Toward an understanding of the conformational plasticity of S100A8 and S100A9 Ca2+-binding proteins.

S100A8 and S100A9 are small, human, Ca2+-binding proteins with multiple intracellular and extracellular functions in signaling, regulation, and defense. The two proteins are not detected as monomers but form various noncovalent homo- or hetero-oligomers related to specific activities in human physiology. Because of their significant roles in numerous medical conditions, there has been intense research on the conformational properties of various S100A8 and S100A9 proteoforms as essential targets of drug discovery. NMR or crystal structures are currently available only for mutated or truncated protein complexes, mainly with bound metal ions, that may well reflect the proteins' properties outside cells but not in other biological contexts in which they perform. Here, we used structural mass spectrometry methods combined with molecular dynamics simulations to compare the conformations of wildtype full-length S100A8 and S100A9 subunits in biologically relevant homo- and heterodimers and in higher oligomers formed in the presence of calcium or zinc ions. We provide, first, rationales for their functional response to changing environmental conditions, by elucidating differences between proteoforms in flexible protein regions that may provide the plasticity of the binding sites for the multiple targets, and second, the key factors contributing to the variable stability of the oligomers. The described methods and a systematic view of the conformational properties of S100A8 and S100A9 complexes provide a basis for further research to characterize and modulate their functions for basic science and therapies.

Vimentin S-glutathionylation at Cys328 inhibits filament elongation and induces severing of mature filaments in vitro.

Vimentin intermediate filaments are a significant component of the cytoskeleton in cells of mesenchymal origin. In vivo, filaments assemble and disassemble and thus participate in the dynamic processes of the cell. Post-translational modifications (PTMs) such as protein phosphorylation regulate the multiphasic association of vimentin from soluble complexes to insoluble filaments and the reverse processes. The thiol side chain of the single vimentin cysteine at position 328 (Cys328) is a direct target of oxidative modifications inside cells. Here, we used atomic force microscopy, electron microscopy and a novel hydrogen-deuterium exchange mass spectrometry (HDex-MS) procedure to investigate the structural consequences of S-nitrosylation and S-glutathionylation of Cys328 for in vitro oligomerisation of human vimentin. Neither modification affects the lateral association of tetramers to unit-length filaments (ULF). However, S-glutathionylation of Cys328 blocks the longitudinal assembly of ULF into extended filaments. S-nitrosylation of Cys328 does not hinder but slows down the elongation. Likewise, S-glutathionylation of preformed vimentin filaments causes their extensive fragmentation to smaller oligomeric species. Chemical reduction of the S-glutathionylated Cys328 thiols induces reassembly of the small fragments into extended filaments. In conclusion, our in vitro results suggest S-glutathionylation as a candidate PTM for an efficient molecular switch in the dynamic rearrangements of vimentin intermediate filaments, observed in vivo, in response to changes in cellular redox status. Finally, we demonstrate that HDex-MS is a powerful method for probing the kinetics of vimentin filament formation and filament disassembly induced by PTMs.

Synergistic Action of Sodium Selenite with some Antidepressants and Diazepam in Mice.

BACKGROUND: The antidepressant and anxiolytic effects of selenium (Se) have been proven in many studies. This work was aimed at confirming these activities of its inorganic form-sodium selenite-and examining the possible synergy of action with antidepressants and diazepam. METHODS: The antidepressant- and anxiolytic-like activity of Se was assessed using forced swim tests (FSTs) and elevated plus-maze test (EPMs). Spontaneous locomotor activity was measured using photoresistor actimeters. The experiments were conducted on male Albino Swiss mice. RESULTS: Sodium selenite (0.5 mg/kg) reduced the immobility time in the FSTs and extended time spent in the open arms of EPMs without affecting locomotor activity The combined administration of Se at an ineffective dose (0.25 mg/kg) together with imipramine (15 mg/kg), fluoxetine (5 mg/kg), tianeptine (10 mg/kg), but not with reboxetine (2.5 mg/kg), resulted in a reduction of immobility time in FSTs, and with a threshold dose of diazepam (0.25 mg/kg) led to the prolongation of time spent in the open arms of the EPM. Moreover, the antidepressant-like effect of Se (0.5 mg/kg) was significantly reduced by pretreatment with p-chlorophenylalanine (100 mg/kg). CONCLUSIONS: The results may indicate the participation of serotonergic transmission to antidepressant action of Se and GABA-ergic transmission to its anxiolytic effects.