Interlaboratory Measurement of Adeno-Associated Virus: Comparative Quantification of Full and Empty Capsids.

Recombinant adeno-associated virus (AAV) is one of the main viral vector-based gene therapy platforms. AAV is a virus consisting of a ≈25 nm diameter capsid with a ≈4.7 kb cargo capacity.

Variable gain DNA nanostructure charge amplifiers for biosensing.

Electronic measurements of engineered nanostructures comprised solely of DNA (DNA origami) enable new signal conditioning modalities for use in biosensing. DNA origami, designed to take on arbitrary shapes and allow programmable motion triggered by conjugated biomolecules, have sufficient mass and charge to generate a large electrochemical signal. Here, we demonstrate the ability to electrostatically control the DNA origami conformation, and thereby the resulting signal amplification, when the structure binds a nucleic acid analyte.

Macaque antibodies targeting Marburg virus glycoprotein induced by multivalent immunization.

Unlabelled: Marburg virus infection in humans is associated with case fatality rates that can reach up to 90%, but to date, there are no approved vaccines or monoclonal antibody (mAb) countermeasures. Here, we immunized Rhesus macaques with multivalent combinations of filovirus glycoprotein (GP) antigens belonging to Marburg, Sudan, and Ebola viruses to generate monospecific and cross-reactive antibody responses against them. From the animal that developed the highest titers of Marburg virus GP-specific neutralizing antibodies, we sorted single memory B cells using a heterologous Ravn virus GP probe and cloned and characterized a panel of 34 mAbs belonging to 28 unique lineages.

Glycoengineered recombinant alpha1-antitrypsin results in comparable and activities to human plasma-derived protein.

Alpha-1-antitrypsin (A1AT) is a multifunctional, clinically important, high value therapeutic glycoprotein that can be used for the treatment of many diseases such as alpha-1-antitrypsin deficiency, diabetes, graft-versus-host-disease, cystic fibrosis and various viral infections. Currently, the only FDA-approved treatment for A1AT disorders is intravenous augmentation therapy with human plasma-derived A1AT. In addition to its limited supply, this approach poses a risk of infection transmission, since it uses therapeutic A1AT harvested from donors.

Particle Metrology Approach to Understanding How Storage Conditions Affect Long-Term Liposome Stability.

Lipid nanoparticles are a generic type of nanomaterial with broad applicability in medicine as drug delivery vehicles. Liposomes are a subtype of lipid nanoparticles and, as a therapeutic platform, can be loaded with a genetic material or pharmaceutical agents for use as drug treatments. An open question for these types of lipid nanoparticles is what factor(s) affect the long-term stability of the particles.

A round-robin approach provides a detailed assessment of biomolecular small-angle scattering data reproducibility and yields consensus curves for benchmarking.

Through an expansive international effort that involved data collection on 12 small-angle X-ray scattering (SAXS) and four small-angle neutron scattering (SANS) instruments, 171 SAXS and 76 SANS measurements for five proteins (ribonuclease A, lysozyme, xylanase, urate oxidase and xylose isomerase) were acquired. From these data, the solvent-subtracted protein scattering profiles were shown to be reproducible, with the caveat that an additive constant adjustment was required to account for small errors in solvent subtraction. Further, the major features of the obtained consensus SAXS data over the q measurement range 0-1 Å are consistent with theoretical prediction.

Ionization and structural properties of mRNA lipid nanoparticles influence expression in intramuscular and intravascular administration.

Lipid Nanoparticles (LNPs) are used to deliver siRNA and COVID-19 mRNA vaccines. The main factor known to determine their delivery efficiency is the pKa of the LNP containing an ionizable lipid. Herein, we report a method that can predict the LNP pKa from the structure of the ionizable lipid.

Design of a native-like secreted form of the hepatitis C virus E1E2 heterodimer.

Hepatitis C virus (HCV) is a major worldwide health burden, and a preventive vaccine is needed for global control or eradication of this virus. A substantial hurdle to an effective HCV vaccine is the high variability of the virus, leading to immune escape. The E1E2 glycoprotein complex contains conserved epitopes and elicits neutralizing antibody responses, making it a primary target for HCV vaccine development.

Spectral-Based Screening Approach Evaluating Two Specific Maize Lines With Divergent Resistance to Invasion by Aflatoxigenic Fungi.

In an effort to control aflatoxin contamination in food and/or feed grains, a segment of research has focused on host resistance to eliminate aflatoxin from susceptible crops, including maize. To this end, screening tools are key to identifying resistant maize genotypes. The traditional field screening techniques, the kernel screening laboratory assay (KSA), and analytical methods (e.

Single-Molecule 3D Images of "Hole-Hole" IgG1 Homodimers by Individual-Particle Electron Tomography.

The engineering of immunoglobulin-G molecules (IgGs) is of wide interest for improving therapeutics, for example by modulating the activity or multiplexing the specificity of IgGs to recognize more than one antigen. Optimization of engineered IgG requires knowledge of three-dimensional (3D) structure of synthetic IgG. However, due to flexible nature of the molecules, their structural characterization is challenging.

Small-angle X-ray and neutron scattering demonstrates that cell-free expression produces properly formed disc-shaped nanolipoprotein particles.

Nanolipoprotein particles (NLPs), composed of membrane scaffold proteins and lipids, have been used to support membrane proteins in a native-like bilayer environment for biochemical and structural studies. Traditionally, these NLPs have been prepared by the controlled removal of detergent from a detergent-solubilized protein-lipid mixture. Recently, an alternative method has been developed using direct cell-free expression of the membrane scaffold protein in the presence of preformed lipid vesicles, which spontaneously produces NLPs without the need for detergent at any stage.

Temporal Effects on Internal Fluorescence Emissions Associated with Aflatoxin Contamination from Corn Kernel Cross-Sections Inoculated with Toxigenic and Atoxigenic .

Non-invasive, easy to use and cost-effective technology offers a valuable alternative for rapid detection of carcinogenic fungal metabolites, namely aflatoxins, in commodities. One relatively recent development in this area is the use of spectral technology. Fluorescence hyperspectral imaging, in particular, offers a potential rapid and non-invasive method for detecting the presence of aflatoxins in maize infected with the toxigenic fungus .

Isotopic labeling of proteins in Halobacterium salinarum.

It is often necessary to obtain isotopically labeled proteins containing (15)N, (13)C, or (2)H for nuclear magnetic resonance; and (2)H for small-angle neutron scattering or neutron diffraction studies. To achieve uniform isotopic labeling, protein expression is most commonly performed in Escherichia coli or yeast using labeled media. However, proteins from extreme halophiles sometimes require a cellular environment with high ionic strength and cannot be heterologously expressed in E.

Genome Sequence of Aspergillus flavus NRRL 3357, a Strain That Causes Aflatoxin Contamination of Food and Feed.

Aflatoxin contamination of food and livestock feed results in significant annual crop losses internationally. Aspergillus flavus is the major fungus responsible for this loss. Additionally, A.

Detergent-solubilized Patched purified from Sf9 cells fails to interact strongly with cognate Hedgehog or Ihog homologs.

Patched (Ptc) is a twelve-pass transmembrane protein that functions as a receptor for the Hedgehog (Hh) family of morphogens. In addition to Ptc, several accessory proteins including the CDO/Ihog family of co-receptors are necessary for proper Hh signaling. Structures of Hh proteins bound to members of the CDO/Ihog family are known, but the nature of the full Hh receptor complex is not well understood.

ZmGns, a maize class I β-1,3-glucanase, is induced by biotic stresses and possesses strong antimicrobial activity.

Plant β-1,3-glucanases are members of the pathogenesis-related protein 2 (PR-2) family, which is one of the 17 PR protein families and plays important roles in biotic and abiotic stress responses. One of the differentially expressed proteins (spot 842) identified in a recent proteomic comparison between five pairs of closely related maize (Zea mays L.) lines differing in aflatoxin resistance was further investigated in the present study.

Co-inoculation of aflatoxigenic and non-aflatoxigenic strains of Aspergillus flavus to study fungal invasion, colonization, and competition in maize kernels.

A currently utilized pre-harvest biocontrol method involves field inoculations with non-aflatoxigenic Aspergillus flavus strains, a tactic shown to strategically suppress native aflatoxin-producing strains and effectively decrease aflatoxin contamination in corn. The present in situ study focuses on tracking the invasion and colonization of an aflatoxigenic A. flavus strain (AF70), labeled with green fluorescent protein (GFP), in the presence of a non-aflatoxigenic A.

Fluorescence imaging spectroscopy (FIS) for comparing spectra from corn ears naturally and artificially infected with aflatoxin producing fungus.

In an effort to address the problem of rapid detection of aflatoxin in grain, particularly oilseeds, the current study assessed the spectral differences of aflatoxin production in kernels from a cornfield inoculated with spores from 2 different strains of toxigenic Aspergillus flavus. Aflatoxin production in corn from the same field due to natural infestation was also assessed. A small corn plot in Baton Rouge, La.

Breeding aflatoxin-resistant maize lines using recent advances in technologies - a review.

Aflatoxin contamination caused by Aspergillus flavus infection of corn is a significant and chronic threat to corn being used as food or feed. Contamination of crops at levels of 20 ng g(-1) or higher (as regulated by the USFDA) by this toxin and potent carcinogen makes the crop unsalable, resulting in a significant economic burden on the producer. This review focuses on elimination of this contamination in corn which is a major US crop and the basis of many products.

A single ligand is sufficient to activate EGFR dimers.

Crystal structures of human epidermal growth factor receptor (EGFR) with bound ligand revealed symmetric, doubly ligated receptor dimers thought to represent physiologically active states. Such complexes fail to rationalize negative cooperativity of epidermal growth factor (EGF) binding to EGFR and the behavior of the ligandless EGFR homolog ErbB2/HER2, however. We report cell-based assays that provide evidence for active, singly ligated dimers of human EGFR and its homolog, ErbB4/HER4.

Tight control of mycotoxin biosynthesis gene expression in Aspergillus flavus by temperature as revealed by RNA-Seq.

To better understand the effect of temperature on mycotoxin biosynthesis, RNA-Seq technology was used to profile the Aspergillus flavus transcriptome under different temperature conditions. This approach allowed us to quantify transcript abundance for over 80% of fungal genes including 1153 genes that were differentially expressed at 30 and 37 °C. Eleven of the 55 secondary metabolite clusters were upregulated at the lower temperature, including aflatoxin biosynthesis genes, which were among the most highly upexpressed genes.

Biomimetic syntheses and antiproliferative activities of racemic, natural (-), and unnnatural (+) glyceollin I.

A 14-step biomimetic synthetic route to glyceollin I (1.5% overall yield) was developed and deployed to produce the natural enantiomeric form in soy, its unnatural stereoisomer, and a racemic mixture. Enantiomeric excess was assessed by asymmetric NMR shift reagents and chiral HPLC.

Volatile profiles of toxigenic and non-toxigenic Aspergillus flavus using SPME for solid phase extraction.

Toxigenic and atoxigenic strains of Aspergillus flavus were grown on potato dextrose agar (PDA) and wetted (23% moisture) sterile, cracked corn for 14 and 21 days, respectively. Volatile compounds produced by A. flavus, as well as those present in the PDA controls and sterile cracked maize, were collected using solid-phase micro-extraction (SPME) and identified by gas chromatography/mass spectrometry.

Glyceollin I enantiomers distinctly regulate ER-mediated gene expression.

Glyceollins are pterocarpan phytoalexins elicited in high concentrations when soybeans are stressed. We have previously reported that the three glyceollin isomers (GLY I-III) exhibit antiestrogenic properties, which may have significant biological effects upon human exposure. Of the three isomers, we have recently shown that glyceollin I is the most potent antiestrogen.

PR10 expression in maize and its effect on host resistance against Aspergillus flavus infection and aflatoxin production.

Maize (Zea mays L.) is a major crop susceptible to Aspergillus flavus infection and subsequent contamination with aflatoxins, the potent carcinogenic secondary metabolites of the fungus. Protein profiles of maize genotypes resistant and susceptible to A.