Specific protein-RNA interactions are mostly preserved in biomolecular condensates.

Many biomolecular condensates are enriched in and depend on RNAs and RNA binding proteins (RBPs). So far, only a few studies have addressed the characterization of the intermolecular interactions responsible for liquid-liquid phase separation (LLPS) and the impact of condensation on RBPs and RNAs. Here, we present an approach to study protein-RNA interactions inside biomolecular condensates by applying cross-linking of isotope labeled RNA and tandem mass spectrometry to phase-separating systems (LLPS-CLIR-MS).

A hybrid structure determination approach to investigate the druggability of the nucleocapsid protein of SARS-CoV-2.

The pandemic caused by SARS-CoV-2 has called for concerted efforts to generate new insights into the biology of betacoronaviruses to inform drug screening and development. Here, we establish a workflow to determine the RNA recognition and druggability of the nucleocapsid N-protein of SARS-CoV-2, a highly abundant protein crucial for the viral life cycle. We use a synergistic method that combines NMR spectroscopy and protein-RNA cross-linking coupled to mass spectrometry to quickly determine the RNA binding of two RNA recognition domains of the N-protein.

Protein-RNA interactions: from mass spectrometry to drug discovery.

Proteins and RNAs are fundamental parts of biological systems, and their interactions affect many essential cellular processes. Therefore, it is crucial to understand at a molecular and at a systems level how proteins and RNAs form complexes and mutually affect their functions. In the present mini-review, we will first provide an overview of different mass spectrometry (MS)-based methods to study the RNA-binding proteome (RBPome), most of which are based on photochemical cross-linking.

Cross-linking and mass spectrometry as a tool for studying the structural biology of ribonucleoproteins.

Cross-linking and mass spectrometry (XL-MS) workflows represent an increasingly popular technique for low-resolution structural studies of macromolecular complexes. Cross-linking reactions take place in the solution state, capturing contact sites between components of a complex that represent the native, functionally relevant structure. Protein-protein XL-MS protocols are widely adopted, providing precise localization of cross-linking sites to single amino acid positions within a pair of cross-linked peptides.

"Thermal Peroxidation" of Dietary Pentapeptides Yields N-Terminal 1,2-Dicarbonyls.

In this contribution we investigate the thermal degradation of dietary-relevant pentapeptides. Most unsaturated lipids degrade by the well-known peroxidation mechanism. Here we show a degradation mechanism of peptides analogous to lipid peroxidation, forming a series of novel degradation products with possible toxicological relevance.

Heat induced hydrolytic cleavage of the peptide bond in dietary peptides and proteins in food processing.

We investigate the hypothesis that proteins and peptides are thermally degraded by hydrolytic bond cleavage of amide bonds, hence yielding shorter peptides as main degradation products. A series of fifteen pentapeptides with varying sequences was subjected to heating. Products were investigated by targeted UHPLC-ESI-tandem mass spectrometry and targeted analysis revealed formation of 2,5-diketopiperazines, di- and tri-peptides.

A proteolytic nanobiocatalyst with built-in disulphide reducing properties.

We report a method to equip proteolytic nanobiocatalysts with intrinsic disulphide bond reducing properties. After immobilisation onto silica particles, selected protease enzymes are partially shielded in a nanometre-thick mercaptosilica layer acting not only as a protective system but also as a substrate reducing agent. The biocatalysts produced efficiently perform simultaneous disulphide bond reduction and protein digestion.

Identification of Products from Thermal Degradation of Tryptophan Containing Pentapeptides: Oxidation and Decarboxylation.

In this contribution, we investigate the thermal decomposition of four pentapeptides containing a tryptophan moiety. Pentapeptides were heated at 220 °C, and the resulting reaction mixtures were investigated by HPLC coupled to high-resolution mass spectrometry and tandem mass spectrometry. A total of 95 thermal decomposition products could be observed and resolved by chromatography.