Characterization of BNT162b2 mRNA to Evaluate Risk of Off-Target Antigen Translation.

mRNA vaccines have been established as a safe and effective modality, thanks in large part to the expedited development and approval of COVID-19 vaccines. In addition to the active, full-length mRNA transcript, mRNA fragment species can be present as a byproduct of the cell-free transcription manufacturing process or due to mRNA hydrolysis. In the current study, mRNA fragment species from BNT162b2 mRNA were isolated and characterized.

Hormone replacement therapy subcutaneous implants for refractory menopause symptoms; the patient's perspective.

Objective: For women with menopause symptoms refractory to standard hormone replacement therapy (HRT) preparations, HRT implants offer an alternative. The primary objective of this study was to evaluate women's perceptions regarding efficacy, tolerability, satisfaction and safety of implant therapy.

Study Design: A single centre service evaluation study performed at Birmingham Women's & Children's Foundation Hospital Trust.

Utility of High Resolution NMR Methods to Probe the Impact of Chemical Modifications on Higher Order Structure of Monoclonal Antibodies in Relation to Antigen Binding.

Purpose: An understanding of higher order structure (HOS) of monoclonal antibodies (mAbs) could be critical to predicting its function. Amongst the various factors that can potentially affect HOS of mAbs, chemical modifications that are routinely encountered during production and long-term storage are of significant interest.

Methods: To this end, two Pfizer mAbs were subjected to forced deamidation stress for a period of eight weeks.

The Atypical MAP Kinase ErkB Transmits Distinct Chemotactic Signals through a Core Signaling Module.

Signaling from chemoattractant receptors activates the cytoskeleton of crawling cells for chemotaxis. We show using phosphoproteomics that different chemoattractants cause phosphorylation of the same core set of around 80 proteins in Dictyostelium cells. Strikingly, the majority of these are phosphorylated at an [S/T]PR motif by the atypical MAP kinase ErkB.

Flotillin proteins recruit sphingosine to membranes and maintain cellular sphingosine-1-phosphate levels.

Sphingosine-1-phosphate (S1P) is an important lipid signalling molecule. S1P is produced via intracellular phosphorylation of sphingosine (Sph). As a lipid with a single fatty alkyl chain, Sph may diffuse rapidly between cellular membranes and through the aqueous phase.

Quantitative proteomics screen identifies a substrate repertoire of rhomboid protease RHBDL2 in human cells and implicates it in epithelial homeostasis.

Rhomboids are intramembrane serine proteases conserved in all kingdoms of life. They regulate epidermal growth factor receptor signalling in Drosophila by releasing signalling ligands from their transmembrane tethers. Their functions in mammals are poorly understood, in part because of the lack of endogenous substrates identified thus far.

Hekate: software suite for the mass spectrometric analysis and three-dimensional visualization of cross-linked protein samples.

Chemical cross-linking of proteins combined with mass spectrometry provides an attractive and novel method for the analysis of native protein structures and protein complexes. Analysis of the data however is complex. Only a small number of cross-linked peptides are produced during sample preparation and must be identified against a background of more abundant native peptides.

Lissencephaly-1 promotes the recruitment of dynein and dynactin to transported mRNAs.

Microtubule-based transport mediates the sorting and dispersal of many cellular components and pathogens. However, the mechanisms by which motor complexes are recruited to and regulated on different cargos remain poorly understood. Here we describe a large-scale biochemical screen for novel factors associated with RNA localization signals mediating minus end-directed mRNA transport during Drosophila development.

Bicaudal-D uses a parallel, homodimeric coiled coil with heterotypic registry to coordinate recruitment of cargos to dynein.

Cytoplasmic dynein is the major minus end-directed microtubule motor in eukaryotes. However, there is little structural insight into how different cargos are recognized and linked to the motor complex. Here we describe the 2.

Abnormal glycosphingolipid mannosylation triggers salicylic acid-mediated responses in Arabidopsis.

The Arabidopsis thaliana protein GOLGI-LOCALIZED NUCLEOTIDE SUGAR TRANSPORTER (GONST1) has been previously identified as a GDP-d-mannose transporter. It has been hypothesized that GONST1 provides precursors for the synthesis of cell wall polysaccharides, such as glucomannan. Here, we show that in vitro GONST1 can transport all four plant GDP-sugars.

Structural characterization of Arabidopsis leaf arabinogalactan polysaccharides.

Proteins decorated with arabinogalactan (AG) have important roles in cell wall structure and plant development, yet the structure and biosynthesis of this polysaccharide are poorly understood. To facilitate the analysis of biosynthetic mutants, water-extractable arabinogalactan proteins (AGPs) were isolated from the leaves of Arabidopsis (Arabidopsis thaliana) plants and the structure of the AG carbohydrate component was studied. Enzymes able to hydrolyze specifically AG were utilized to release AG oligosaccharides.

Determination of the N-glycosylation patterns of seed proteins: applications to determine the authenticity and substantial equivalence of genetically modified (GM) crops.

Methods have been developed to determine the N-glycosylation pattern of proteins at the single-seed level in two different biological systems. These were the well-characterized and widely consumed storage protein phaseolin from several species of Phaseolus (bean) and the α-amylase inhibitor from the same Phaseolus species expressed transgenically in pea. The N-glycosylation pattern of the α-amylase inhibitor expressed transgenically in pea was different from that of the inhibitor present in common bean (P.

Carbohydrate structural analysis of wheat flour arabinogalactan protein.

The water-extractable arabinogalactan protein (AGP) was isolated from bread wheat flour (Triticum aestivum L. variety Cadenza) and the structure of the arabinogalactan (AG) carbohydrate component was studied. Oligosaccharides, released by hydrolysis of the AG with a range of AGP-specific enzymes, were characterised by Matrix Assisted Laser Desorption Ionisation (MALDI)-Time of Flight (ToF)-Mass Spectrometry (MS), MALDI-ToF/ToF high energy collision induced dissociation (CID) and Polysaccharide Analysis by Carbohydrate gel Electrophoresis (PACE).

Stabilization of mutant p53 via alkylation of cysteines and effects on DNA binding.

Oncogenic mutations inactivate the tumor suppressor p53 by lowering its stability or by weakening its binding to DNA. Alkylating agents that reactivate mutant p53 are currently being explored for cancer therapy. We have discovered ligands containing an α,β-unsaturated double bond, characteristic of Michael acceptors, that bind covalently to generic cysteine sites in the p53 core domain.

Absence of branches from xylan in Arabidopsis gux mutants reveals potential for simplification of lignocellulosic biomass.

As one of the most abundant polysaccharides on Earth, xylan will provide more than a third of the sugars for lignocellulosic biofuel production when using grass or hardwood feedstocks. Xylan is characterized by a linear β(1,4)-linked backbone of xylosyl residues substituted by glucuronic acid, 4-O-methylglucuronic acid or arabinose, depending on plant species and cell types. The biological role of these decorations is unclear, but they have a major influence on the properties of the polysaccharide.

Analysis of carbohydrates on proteins by offline normal-phase liquid chromatography MALDI-TOF/TOF-MS/MS.

Glycoproteins typically exist as a diverse population of glycoforms, which can consist of a great number of different glycan structures (including structural isomers) that vary in their degree of occupancy at given glycosylation sites. Hence, the detailed characterization of such glycans can be a very demanding task. Liquid chromatography in combination with mass spectrometric techniques can provide a fast and sensitive tool for the analysis of these structurally complex molecules.

Aberrant attachment of heme to cytochrome by the Ccm system results in a cysteine persulfide linkage.

The system I cytochrome c maturation (Ccm) apparatus has been shown to handle a wide variety of apocytochrome substrates containing the CX(n)CH heme attachment sequence, where n = 2, 3, or 4 in natural sequences. When n = 5 or 6, the apparatus also appears to handle these substrates correctly, but close inspection reveals that the resulting mature cytochromes are mixtures of species containing extra mass. We have used accurate mass spectrometry to analyze peptide digests of matured Escherichia coli cytochrome cb(562) with n = 1, 5, or 6 and shown that an extra sulfur is sometimes incorporated into the heme-protein linkage.

Synthetic chain terminators off-load intermediates from a type I polyketide synthase.

Modular biocatalysis is responsible for the generation of countless bioactive products and its mining remains a major focus for drug discovery purposes. One of the enduring hurdles is the isolation of biosynthetic intermediates in a readily-analysed form. We prepared a series of nonhydrolysable pantetheine and N-acetyl cysteamine mimics of the natural (methyl)malonyl extender units recruited for polyketide formation.