Visible light triggers the formation of reactive oxygen species in monoclonal antibody formulations.

Most monoclonal antibody formulations require the presence of a surfactant, such as polysorbate, to ensure protein stability. The presence of high concentrations of polysorbate have been shown to enhance photooxidation of certain protein drug products when exposed to visible light. The current literature, however, suggest that photooxidation of polysorbate only occurs when exposed to visible light in combination with UVA light.

Repurposing the mammalian RNA-binding protein Musashi-1 as an allosteric translation repressor in bacteria.

The RNA recognition motif (RRM) is the most common RNA-binding protein domain identified in nature. However, RRM-containing proteins are only prevalent in eukaryotic phyla, in which they play central regulatory roles. Here, we engineered an orthogonal post-transcriptional control system of gene expression in the bacterium with the mammalian RNA-binding protein Musashi-1, which is a stem cell marker with neurodevelopmental role that contains two canonical RRMs.

Kinetic FRET Assay to Measure Binding-Induced Conformational Changes of Nucleic Acids.

The interaction of small molecules or proteins with RNA or DNA often involves changes in the nucleic acid (NA) folding and structure. A biophysical characterization of these processes helps us to understand the underlying molecular mechanisms. Here, we propose kinFRET (kinetics Förster resonance energy transfer), a real-time ensemble FRET methodology to measure binding and folding kinetics.

Protein photodegradation in the visible range? Insights into protein photooxidation with respect to protein concentration.

Visible light (400-800 nm) can lead to photooxidation of protein formulations, which might impair protein integrity. However, the relevant mechanism of photooxidation upon visible light exposure is still unclear for therapeutic proteins, since proteinogenic structures do not absorb light in the visible range. Here, we show that exposure of monoclonal antibody formulations to visible light, lead to the formation of reactive oxygen species (ROS), which subsequently induce specific protein degradations.

Photo-Oxidation of Therapeutic Protein Formulations: From Radical Formation to Analytical Techniques.

UV and ambient light-induced modifications and related degradation of therapeutic proteins are observed during manufacturing and storage. Therefore, to ensure product quality, protein formulations need to be analyzed with respect to photo-degradation processes and eventually protected from light exposure. This task usually demands the application and combination of various analytical methods.

Investigating photodegradation of antibodies governed by the light dosage.

The present study investigated the photodegradation of three different monoclonal antibodies (mAb) by visible light. Several chromatographic techniques, such as size-exclusion and hydrophobic interaction chromatography as well as mass spectrometry were used to measure relative changes of various oxidation related monoclonal antibody species. The results show that visible light is indeed capable of inducing the formation of protein photo-oxidation products, such as acidic, basic, hydrophilic, and several other protein species with altered physicochemical properties.

Nucleotide binding kinetics and conformational change analysis of tissue transglutaminase with switchSENSE.

Function, activity, and interactions of proteins crucially depend on their three-dimensional structure and are often regulated by effector binding and environmental changes. Tissue transglutaminase (Transglutaminase 2, TG2) is a multifunctional protein, allosterically regulated by nucleotides and Ca ions, which trigger opposing conformational changes. Here we introduce switchSENSE as a versatile tool for TG2 characterization and provide novel insights into protein conformation as well as analyte binding kinetics.

Electrical Actuation of a DNA Origami Nanolever on an Electrode.

Development of electrically powered DNA origami nanomachines requires effective means to actuate moving origami parts by externally applied electric fields. We demonstrate how origami nanolevers on an electrode can be manipulated (switched) at high frequency by alternating voltages. Orientation switching is long-time stable and can be induced by applying low voltages of 200 mV.

Protein analysis by time-resolved measurements with an electro-switchable DNA chip.

Measurements in stationary or mobile phases are fundamental principles in protein analysis. Although the immobilization of molecules on solid supports allows for the parallel analysis of interactions, properties like size or shape are usually inferred from the molecular mobility under the influence of external forces. However, as these principles are mutually exclusive, a comprehensive characterization of proteins usually involves a multi-step workflow.

Quantitation of affinity, avidity, and binding kinetics of protein analytes with a dynamically switchable biosurface.

A label-free method for the analysis of interactions of proteins with surface-tethered ligands is introduced. Short DNA levers are electrically actuated on microelectrodes by ac potentials, and their switching dynamics are measured in real-time by fluorescence energy transfer. Binding of proteins to ligands attached to the top of the DNA levers is detected by time-resolved measurements of the levers' dynamic motion.

Conformations of end-tethered DNA molecules on gold surfaces: influences of applied electric potential, electrolyte screening, and temperature.

We describe the behavior of 72mer oligonucleotides that are end-tethered to gold surfaces under the influence of applied electric fields. The DNA extension is measured by fluorescence energy transfer as a function of the DNA hybridization state (single- and double-stranded), the concentration of monovalent salt in solution (100 microM to 1 M NaCl), the applied electrode potential (-0.6 to +0.

Detection and size analysis of proteins with switchable DNA layers.

We introduce a chip-compatible scheme for the label-free detection of proteins in real-time that is based on the electrically driven conformation switching of DNA oligonucleotides on metal surfaces. The switching behavior is a sensitive indicator for the specific recognition of IgG antibodies and antibody fragments, which can be detected in quantities of less than 10(-18) mol on the sensor surface. Moreover, we show how the dynamics of the induced molecular motion can be monitored by measuring the high-frequency switching response.

Remember judgments and the constraint of direct experience.

The most direct assessment of episodic memory is provided by Remember versus Know judgments of recalled or recognised items. We investigate whether Remember judgments reflect episodic memories as a re-experience of formerly experienced events (mental time travel). If they do, they must obey the direct experience constraint: only directly experienced events can be re-experienced but not when the event is known through indirectly conveyed information.

Subjective evaluation: is there more than one criterion?

Previous cross-sectional investigations have shown that subjective evaluation criteria (criteria that are used in psychiatric research for evaluating care based on patients' statements) do overlap and that there exists a single general factor underlying all these criteria. In this study, we tested longitudinally and in two different samples of schizophrenia patients the distinctness and covariation at baseline and at followup of three common subjective evaluation criteria (subjective quality of life, self-rated needs, and self-reported symptoms). Scores were intercorrelated at both baseline and followup and showed some intercorrelations over time, suggesting temporal covariation.

Do long-term hospitalised patients benefit from discharge into the community?

Objective: The study investigated whether long-stay patients would benefit from discharge into the community in Berlin, Germany.

Method: In a prospective controlled study, all long-term hospitalised psychiatric patients from a defined catchment area were assessed using established standardised instruments. Quality of life, treatment satisfaction, needs and psychopathology were re-assessed in 63 non-discharged patients 1.