Label-free quantitative proteomic analysis of the inhibitory activities of juglone against translation and energy metabolism in Escherichia coli

Plant-derived antimicrobial agents have received increasing attention owing to their potential to control pathogens and excellent efficacy despite the growing prevalence of antibiotic resistance. However, the antibacterial mechanism of juglone, a traditional medicine used to cure skin infections, is still unclear. Therefore, in this study, in order to elucidate the mechanisms underlying the antibacterial activity of juglone, label-free quantitative proteomic technology was applied for analysis of the 417 proteins that were differentially expressed in Escherichia coli after treatment with juglone at one-half of the minimum inhibitory concentration. Gene ontology enrichment analysis of differentially expressed proteins suggested that juglone effectively repressed the expression of dehydrogenase and cytochrome oxidase, indicating that energy generation was blocked. Additionally, juglone induced RNA formation and ribosome assembly, resulting in inhibition of translation. This is the first study to adopt a proteomic approach to investigate the antibacterial mechanism of action of juglone against E. coli.     

Immunomic Identification of Malaria Antigens Associated With Protection in Mice

1. Download : Download high-res image (84KB)
2. Download : Download full-size image

Highlights

• •Production of sera with different levels of protection against rodent Plasmodium.
• •Generation of immunomic and proteomic data sets enriched in protective antigens.
• •Prediction of the most likely protective antigens using a weighted scoring system.

     

Spatiotemporal Changes of the Phagosomal Proteome in Dendritic Cells in Response to LPS Stimulation*

1. Download : Download high-res image (115KB)
2. Download : Download full-size image

Highlights

• •Characterization of the phagosomal proteome comparing resting and LPS-treated BMDCs.
• •Label-free quantification determined 2843 phagosomal proteins.
• •Reduced recruitment of hydrolases and V-ATPase to phagosomes of LPS-treated cells.
• •Increased recruitment of antigen cross-presentation molecules to these phagosomes.

     

Insulin Induces Microtubule Stabilization and Regulates the Microtubule Plus-end Tracking Protein Network in Adipocytes

1. Download : Download high-res image (182KB)
2. Download : Download full-size image

Highlights

• •Insulin Affects the Phosphorylation of G2L1, MARK2, CLIP2, EB1, AGAP3, and CKAP5.
• •Insulin Increases CLASP2 +TIP Density and Decreases CLASP2 +TIP Velocity.
• •Insulin Stimulates CLASP2 and G2L1 Trailing Along Microtubules.
• •Insulin Stimulates α-Tubulin Acetylation at Lysine 40 and Microtubule Stabilization.

     

Redundancy and Complementarity between ERAP1 and ERAP2 Revealed by their Effects on the Behcet's Disease-associated HLA-B*51 Peptidome,

1. Download : Download high-res image (120KB)
2. Download : Download full-size image

Highlights

• •HLA-B*51 and ERAP1, but not ERAP2, are risk factors for Behçet's disease.
• •The HLA-B*51 peptidome and the effects of ERAP1 and ERAP2 on it are analyzed.
• •ERAP1 and ERAP2 alter multiple features of the HLA-B*51 peptidome in distinct ways.
• •Both enzymes act independently with complementary and partially redundant functions.

     

Characterising biomolecular interactions and dynamics with mass photometry

We review recent advances in our ability to characterise biomolecular structure, interactions and associated dynamics by mass photometry (MP), the label-free detection and mass measurement of individual biomolecules in solution. Molecular counting and identification provides direct access to relative abundance, and thereby affinities, while associated dynamics yield on- and off-rates. The molecular resolution afforded by MP enables these measurements as a function of stoichiometry and assembly at equilibrium, as opposed to the majority of existing solution-based methods. Together with future improvements in terms of assays and technological performance, MP is likely to provide mechanistic details of complex biomolecular processes.     

Label-free and amplified quantitation of proteins in complex mixtures using diffractive optics technology

Immunoaffinity approaches remain invaluable tools for characterization and quantitation of biopolymers. Their application in separation science is often limited due to the challenges of immunoassay development. Typical end-point immunoassays require time consuming and labor-intensive approaches for optimization. Real-time label-free analysis using diffractive optics technology (dot^®) helps guide a very effective iterative process for rapid immunoassay development. Both label-free and amplified approaches can be used throughout feasibility testing and ultimately in the final assay, providing a robust platform for biopolymer analysis over a very broad dynamic range. We demonstrate the use of dot in rapidly developing assays for quantitating (1) human IgG in complex media, (2) a fusion protein in production media and (3) protein A contamination in purified immunoglobulin preparations.     

Optically responsive nanoparticle layers for the label-free analysis of biospecific interactions in array formats

A novel nanocomposite surface is prepared by coating surface-adsorbed dielectric colloidal particles with a contiguous layer of gold nanoparticles. The resulting surface shows pronounced optical extinction in reflection with the extinction peaks located in the UV–Vis and NIR region of the electromagnetic spectrum. The peak positions of these maxima change very sensitively with the adsorption of organic molecules onto the surface. For the adsorption of a monolayer of octadecanethiol, we observe a peak shift of 55 nm on average, which is about five times that of established label-free sensing methods based on propagating and localized surface plasmons. In a first proof-of-principle experiment, the interaction of peptides with specific antibodies has been detected without labeling by means of a fiber-optical set-up with microscopic lateral resolution. To avoid crosstalk in high-density arrays, the optically responsive surface areas can be locally separated on a micro- or even nanometer scale. Accordingly, the newly developed optically responsive surfaces are well suited for integration into high-density peptide or DNA arrays as demanded in genomics, proteomics, and biomedical research in general.     

Label-free detection of single nucleotide polymorphisms utilizing the differential transfer function of field-effect transistors

We present a label-free method for the detection of DNA hybridization, which is monitored by non-metallized silicon field-effect transistors (FET) in a microarray approach. The described method enables a fast and fully electronic readout of ex situ binding assays. The label-free detection utilizing the field-effect is based on the intrinsic charge of the DNA molecules and/or on changes of the solid–liquid interface impedance, when biomolecules bind to the sensor surface. With our sensor system, usually a time-resolved, dc readout is used. In general, this FET signal suffers from sensor drift, temperature drift, changes in electrolyte composition or pH value, influence of the reference electrode, etc. In this article, we present a differential ac readout concept for FET microarrays, which enables a stable operation of the sensor against many of these side-parameters, reliable readout and a possibility for a quick screening of large sensor arrays. We present the detection of point mutations in short DNA samples with this method in an ex situ binding assay.