Fast Fluoroalkylation of Proteins Uncovers the Structure and Dynamics of Biological Macromolecules.

Covalent labeling of proteins in combination with mass spectrometry has been established as a complementary technique to classical structural methods, such as X-ray, NMR, or cryogenic electron microscopy (Cryo-EM), used for protein structure determination. Although the current covalent labeling techniques enable the protein solvent accessible areas with sufficient spatial resolution to be monitored, there is still high demand for alternative, less complicated, and inexpensive approaches. Here, we introduce a new covalent labeling method based on fast fluoroalkylation of proteins (FFAP). FFAP uses fluoroalkyl radicals formed by reductive decomposition of Togni reagents with ascorbic acid to label proteins on a time scale of seconds. The feasibility of FFAP to effectively label proteins was demonstrated by monitoring the differential amino acids modification of native horse heart apomyoglobin/holomyoglobin and the human haptoglobin-hemoglobin complex. The obtained data confirmed the Togni reagent-mediated FFAP is an advantageous alternative method for covalent labeling in applications such as protein footprinting and epitope mapping of proteins (and their complexes) in general. Data are accessible via the ProteomeXchange server with the data set identifier PXD027310.

Simple and fast method for recognition of reducing and nonreducing neutral carbohydrates by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.

Negative-ion mode matrix-assisted laser desorption/ionization (MALDI) time-of-flight mass spectrometry (TOF-MS) was used for the characterization of storage, neutral oligosaccharides extracted from Jerusalem artichoke, red onion, and wheat. The oligosaccharides from the real samples were analyzed with 2,4,6-trihydroxyacetophenone as the most convenient matrix that was selected in advance with the standard carbohydrate samples (inulin and maltooligosaccharides). The oligosaccharides from Jerusalem artichoke and red onion (similarly as inulin) produced [M - H](-) peaks as the main distribution, which reflects their nonreducing composition. On the contrary, the cross-ring fragmentations [M - H - 120](-) formed the main distribution in the mass spectra of hydrolyzed wheat starch similarly to reducing maltooligosaccharides and dextrans. The negative-ion mode MALDI-TOF MS is capable of recognizing reducing and nonreducing oligosaccharides. Such a simple differentiation of malto or inulin type of oligosaccharides is not possible in the positive-ion mode.