research being done in academia and industry that can be leveraged by the DoD, and fostered collaborative opportunities. This report summarizes the presentations and outcomes of the 3rd annual TSMC symposium.
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- 2020
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10. A Pilot Study of the Effect of Deployment on the Gut Microbiome and Traveler’s Diarrhea Susceptibility
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Blake W. Stamps, Wanda J. Lyon, Adam P. Irvin, Nancy Kelley-Loughnane, and Michael S. Goodson
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gut microbiome ,16S rRNA gene ,diarrhea ,dysbiosis ,deployment ,Microbiology ,QR1-502 - Abstract
Traveler’s diarrhea (TD) is a recurrent and significant issue for many travelers including the military. While many known enteric pathogens exist that are causative agents of diarrhea, our gut microbiome may also play a role in TD susceptibility. To this end, we conducted a pilot study of the microbiome of warfighters prior to- and after deployment overseas to identify marker taxa relevant to TD. This initial study utilized full-length 16S rRNA gene sequencing to provide additional taxonomic resolution toward identifying predictive taxa.16S rRNA analyses of pre- and post-deployment fecal samples identified multiple marker taxa as significantly differentially abundant in subjects that reported diarrhea, including Weissella, Butyrivibrio, Corynebacterium, uncultivated Erysipelotrichaceae, Jeotgallibaca, unclassified Ktedonobacteriaceae, Leptolinea, and uncultivated Ruminiococcaceae. The ability to identify TD risk prior to travel will inform prevention and mitigation strategies to influence diarrhea susceptibility while traveling.
- Published
- 2020
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11. Epidemiology and associated microbiota changes in deployed military personnel at high risk of traveler's diarrhea.
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William A Walters, Faviola Reyes, Giselle M Soto, Nathanael D Reynolds, Jamie A Fraser, Ricardo Aviles, David R Tribble, Adam P Irvin, Nancy Kelley-Loughnane, Ramiro L Gutierrez, Mark S Riddle, Ruth E Ley, Michael S Goodson, and Mark P Simons
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Medicine ,Science - Abstract
Travelers' diarrhea (TD) is the most prevalent illness encountered by deployed military personnel and has a major impact on military operations, from reduced job performance to lost duty days. Frequently, the etiology of TD is unknown and, with underreporting of cases, it is difficult to accurately assess its impact. An increasing number of ailments include an altered or aberrant gut microbiome. To better understand the relationships between long-term deployments and TD, we studied military personnel during two nine-month deployment cycles in 2015-2016 to Honduras. To collect data on the prevalence of diarrhea and impact on duty, a total of 1173 personnel completed questionnaires at the end of their deployment. 56.7% reported reduced performance and 21.1% reported lost duty days. We conducted a passive surveillance study of all cases of diarrhea reporting to the medical unit with 152 total cases and a similar pattern of etiology. Enteroaggregative E. coli (EAEC, 52/152), enterotoxigenic E. coli (ETEC, 50/152), and enteropathogenic E. coli (EPEC, 35/152) were the most prevalent pathogens detected. An active longitudinal surveillance of 67 subjects also identified diarrheagenic E. coli as the primary etiology (7/16 EPEC, 7/16 EAEC, and 6/16 ETEC). Eleven subjects were recruited into a nested longitudinal substudy to examine gut microbiome changes associated with deployment. A 16S rRNA amplicon survey of fecal samples showed differentially abundant baseline taxa for subjects who contracted TD versus those who did not, as well as detection of taxa positively associated with self-reported gastrointestinal distress. Disrupted microbiota was also qualitatively observable for weeks preceding and following the incidents of TD. These findings illustrate the complex etiology of diarrhea amongst military personnel in deployed settings and its impacts on job performance. Potential factors of resistance or susceptibility can provide a foundation for future clinical trials to evaluate prevention and treatment strategies.
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- 2020
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12. Hybridization State Detection of DNA-Functionalized Gold Nanoparticles Using Hyperspectral Imaging
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Richard C. Murdock, Omar A. Khan, Thomas J. Lamkin, Saber M. Hussain, and Nancy Kelley-Loughnane
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Optics. Light ,QC350-467 - Abstract
Hyperspectral imaging has the unique ability of capturing spectral data for multiple wavelengths at each pixel in an image. This gives the ability to distinguish, with certainty, different nanomaterials and/or distinguish nanomaterials from biological materials. In this study, 4 nm and 13 nm gold nanoparticles (Au NPs) were synthesized, functionalized with complimentary oligonucleotides, and hybridized to form large networks of NPs. Scattering spectra were collected from each sample (unfunctionalized, functionalized, and hybridized) and evaluated. The spectra showed unique peaks for each size of Au NP sample and also exhibited narrowing and intensifying of the spectra as the NPs were functionalized and then subsequently hybridized. These spectra are different from normal aggregation effects where the LSPR and reflected spectrum broaden and are red-shifted. Rather, this appears to be dependent on the ability to control the interparticle distance through oligonucleotide length, which is also investigated through the incorporation of a poly-A spacer. Also, hybridized Au NPs were exposed to cells with no adverse effects and retained their unique spectral signatures. With the ability to distinguish between hybridization states at nearly individual NP levels, this could provide a new method of tracking the intracellular actions of nanomaterials as well as extracellular biosensing applications.
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- 2017
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13. Microarrays as Model Biosensor Platforms to Investigate the Structure and Affinity of Aptamers
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Jennifer A. Martin, Yaroslav Chushak, Jorge L. Chávez, Joshua A. Hagen, and Nancy Kelley-Loughnane
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Genetics ,QH426-470 ,Biochemistry ,QD415-436 - Abstract
Immobilization of nucleic acid aptamer recognition elements selected free in solution onto the surface of biosensor platforms has proven challenging. This study investigated the binding of multiple aptamer/target pairs immobilized on a commercially available microarray as a model system mimicking biosensor applications. The results indicate a minimum distance (linker length) from the surface and thymine nucleobase linker provides reproducible binding across varying conditions. An indirect labeling method, where the target was labeled with a biotin followed by a brief Cy3-streptavidin incubation, provided a higher signal-to-noise ratio and over two orders of magnitude improvement in limit of detection, compared to direct Cy3-protein labeling. We also showed that the affinities of the aptamer/target interaction can change between direct and indirect labeling and conditions to optimize for the highest fluorescence intensity will increase the sensitivity of the assay but will not change the overall affinity. Additionally, some sequences which did not initially bind demonstrated binding when conditions were optimized. These results, in combination with studies demonstrating enhanced binding in nonselection buffers, provided insights into the structure and affinity of aptamers critical for biosensor applications and allowed for generalizations in starting conditions for researchers wishing to investigate aptamers on a microarray surface.
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- 2016
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14. Biofunctionalized Zinc Oxide Field Effect Transistors for Selective Sensing of Riboflavin with Current Modulation
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Morley O. Stone, Kevin Leedy, Rajesh R. Naik, Nancy Kelley-Loughnane, Jorge L. Chávez, Burhan Bayraktaroglu, Sang N. Kim, and Joshua A. Hagen
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aptamer ,biomolecular detection ,ZnO-FET ,sensor ,riboflavin ,selectivity ,label-free ,biofunctionalization ,Chemical technology ,TP1-1185 - Abstract
Zinc oxide field effect transistors (ZnO-FET), covalently functionalized with single stranded DNA aptamers, provide a highly selective platform for label-free small molecule sensing. The nanostructured surface morphology of ZnO provides high sensitivity and room temperature deposition allows for a wide array of substrate types. Herein we demonstrate the selective detection of riboflavin down to the pM level in aqueous solution using the negative electrical current response of the ZnO-FET by covalently attaching a riboflavin binding aptamer to the surface. The response of the biofunctionalized ZnO-FET was tuned by attaching a redox tag (ferrocene) to the 3’ terminus of the aptamer, resulting in positive current modulation upon exposure to riboflavin down to pM levels.
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- 2011
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15. Single-Round Patterned DNA Library Microarray Aptamer Lead Identification
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Jennifer A. Martin, Peter A. Mirau, Yaroslav Chushak, Jorge L. Chávez, Rajesh R. Naik, Joshua A. Hagen, and Nancy Kelley-Loughnane
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Analytical chemistry ,QD71-142 - Abstract
A method for identifying an aptamer in a single round was developed using custom DNA microarrays containing computationally derived patterned libraries incorporating no information on the sequences of previously reported thrombin binding aptamers. The DNA library was specifically designed to increase the probability of binding by enhancing structural complexity in a sequence-space confined environment, much like generating lead compounds in a combinatorial drug screening library. The sequence demonstrating the highest fluorescence intensity upon target addition was confirmed to bind the target molecule thrombin with specificity by surface plasmon resonance, and a novel imino proton NMR/2D NOESY combination was used to screen the structure for G-quartet formation. We propose that the lack of G-quartet structure in microarray-derived aptamers may highlight differences in binding mechanisms between surface-immobilized and solution based strategies. This proof-of-principle study highlights the use of a computational driven methodology to create a DNA library rather than a SELEX based approach. This work is beneficial to the biosensor field where aptamers selected by solution based evolution have proven challenging to retain binding function when immobilized on a surface.
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- 2015
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16. Cross-Reactive Plasmonic Aptasensors for Controlled Substance Identification
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Joshua N. Yoho, Brian Geier, Claude C. Grigsby, Joshua A. Hagen, Jorge L. Chávez, and Nancy Kelley-Loughnane
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illicit drugs ,plasmonic sensor ,aptamer ,gold nanoparticle ,cross-reactive ,colorimetric ,Chemical technology ,TP1-1185 - Abstract
In this work, we developed an assay to determine if an arbitrary white powder is a controlled substance, given the plasmonic response of aptamer-gold nanoparticle conjugates (Apt-AuNPs). Toward this end, we designed Apt-AuNPs with specific a response to common controlled substances without cross reactivity to chemicals typically used as fillers in street formulations. Plasmonic sensor variation was shown to produce unique data fingerprints for each chemical analyzed, supporting the application of multivariate statistical techniques to annotate unknown samples by chemical similarity. Importantly, the assay takes less than fifteen minutes to run, and requires only a few micrograms of the material, making the proposed assay easily deployable in field operations.
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- 2017
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17. Artificial intelligence for synthetic biology.
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Mohammed Eslami, Aaron Adler, Rajmonda Sulo Caceres, Joshua G. Dunn, Nancy Kelley-Loughnane, Vanessa A. Varaljay, and Héctor García Martín
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- 2022
- Full Text
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18. Aerosol Jet Printed Surface-Enhanced Raman Substrates: Application for High-Sensitivity Detection of Perfluoroalkyl Substances
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Colleen McDonnell, Faris M. Albarghouthi, Ryan Selhorst, Nancy Kelley-Loughnane, Aaron D. Franklin, and Rahul Rao
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General Chemical Engineering ,General Chemistry - Published
- 2022
19. Effect of a Mixed Peptide Ligand Layer on Au Nanoparticles for Optical Control of Catalysis
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Maichong Xie, Joseph M. Slocik, Nancy Kelley-Loughnane, and Marc R. Knecht
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General Materials Science - Published
- 2022
20. System-level design of an RFID sweat electrolyte sensor patch.
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Daniel P. Rose, Michael E. Ratterman, Daniel K. Griffin, Linlin Hou, Nancy Kelley-Loughnane, Rajesh K. Naik, Joshua A. Hagen, Ian Papautsky, and Jason Heikenfeld
- Published
- 2014
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21. Biochemical and microbiological characterization of a thermotolerant bacterial consortium involved in the corrosion of Aluminum Alloy 7075
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Joaquin Atalah, Lotsé Blamey, Maximiliano J. Amenabar, Nancy Kelley-Loughnane, and Jenny M. Blamey
- Abstract
Microorganisms can play a significant role in material corrosion, with bacterial biofilms as major participants in microbially influenced corrosion (MIC). The exact mechanisms by which this takes place are poorly understood, resulting in a scarcity of information regarding MIC detection and prevention. In this work, a consortium of moderately thermophilic bacteria isolated from a biofilm growing over aluminum alloy 7075 was characterized. Its effect over the alloy was evaluated on a 40-day period using Electron Microscopy, demonstrating acceleration of corrosion in comparison to the abiotic control. The bacterial consortium was biochemically and microbiologically characterized as an as an attempt to elucidate factors contributing to corrosion. Molecular analysis revealed that the consortium consisted mainly of members of the Bacillusgenus, with lower abundance of other genera such as Thermoanaerobacterium, Anoxybacillus and Paenibacillus. The EPS polysaccharide presented mainly mannose, galactose, rhamnose and ribose. Our observations suggest that the acidification of the culture media resulting from bacterial metabolism acted as the main contributor to corrosion, hinting at an unspecific mechanism. The consortium was not sulfate-reducing, but it was found to produce hydrogen, which could also be a compounding factor for corrosion.
- Published
- 2022
22. Adhesive RFID Sensor Patch for Monitoring of Sweat Electrolytes.
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Daniel P. Rose, Michael E. Ratterman, Daniel K. Griffin, Linlin Hou, Nancy Kelley-Loughnane, Rajesh R. Naik, Joshua A. Hagen, Ian Papautsky, and Jason C. Heikenfeld
- Published
- 2015
- Full Text
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23. Accurate prediction by AlphaFold2 for ligand binding in a reductive dehalogenase: Implications for PFAS (per- and polyfluoroalkyl substance) biodegradation
- Author
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Hao-Bo Guo, Vanessa Varaljay, Gary Kedziora, Kimberly Taylor, Sanaz Farajollahi, Nina Lombardo, Eric Harper, Chia Hung, Marie Gross, Alexander Perminov, Patrick Dennis, Nancy Kelley-Loughnane, and Rajiv Berry
- Abstract
Despite the success of AlphaFold2 (AF2), it is unclear how AF2 models accommodate for ligand binding. Here, we start with a protein sequence from Acidimicrobiaceae TMED77 (T7RdhA) with potential for catalyzing the degradation of per- and polyfluoroalkyl substances (PFASs). AF2 models and experiments identified T7RdhA as a corrinoid iron-sulfur protein (CoFeSP) which uses a norpseudo-cobalamin (BVQ) cofactor and two [4Fe4S] iron-sulfur clusters (SF4) for catalysis. Docking and molecular dynamics simulations suggest that T7RdhA uses perfluorooctanoic acetate (PFOA) as a substrate, supporting the reported defluorination activity of its homolog, A6RdhA. We showed that AF2 provides processual (dynamic) predictions for the binding pockets of ligands (cofactors and/or substrates). Because the pLDDT scores provided by AF2 reflect the protein native states in complex with ligands as the evolutionary constraints, the Evoformer network of AF2 predicts protein structures and residue flexibility in complex with the ligands, i.e., in their native states.
- Published
- 2022
24. Draft Genome Sequence of the Nonmotile Tremellomycetes Yeast Naganishia albida, Isolated from Aircraft
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Dominique N. Wagner, Vanessa A. Varaljay, Wanda J. Lyon, Audra L. Crouch, Carrie A. Drake, Clayton Allex-Buckner, Justin C. Biffinger, Nancy Kelley-Loughnane, and Wendy J. Crookes-Goodson
- Subjects
Immunology and Microbiology (miscellaneous) ,Genetics ,Molecular Biology - Abstract
The Basidiomycota yeast Naganishia albida strain 5307AI was isolated from an aircraft polymer-coated surface. The genome size is 20,642,279 bp, with a G+C content of 53.99%. The genome contains fatty acid transporters, cutinases, hydroxylases, and lipases that are likely used for survival on polymer coatings on aircraft.
- Published
- 2022
25. Molecular Dynamics Investigation into pH Dependent Metal Binding of the Intrinsically Disordered Worm Jaw Protein, Nvjp-1
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Selemon Bekele, Kristi Singh, Evan Helton, Sanaz Farajollahi, Rajesh R. Naik, Patrick Dennis, Nancy Kelley-Loughnane, and Rajiv Berry
- Subjects
Intrinsically Disordered Proteins ,Ions ,Zinc ,Binding Sites ,Materials Chemistry ,Histidine ,Hydrogels ,Physical and Theoretical Chemistry ,Hydrogen-Ion Concentration ,Molecular Dynamics Simulation ,Surfaces, Coatings and Films ,Chelating Agents ,Protein Binding - Abstract
Sclerotization of the
- Published
- 2022
26. Characterization of synthetic riboswitch in cell-free protein expression systems
- Author
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Nancy Kelley-Loughnane, Kathryn Zimlich, Yaroslav Chushak, Bryan Alfred, Svetlana Harbaugh, and Jorge L. Chávez
- Subjects
Riboswitch ,0303 health sciences ,Messenger RNA ,Cell-Free System ,Green Fluorescent Proteins ,Free protein ,RNA ,Gene Expression Regulation, Bacterial ,Cell Biology ,Aptamers, Nucleotide ,Biology ,Cell biology ,03 medical and health sciences ,0302 clinical medicine ,Theophylline ,030220 oncology & carcinogenesis ,Escherichia coli ,Synthetic Biology ,sense organs ,Cell Engineering ,Molecular Biology ,Gene ,Research Paper ,030304 developmental biology - Abstract
Riboswitches are RNA-based regulatory elements that utilize ligand-induced structural changes in the 5ʹ-untranslated region of mRNA to regulate the expression of associated genes. The majority of synthetic riboswitches have been selected and tested in cell-based systems. Cell-free protein expression systems (CFPS) have several advantages for the development and testing of synthetic riboswitches, including eliminating interactions with complex cellular networks, and the decoupling of transcription and translation processes. To gain a better understanding of the riboswitch regulatory mechanism, to allow for more efficient riboswitch optimization and use for biosensing applications, we studied the performance of a theophylline-responsive synthetic riboswitch coupled with the superfolder green fluorescent protein (sfGFP) reporter gene in E. coli cellular extract and PURE cell-free systems. To monitor the mRNA dynamics, a malachite green aptamer sequence was added to the 3ʹ-untranslated region of sfGFP mRNA. Performance of the theophylline riboswitch was compared with a constitutively expressed sfGFP (control). Transcription dynamics of the riboswitch mRNA was very similar to the transcription of the control mRNA for all theophylline concentrations tested in both E. coli extract and PURE CFPS. However, sfGFP expression in the riboswitch construct was one order of magnitude lower, even at the highest concentration of theophylline. A mathematical model of riboswitch activation governed by the kinetic trapping mechanism was developed. Two factors – a reduced fraction of mRNA in the ‘ON’ state and a considerably lower translation initiation rate in the riboswitch – contribute to the much lower level of protein expression in the theophylline riboswitch compared to the control construct.
- Published
- 2021
27. Biofunctionalized Zinc Oxide Field Effect Transistors for Selective Sensing of Riboflavin with Current Modulation.
- Author
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Joshua A. Hagen, Sang N. Kim, Burhan Bayraktaroglu, Kevin D. Leedy, Jorge L. Chávez, Nancy Kelley-Loughnane, Rajesh R. Naik, and Morley O. Stone
- Published
- 2011
- Full Text
- View/download PDF
28. Meeting report of the third annual Tri-Service Microbiome Consortium symposium
- Author
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Nancy Kelley-Loughnane, Laurel A Doherty, Sarah M. Glaven, Camilla A Mauzy, Andrew J. Hoisington, Dagmar H. Leary, Robyn A. Barbato, R. L. Mickol, Robert J Kokoska, Michael S. Goodson, Aarti Gautam, J. Philip Karl, Edward J Van Opstal, Vanessa A. Varaljay, Matthew Perisin, and Jason W. Soares
- Subjects
Engineering ,Service (systems architecture) ,lcsh:QR1-502 ,Library science ,Meeting Report ,Applied Microbiology and Biotechnology ,Microbiology ,lcsh:Microbiology ,03 medical and health sciences ,Military ,Genetics ,Microbiome ,Synthetic biology ,lcsh:Environmental sciences ,030304 developmental biology ,Environmental microbiome ,lcsh:GE1-350 ,0303 health sciences ,Scope (project management) ,030306 microbiology ,business.industry ,Information sharing ,Microbiota ,Human performance ,Microbiome engineering ,Polymicrobial communities ,business ,Biotechnology - Abstract
The Tri-Service Microbiome Consortium (TSMC) was founded to enhance collaboration, coordination, and communication of microbiome research among U.S. Department of Defense (DoD) organizations and to facilitate resource, material and information sharing among consortium members. The 2019 annual symposium was held 22–24 October 2019 at Wright-Patterson Air Force Base in Dayton, OH. Presentations and discussions centered on microbiome-related topics within five broad thematic areas: 1) human microbiomes; 2) transitioning products into Warfighter solutions; 3) environmental microbiomes; 4) engineering microbiomes; and 5) microbiome simulation and characterization. Collectively, the symposium provided an update on the scope of current DoD microbiome research efforts, highlighted innovative research being done in academia and industry that can be leveraged by the DoD, and fostered collaborative opportunities. This report summarizes the presentations and outcomes of the 3rd annual TSMC symposium.
- Published
- 2020
29. Abstract 1168: Purification of Recombinant Histidine-Tagged Lanmodulin without Size Exclusion Chromatography and Metal Binding Affinities for Eu(III) or Fe(III)
- Author
-
Courtney Henthorn, Emily Koerner, Martha Carter, JD Gillum, Nancy Kelley-Loughnane, Chia-Suei Hung, and Justin Biffinger
- Subjects
Cell Biology ,Molecular Biology ,Biochemistry - Published
- 2023
30. Point-of-Use Detection of Environmental Fluoride via a Cell-Free Riboswitch-Based Biosensor
- Author
-
Adam D. Silverman, Matthew S. Verosloff, Walter Thavarajah, Michael C. Jewett, Nancy Kelley-Loughnane, and Julius B. Lucks
- Subjects
0106 biological sciences ,Riboswitch ,0303 health sciences ,Water source ,Biomedical Engineering ,Nanotechnology ,General Medicine ,Cell free ,Molecular systems ,01 natural sciences ,Biochemistry, Genetics and Molecular Biology (miscellaneous) ,03 medical and health sciences ,chemistry.chemical_compound ,chemistry ,13. Climate action ,010608 biotechnology ,Biosensor ,Fluoride ,030304 developmental biology ,Field conditions - Abstract
Advances in biosensor engineering have enabled the design of programmable molecular systems to detect a range of pathogens, nucleic acids, and chemicals. Here, we engineer and field-test a biosensor for fluoride, a major groundwater contaminant of global concern. The sensor consists of a cell-free system containing a DNA template that encodes a fluoride-responsive riboswitch regulating genes that produce a fluorescent or colorimetric output. Individual reactions can be lyophilized for long-term storage and detect fluoride at levels above 2 ppm, the Environmental Protection Agency's most stringent regulatory standard, in both laboratory and field conditions. Through onsite detection of fluoride in a real-world water source, this work provides a critical proof-of-principle for the future engineering of riboswitches and other biosensors to address challenges for global health and the environment.
- Published
- 2019
31. Organism Engineering for the Bioproduction of the Triaminotrinitrobenzene (TATB) Precursor Phloroglucinol (PG)
- Author
-
Nathan D. Schwalm, Adam D. Silverman, Cassandra Bristol, Maneesh K. Gupta, Julius P. Lucks, Carrie A. Drake, Wendy J. Crookes-Goodson, Michael C. Jewett, Sarah M. Glaven, Steven M Blum, Nancy Kelley-Loughnane, Alexander V. Tobias, Raissa Eluere, Elizabeth L. Onderko, Wais Mojadedi, Felix Moser, Christopher A. Voigt, Matthew P. Lux, Arturo Casini, R. L. Mickol, Katelin Pratt, Katherine L. Akingbade, Vanessa A. Varaljay, D. Benjamin Gordon, Adam J. Meyer, Shangtao Liu, and Ishtiaq Saaem
- Subjects
0106 biological sciences ,Phloroglucinol ,Biomedical Engineering ,Computational biology ,Biology ,01 natural sciences ,Biochemistry, Genetics and Molecular Biology (miscellaneous) ,Genome ,Metabolic engineering ,03 medical and health sciences ,chemistry.chemical_compound ,Synthetic biology ,010608 biotechnology ,Escherichia coli ,Genetic Testing ,Gene ,Nitrobenzenes ,Selection (genetic algorithm) ,Organism ,030304 developmental biology ,0303 health sciences ,General Medicine ,Bioproduction ,Metabolic Engineering ,chemistry - Abstract
Organism engineering requires the selection of an appropriate chassis, editing its genome, combining traits from different source species, and controlling genes with synthetic circuits. When a strain is needed for a new target objective, for example, to produce a chemical-of-need, the best strains, genes, techniques, software, and expertise may be distributed across laboratories. Here, we report a project where we were assigned phloroglucinol (PG) as a target, and then combined unique capabilities across the United States Army, Navy, and Air Force service laboratories with the shared goal of designing an organism to produce this molecule. In addition to the laboratory strain
- Published
- 2019
32. Erratum for Harris et al., 'Draft Genome Sequence of the Bacterium Delftia acidovorans Strain D4B, Isolated from Soil'
- Author
-
John C Sitko, J. Jordan Steel, Marie Gross, Patrick B. Dennis, Sanaz Farajollahi, Nancy Kelley-Loughnane, Jeremy Kemball, Vanessa A. Varaljay, Erin A. Almand, and Jackson Harris
- Subjects
Genetics ,Whole genome sequencing ,Immunology and Microbiology (miscellaneous) ,Delftia acidovorans ,Strain (chemistry) ,Genome Sequences ,Erratum ,Biology ,biology.organism_classification ,Molecular Biology ,Bacteria - Abstract
Delftia acidovorans strain D4B is an aerobic bacterium within the Betaproteobacteria lineage that was isolated from soil. The genome size is 6.26 Mbp, with a G+C content of 67%. The genome encodes enzymes potentially involved in the degradation of fluorinated compounds.
- Published
- 2021
33. Draft Genome Sequence of the Bacterium Delftia acidovorans Strain D4B, Isolated from Soil
- Author
-
Erin A. Almand, Vanessa A. Varaljay, Marie Gross, Jeremy Kemball, John C Sitko, Nancy Kelley-Loughnane, Sanaz Farajollahi, J. Jordan Steel, Jackson Harris, and Patrick B. Dennis
- Subjects
chemistry.chemical_classification ,Whole genome sequencing ,Delftia acidovorans ,biology ,Strain (chemistry) ,biology.organism_classification ,Genome ,Microbiology ,Enzyme ,Immunology and Microbiology (miscellaneous) ,chemistry ,Genetics ,Molecular Biology ,Genome size ,Betaproteobacteria ,Bacteria - Abstract
Delftia acidovorans strain D4B is an aerobic bacterium within the Betaproteobacteria lineage that was isolated from soil. The genome size is 6.26 Mbp, with a G+C content of 67%. The genome encodes enzymes potentially involved in the degradation of fluorinated compounds.
- Published
- 2021
34. Preparation of Multifunctional Silk-Based Microcapsules loaded with DNA Plasmids Encoding RNA Aptamers and Riboswitches
- Author
-
Svetlana Harbaugh, Nancy Kelley-Loughnane, Jorge L. Chávez, and Irina Drachuk
- Subjects
General Immunology and Microbiology ,Chemistry ,General Chemical Engineering ,General Neuroscience ,Aptamer ,Silk ,Fibroin ,RNA ,Metal Nanoparticles ,Capsules ,DNA ,Aptamers, Nucleotide ,General Biochemistry, Genetics and Molecular Biology ,chemistry.chemical_compound ,Plasmid ,Membrane ,Riboswitch ,Biophysics ,Gold ,Primer (molecular biology) ,Macromolecule ,Plasmids - Abstract
We introduce a protocol for the preparation of DNA-laden silk fibroin microcapsules via the Layer-by-Layer (LbL) assembly method on sacrificial spherical cores. Following adsorption of a prime layer and DNA plasmids, the formation of robust microcapsules was facilitated by inducing β-sheets in silk secondary structure during acute dehydration of a single silk layer. Hence, the layering occurred via multiple hydrogen bonding and hydrophobic interactions. Upon adsorption of multilayered shells, the core-shell structures can be further functionalized with gold nanoparticles (AuNPs) and/or antibodies (IgG) to be used for remote sensing and/or targeted delivery. Adjusting several key parameters during sequential deposition of key macromolecules on silica cores such as the presence of a polymer primer, the concentration of DNA and silk protein, as well as a number of adsorbed layers resulted in biocompatible, DNA-laden microcapsules with variable permeability and DNA loadings. Upon dissolution of silica cores, the protocol demonstrated the formation of hollow and robust microcapsules with DNA plasmids immobilized to the inner surface of the capsule membrane. Creating a selectively permeable biocompatible membrane between the DNA plasmids and the external environment preserved the DNA during long-term storage and played an important role in the improved output response from spatially confined plasmids. The activity of DNA templates and their accessibility were tested during in vitro transcription and translation reactions (cell-free systems). DNA plasmids encoding RNA light-up aptamers and riboswitches were successfully activated with corresponding analytes, as was visualized during localization of fluorescently labeled RNA transcripts or GFPa1 protein in the shell membranes.
- Published
- 2021
35. Riboswitches as Sensor Entities
- Author
-
Michael S. Goodson, Svetlana Harbaugh, Yaroslav Chushak, Jorge L. Chavez, and Nancy Kelley-Loughnane
- Subjects
Chemistry - Published
- 2021
36. Fast and Selective Plasmonic Serotonin Detection with Aptamer-Gold Nanoparticle Conjugates.
- Author
-
Jorge L. Chávez, Joshua A. Hagen, and Nancy Kelley-Loughnane
- Published
- 2017
- Full Text
- View/download PDF
37. In Operando Observation of Neuropeptide Capture and Release on Graphene Field-Effect Transistor Biosensors with Picomolar Sensitivity
- Author
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Cameron M. Crasto, Joshua A. Hagen, Rajesh R. Naik, Oksana M. Pavlyuk, Joseph M. Slocik, Nancy Kelley-Loughnane, Ming-Siao Hsiao, Lawrence F. Drummy, Yen Ngo, Steve S. Kim, Li Xing, Rhett L. Martineau, Ahmad E. Islam, Cheri M. Hampton, and Madhavi P. Kadakia
- Subjects
Analyte ,Materials science ,Transistors, Electronic ,Cryo-electron microscopy ,Nanotechnology ,Biosensing Techniques ,02 engineering and technology ,010402 general chemistry ,01 natural sciences ,law.invention ,Responsivity ,Peptide Library ,law ,Humans ,Molecule ,Neuropeptide Y ,General Materials Science ,Sweat ,Graphene ,Cryoelectron Microscopy ,Transistor ,021001 nanoscience & nanotechnology ,0104 chemical sciences ,Coupling (electronics) ,ROC Curve ,Area Under Curve ,Graphite ,0210 nano-technology ,Biosensor ,Protein Binding - Abstract
Transmission electron microscopy (TEM) is being pushed to new capabilities which enable studies on systems that were previously out of reach. Among recent innovations, TEM through liquid cells (LC-TEM) enables in operando observation of biological phenomena. This work applies LC-TEM to the study of biological components as they interact on an abiotic surface. Specifically, analytes or target molecules like neuropeptide Y (NPY) are observed in operando on functional graphene field-effect transistor (GFET) biosensors. Biological recognition elements (BREs) identified using biopanning with affinity to NPY are used to functionalize graphene to obtain selectivity. On working devices capable of achieving picomolar responsivity to neuropeptide Y, LC-TEM reveals translational motion, stochastic positional fluctuations due to constrained Brownian motion, and rotational dynamics of captured analyte. Coupling these observations with the electrical responses of the GFET biosensors in response to analyte capture and/or release will potentially enable new insights leading to more advanced and capable biosensor designs.
- Published
- 2019
38. Immobilization of Recombinant
- Author
-
Irina, Drachuk, Svetlana, Harbaugh, Ren, Geryak, David L, Kaplan, Vladimir V, Tsukruk, and Nancy, Kelley-Loughnane
- Abstract
Strategies for the encapsulation of cells for the design of cell-based sensors require efficient immobilization procedures while preserving biological activity of the reporter cells. Here, we introduce an immobilization technique that relies upon the symbiotic relationship between two bacterial strains: cellulose-producing
- Published
- 2021
39. Improving the Activity of DNA-Encoded Sensing Elements through Confinement in Silk Microcapsules
- Author
-
Irina Drachuk, Svetlana Harbaugh, Nancy Kelley-Loughnane, and Jorge L. Chávez
- Subjects
chemistry.chemical_classification ,Cell-free protein synthesis ,Materials science ,Artificial cell ,010405 organic chemistry ,Surface Properties ,Biomolecule ,Silk ,Fibroin ,Nanotechnology ,Capsules ,DNA ,010402 general chemistry ,Bombyx ,01 natural sciences ,0104 chemical sciences ,Membrane ,chemistry ,Colloidal gold ,Animals ,General Materials Science ,Semipermeable membrane ,Particle Size ,Fibroins ,Biosensor - Abstract
Assembling synthetic bioparts into simplified artificial cells holds tremendous promise for advancing studies into the synthesis, biosensing, and delivery of biomolecules. Currently, the most successful techniques for encapsulation of the transcription-translation machinery exploit compartmentalization in liposomal vesicles. However, improvements to these methods may increase permeability to polar molecules, functionalization of the membrane with biologically active elements, and encapsulation efficiency. Microcapsules prepared via templated layer-by-layer (LbL) assembly using natural polymers have the potential to resolve some of the hurdles associated with liposomes. Here, we introduce a design for immobilizing DNA templates encoding translationally activated riboswitches and RNA aptamers into microcapsules prepared from regenerated silk fibroin protein. Adjusting several key parameters such as the presence of a polymer primer, concentration of silk protein, and DNA loadings during LbL assembly resulted in biocompatible, semipermeable, DNA-laden microcapsules. To preserve bioactivity, DNA was immobilized inside of the capsule membrane, which not only promoted stability during long-term storage at ambient conditions but also improved output response from spatially confined DNA-encoded sensing elements (SEs). Multiple copies of mRNA and GFPa1 protein were synthesized upon activation with specific analytes during in vitro transcription/translation reactions, demonstrating that selective permeability of silk microcapsules was essential for the diffusion of components of the cell-free system inside of the capsules. Further functionalization of capsule shells with gold nanoparticles (AuNPs) and antibodies (IgG) demonstrated the applicability of microcompartmentalized colloidal objects carrying SEs for remote sensing and/or targeted delivery. In the future, multifunctional, biocompatible silk-based microcapsules loaded with different RNA sensors can help advance the design of multiplexed biosensors tracking multiple biomarkers in complex media.
- Published
- 2020
40. Gut‐brain axis serotonergic responses to acute stress exposure are microbiome‐dependent
- Author
-
Gerard Clarke, Cassandra E. Gheorghe, John F. Cryan, Nancy Kelley-Loughnane, Michael S. Goodson, Joshua M. Lyte, and Timothy G. Dinan
- Subjects
Male ,Restraint, Physical ,0301 basic medicine ,Serotonin ,medicine.medical_specialty ,Colon ,Physiology ,Gut–brain axis ,Gene Expression ,Ileum ,Disease ,Tryptophan Hydroxylase ,Biology ,Serotonergic ,Mice ,03 medical and health sciences ,Basal (phylogenetics) ,Sex Factors ,0302 clinical medicine ,Internal medicine ,Gene expression ,medicine ,Animals ,Germ-Free Life ,Indoleamine-Pyrrole 2,3,-Dioxygenase ,RNA, Messenger ,Microbiome ,Endocrine and Autonomic Systems ,Gastroenterology ,Hydroxyindoleacetic Acid ,Frontal Lobe ,Gastrointestinal Microbiome ,030104 developmental biology ,Endocrinology ,medicine.anatomical_structure ,Female ,Corticosterone ,Stress, Psychological ,030217 neurology & neurosurgery - Abstract
Background Understanding the mechanisms underpinning the response to acute stress is critical for determining how this can be modulated in both health and disease and across sexes. Stress can markedly alter the microbiome and gut-brain axis signaling with the serotonergic system being particularly sensitive to acute stress. As the impact of acute stress on regional serotonergic dynamics in the gut-brain axis and the contribution of the microbiome to this are poorly appreciated, we used microbiota-deficient mice to assess whether the serotonergic response to acute stress exposure is microbiome dependent. Methods Adult male and female conventional, germ-free, and colonized germ-free mice underwent a single acute stressor and samples were harvested immediately or 45 minutes following stress. Serotonin and related metabolites and serotonergic gene expression were determined. Key results Our data clearly show the microbiota influenced gastrointestinal serotonergic response to acute stress in a sex- and region-dependent manner. Male-specific poststress increases in colonic serotonin were absent in germ-free mice but normalized following colonization. mRNA serotonergic gene expression was differentially expressed in colon and ileum of germ-free mice on a sex-dependent basis. Within the frontal cortex, absence of the microbiome altered basal serotonin, its main metabolite 5-hydroxyindoleacetic acid, and prevented stress-induced increases in serotonin turnover. Conclusions and inferences The gut microbiome influences the set points of the brain and gastrointestinal serotonergic systems and affected their response to acute stress in a sex- and region-dependent manner.
- Published
- 2020
41. Point-of-Use Detection of Environmental Fluoride
- Author
-
Walter, Thavarajah, Adam D, Silverman, Matthew S, Verosloff, Nancy, Kelley-Loughnane, Michael C, Jewett, and Julius B, Lucks
- Subjects
Cell-Free System ,Transcription, Genetic ,Drinking Water ,Green Fluorescent Proteins ,Biosensing Techniques ,Templates, Genetic ,Global Health ,Article ,Fluorides ,Lakes ,Freeze Drying ,Swimming Pools ,Bacillus cereus ,Bacterial Proteins ,Genes, Reporter ,Riboswitch ,Water Pollutants, Chemical - Abstract
Advances in biosensor engineering have enabled the design of programmable molecular systems to detect a range of pathogens, nucleic acids, and chemicals. Here, we engineer and field-test a biosensor for fluoride, a major groundwater contaminant of global concern. The sensor consists of a cell-free system containing a DNA template that encodes a fluoride-responsive riboswitch regulating genes that produce a fluorescent or colorimetric output. Individual reactions can be lyophilized for long-term storage and detect fluoride at levels above 2 parts per million, the Environmental Protection Agency’s most stringent regulatory standard, in both laboratory and field conditions. Through onsite detection of fluoride in a real-world water source, this work provides a critical proof-of-principle for the future engineering of riboswitches and other biosensors to address challenges for global health and the environment.
- Published
- 2019
42. The current state and future direction of DoD gut microbiome research: a summary of the first DoD gut microbiome informational meeting
- Author
-
Keith W. Whitaker, C. A. P. T. Mark Riddle, J. Philip Karl, Nancy Kelley Loughnane, C. P. T. Blair C. R. Dancy, Kenneth Racicot, Frederick D. Gregory, Michael L. Goodson, Rasha Hammamieh, Robert Kokoska, Linda A. Chrisey, Steven Arcidiacono, and Jason W. Soares
- Subjects
0301 basic medicine ,Knowledge management ,lcsh:QH426-470 ,Performance ,Biology ,Meeting Report ,03 medical and health sciences ,Human health ,0302 clinical medicine ,Genetics ,Warfighter ,Disease ,Microbiome ,Knowledge dissemination ,Nutrition ,business.industry ,Stressors ,PTSD ,Gut microbiome ,lcsh:Genetics ,030104 developmental biology ,Health ,030211 gastroenterology & hepatology ,Nutrition research ,business ,Gut-brain axis ,Biomarkers - Abstract
The gut microbiome is increasingly recognized as integral to human health, and is emerging as a mediator of human physical and cognitive performance. This has led to the recognition that US Department of Defense (DoD) research supporting a healthy and resilient gut microbiome will be critical to optimizing the health and performance of future Warfighters. To facilitate knowledge dissemination and collaboration, identify resource capabilities and gaps, and maximize the positive impact of gut microbiome research on the Warfighter, DoD partners in microbiome research participated in a 2-day informational meeting co-hosted by the Natick Soldier Research, Engineering and Development Center (NSRDEC) and the US Army Research Institute of Environmental Medicine (USARIEM) on 16–17 November 2015. Attendee presentations and discussions demonstrated that multiple DoD organizations are actively advancing gut microbiome research. Common areas of research included the influence of military-relevant stressors on interactions between the microbiome and Warfighter biology, manipulation of the microbiome to influence Warfighter health, and use of the microbiome as a biomarker of Warfighter health status. Although resources and capabilities are available, they vary across laboratories and it was determined that centralizing certain DoD capabilities could accelerate progress. More significantly, the meeting created a foundation for a coordinated gut microbiome and nutrition research program aligning key DoD partners in the area of microbiome research. This report details the presentations and discussions presented during the 1st DoD Gut Microbiome Informational Meeting.
- Published
- 2018
43. Immobilization of Recombinant E. coli Cells in a Bacterial Cellulose–Silk Composite Matrix To Preserve Biological Function
- Author
-
Vladimir V. Tsukruk, Svetlana Harbaugh, Irina Drachuk, Ren Geryak, David L. Kaplan, and Nancy Kelley-Loughnane
- Subjects
0301 basic medicine ,Materials science ,Cell ,Biomedical Engineering ,Gluconacetobacter xylinus ,Fibroin ,02 engineering and technology ,021001 nanoscience & nanotechnology ,law.invention ,Biomaterials ,03 medical and health sciences ,chemistry.chemical_compound ,030104 developmental biology ,Membrane ,SILK ,medicine.anatomical_structure ,chemistry ,Biochemistry ,law ,Bacterial cellulose ,medicine ,Recombinant DNA ,Cellulose ,0210 nano-technology - Abstract
Strategies for the encapsulation of cells for the design of cell-based sensors require efficient immobilization procedures while preserving biological activity of the reporter cells. Here, we introduce an immobilization technique that relies upon the symbiotic relationship between two bacterial strains: cellulose-producing Gluconacetobacter xylinus cells; and recombinant Escherichia coli cells harboring recombinase-based dual-color synthetic riboswitch (RS), as a model for cell-based sensor. Following sequential coculturing of recombinant cells in the cellulose matrix, final immobilization of E. coli cells was completed after reconstituted silk fibroin (SF) protein was added to a “living membrane” generating the composite bacterial cellulose-silk fibroin (BC-SF) scaffold. By controlling incubation parameters for both types of cells, as well as the conformations in SF secondary structure, a variety of robust composite scaffolds were prepared ranging from opaque to transparent. The properties of the scaffol...
- Published
- 2017
44. Amplifying Riboswitch Signal Output Using Cellular Wiring
- Author
-
Annastacia C. Bennett, Michael S. Goodson, Svetlana Harbaugh, Emily Briskin, Brenton R. Jennewine, and Nancy Kelley-Loughnane
- Subjects
0301 basic medicine ,Riboswitch ,Biomedical Engineering ,Biosensing Techniques ,Biology ,Sensitivity and Specificity ,01 natural sciences ,Biochemistry, Genetics and Molecular Biology (miscellaneous) ,Signal ,03 medical and health sciences ,Synthetic biology ,Genes, Reporter ,Escherichia coli ,Signal processing ,010405 organic chemistry ,Reproducibility of Results ,Gene Expression Regulation, Bacterial ,General Medicine ,Molecular biology ,0104 chemical sciences ,Quorum sensing ,Spectrometry, Fluorescence ,030104 developmental biology ,Biophysics ,Sensitivity (electronics) ,Biosensor - Abstract
If fieldable riboswitch-based biological sensors are to fulfill their potential, it is necessary to increase their signal output. Here we report a novel modular amplification system using a riboswitch to initiate signaling between a sensing strain and a reporter strain of E. coli. A quorum sensing signaling molecule biologically wires the sensing and reporter strains together. The amplification circuit increased the amount of fluorescence generated on ligand binding compared to when the riboswitch controlled fluorescence expression directly. This had the corollary effect of increasing the sensitivity of the system, and allowed riboswitch-based reporting in E. coli strains that did not produce a detectable output when the riboswitch directly controlled reporter expression. The amplification circuit also reduced the time required to detect a signal output. The modularity of this amplification system coupled with the achievable increases in output can advance the development of riboswitches and biological sensors.
- Published
- 2017
45. Development of a point-of-care diagnostic for influenza detection with antiviral treatment effectiveness indication
- Author
-
Alison A. Weiss, Karen M. Gallegos, Nancy Kelley-Loughnane, Ian Papautsky, Richard C. Murdock, and Joshua A. Hagen
- Subjects
Paper ,Point-of-Care Systems ,Biomedical Engineering ,Neuraminidase ,Bioengineering ,02 engineering and technology ,Smartphone application ,Biology ,Bioinformatics ,Antiviral Agents ,01 natural sciences ,Biochemistry ,Article ,law.invention ,Antigen ,law ,Lab-On-A-Chip Devices ,Influenza, Human ,Humans ,Antiviral treatment ,Polymerase chain reaction ,Point of care ,010401 analytical chemistry ,General Chemistry ,021001 nanoscience & nanotechnology ,Virology ,0104 chemical sciences ,Treatment Outcome ,Infectious disease (medical specialty) ,Treatment decision making ,0210 nano-technology - Abstract
Currently, diagnosis of influenza is performed either through tedious polymerase chain reaction (PCR) or through rapid antigen detection assays. The rapid antigen detection assays available today are highly specific but not very sensitive, and most importantly, lack the ability to show if the strain of influenza detected is susceptible to antiviral agents, such as Tamiflu and Relenza. The ability to rapidly determine if a patient has an infectious disease and what type of treatment the infection will respond to, would significantly reduce the treatment decision time, shorten the impact of symptoms, and minimize transfer to others. In this study, a novel, point-of-care style μPAD (microfluidic paper-based diagnostic) for influenza has been developed with the ability to determine antiviral susceptibility of the strain for treatment decision. The assay exploits the enzymatic activity of surface proteins present on all influenza strains, and potential false positive responses can be mitigated. A sample can be added to the device, distributed to 4 different reagent zones, and development of the enzymatic substrate under different buffer conditions takes place on bottom of the device. Analysis can be performed by eye or through a colorimetric image analysis smartphone application.
- Published
- 2017
46. Aptamer-functionalized graphene-gold nanocomposites for label-free detection of dielectrophoretic-enriched neuropeptide Y
- Author
-
Nancy Kelley-Loughnane, Nathan S. Swami, Joshua A. Hagen, Renny Edwin Fernandez, Bankim J. Sanghavi, Vahid Farmehini, Jorge L. Chávez, and Chia-Fu Chou
- Subjects
Analyte ,Materials science ,Nanocomposite ,Graphene ,Aptamer ,010401 analytical chemistry ,Microfluidics ,Nanotechnology ,02 engineering and technology ,Cyclic olefin copolymer ,021001 nanoscience & nanotechnology ,Neuropeptide Y receptor ,01 natural sciences ,0104 chemical sciences ,Nanoimprint lithography ,law.invention ,lcsh:Chemistry ,chemistry.chemical_compound ,chemistry ,lcsh:Industrial electrochemistry ,lcsh:QD1-999 ,law ,Electrochemistry ,0210 nano-technology ,lcsh:TP250-261 - Abstract
We present an electrokinetically enhanced aptamer sensing platform on a disposable plastic chip for label-free detection of neuropeptide Y (NPY), which is a key neurological biomarker. The sensor consists of aptamer-functionalized graphene-gold nanocomposites (Gr-AuNs) patterned inside a nanoslit that is embossed on cyclic olefin copolymer via nanoimprint lithography. Analyte molecules are dielectrophoretically focused through the nanoslit onto aptamer-immobilized Gr-AuNs for rapid and selective electrochemical detection of NPY at picomolar levels. Keywords: Aptamer, Neuropeptide, Tyrosine, Graphene, Electrokinetics, Microfluidics
- Published
- 2016
47. The role of the microbiota in acute stress-induced myeloid immune cell trafficking
- Author
-
Timothy G. Dinan, Gerard Clarke, Marzia Sichetti, Michael S. Goodson, Marcus Boehme, Nancy Kelley-Loughnane, Marcel van de Wouw, John F. Cryan, Gerard M. Moloney, and Joshua M. Lyte
- Subjects
0301 basic medicine ,CCR2 ,Myeloid ,Immunology ,Spleen ,Granulocyte ,Monocytes ,03 medical and health sciences ,Behavioral Neuroscience ,Mice ,0302 clinical medicine ,Immune system ,Microbial composition ,Cell Movement ,Stress, Physiological ,medicine ,Mesenteric lymph nodes ,Animals ,Obesity ,Myeloid Progenitor Cells ,Stress-related disorders ,Innate immune system ,Host Microbial Interactions ,Endocrine and Autonomic Systems ,business.industry ,Microbiota ,Immunity, Innate ,Gastrointestinal Microbiome ,Mice, Inbred C57BL ,030104 developmental biology ,medicine.anatomical_structure ,Metagenome ,business ,030217 neurology & neurosurgery - Abstract
There has been a growing recognition of the involvement of the gastrointestinal microbiota in the development of stress-related disorders. Acute stress leads to activation of the neuroendocrine systems, which in turn orchestrate a large-scale redistribution of innate immune cells. Both these response systems are independently known to be primed by the microbiota, even though much is still unclear about the role of the gastrointestinal microbiota in acute stress-induced immune activation. In this study, we investigated whether the microbiota influences acute stress-induced changes in innate immunity using conventionally colonised mice, mice devoid of any microbiota (i.e. germ-free, GF), and colonised GF mice (CGF). We also explored the kinetics of stress-induced immune cell mobilisation in the blood, the spleen and mesenteric lymph nodes (MLNs). Mice were either euthanised prior to stress or underwent restraint stress and were then euthanised at various time points (i.e. 0, 45- and 240-minutes) post-stress. Plasma adrenaline and noradrenaline levels were analysed using ELISA and immune cell levels were quantified using flow cytometry. GF mice had increased baseline levels of adrenaline and noradrenaline, of which adrenaline was normalised in CGF mice. In tandem, GF mice had decreased circulating levels of LY6Chi and LY6Cmid, CCR2+ monocytes, and granulocytes, but not LY6C-, CX3CR1+ monocytes. These deficits were normalised in CGF mice. Acute stress decreased blood LY6Chi and LY6Cmid, CCR2+ monocytes while increasing granulocyte levels in all groups 45 minutes post-stress. However, only GF mice showed stress-induced changes in LY6Chi monocytes and granulocytes 240 minutes post-stress, indicating impairments in the recovery from acute stress-induced changes in levels of specific innate immune cell types. LY6C-, CX3CR1+ monocytes remained unaffected by stress, indicating that acute stress impacts systemic innate immunity in a cell-type-specific manner. Overall, these data reveal novel cell-type-specific changes in the innate immune system in response to acute stress, which in turn are impacted by the microbiota. In conclusion, the microbiota influences the priming and recovery of the innate immune system to an acute stressor and may inform future microbiota-targeted therapeutics aimed at modulating stress-induced immune activation in stress-related disorders.
- Published
- 2019
48. Point-of-Use Detection of Environmental Fluoride via a Cell-Free Riboswitch-Based Biosensor
- Author
-
Michael C. Jewett, Adam D. Silverman, Walter Thavarajah, Matthew S. Verosloff, Julius B. Lucks, and Nancy Kelley-Loughnane
- Subjects
Riboswitch ,0303 health sciences ,Chemistry ,Water source ,Nanotechnology ,Cell free ,Molecular systems ,010402 general chemistry ,01 natural sciences ,0104 chemical sciences ,03 medical and health sciences ,chemistry.chemical_compound ,13. Climate action ,Biosensor ,Fluoride ,030304 developmental biology ,Field conditions - Abstract
Advances in biosensor engineering have enabled the design of programmable molecular systems to detect a range of pathogens, nucleic acids, and chemicals. Here, we engineer and field-test a biosensor for fluoride, a major groundwater contaminant of global concern. The sensor consists of a cell-free system containing a DNA template that encodes a fluoride-responsive riboswitch regulating genes that produce a fluorescent or colorimetric output. Individual reactions can be lyophilized for long-term storage and detect fluoride at levels above 2 parts per million, the EPA’s most stringent regulatory standard, in both laboratory and field conditions. Through onsite detection of fluoride in a real-world water source, this work provides a critical proof-of-principle for the future engineering of riboswitches and other biosensors to address challenges for global health and the environment.
- Published
- 2019
- Full Text
- View/download PDF
49. Biosensor platforms for biomarker detection: plasmonic aptasensors for detection of Neuropeptide Y
- Author
-
Joshua A. Hagen, Jorge L. Chávez, Kelsey Rieger, and Nancy Kelley-Loughnane
- Subjects
Colloidal gold ,Chemistry ,Stress biomarkers ,Aptamer ,mental disorders ,Nanotechnology ,Neuropeptide Y receptor ,Biosensor ,humanities ,Plasmon ,Biomarker (cell) - Abstract
A Neuropeptide Y-binding aptamer (NPY-BA) and 15 nm gold nanoparticles (AuNPs) were used to create plasmonic aptasensors for the selective detection of Neuropeptide Y in a rapid colorimetric assay. In this report, we describe different parameters that can be varied to optimize the sensitivity and selectivity of these assay and demonstrate initial NPY detection in sweat.
- Published
- 2019
50. Influence of Silica Matrix Composition and Functional Component Additives on the Bioactivity and Viability of Encapsulated Living Cells
- Author
-
Darren R. Dunphy, Travis Savage, Svetlana Harbaugh, Jason C. Harper, Nancy Kelley-Loughnane, and C. Jeffrey Brinker
- Subjects
Riboswitch ,Aqueous solution ,Materials science ,Biomedical Engineering ,Sodium silicate ,Silicate ,Biomaterials ,chemistry.chemical_compound ,chemistry ,Chemical engineering ,Biochemistry ,Glycerol ,Viability assay ,Cell encapsulation ,Biosensor - Abstract
The remarkable impact encapsulation matrix chemistry can have on the bioactivity and viability of integrated living cells is reported. Two silica chemistries (aqueous silicate and alkoxysilane), and a functional component additive (glycerol), are employed to generate three distinct silica matrices. These matrices are used to encapsulate living E. coli cells engineered with a synthetic riboswitch for cell-based biosensing. Following encapsulation, membrane integrity, reproductive capability, and riboswitch-based protein expression levels and rates are measured over a 5 week period. Striking differences in E. coli bioactivity, viability, and biosensing performance are observed for cells encapsulated within the different matrices. E. coli cells encapsulated for 35 days in aqueous silicate-based (AqS) matrices showed relatively low membrane integrity, but high reproductive capability in comparison to cells encapsulated in glycerol containing sodium silicate-based (AqS + g) and alkoxysilane-based (PGS) gels. F...
- Published
- 2015
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