Interaction of the α2A domain of integrin with small collagen fragments

We here present a detailed study of the ligand-receptor interactions between single and triple-helical strands of collagen and the α2A domain of integrin (α2A), providing valuable new insights into the mechanisms and dynamics of collagen-integrin binding at a sub-molecular level. The occurrence of single and triple-helical strands of the collagen fragments was scrutinized with atom force microscopy (AFM) techniques. Strong interactions of the triple-stranded fragments comparable to those of collagen can only be detected for the 42mer triple-helical collagen-like peptide under study (which contains 42 amino acid residues per strand) by solid phase assays as well as by surface plasmon resonance (SPR) measurements. However, changes in NMR signals during titration and characteristic saturation transfer difference (STD) NMR signals are also detectable when α2A is added to a solution of the 21mer single-stranded collagen fragment. Molecular dynamics (MD) simulations employing different sets of force field parameters were applied to study the interaction between triple-helical or single-stranded collagen fragments with α2A. It is remarkable that even single-stranded collagen fragments can form various complexes with α2A showing significant differences in the complex stability with identical ligands. The results of MD simulations are in agreement with the signal alterations in our NMR experiments, which are indicative of the formation of weak complexes between single-stranded collagen and α2A in solution. These results provide useful information concerning possible interactions of α2A with small collagen fragments that are of relevance to the design of novel therapeutic A-domain inhibitors.     

Indirect role of Ca2+ in the assembly of extracellular matrix proteins

We have investigated through molecular dynamics the binding properties at the interface between the C-type lectin subdomain (CLD) of aggrecan and the fibronectin type III domains (^4-5FnIII) of tenasin, in particular the mechanistically unknown, but essential role of Ca²⁺ in the binding. The binding between the CLD and ^4-5FnIII is critical in cross-linking aggrecan, hyaluronan, and tenascin to form extended protein networks found in tissues such as cartilage. None of the structurally resolved Ca²⁺ ions in the complex of the CLD and ^4-5FnIII is directly bridging the two proteins. However, one of the Ca²⁺ ions (Ca2) is found to play the role of maintaining the structure of the L4 loop at the CLD binding surface, thus facilitating a high affinity binding between the CLD and ^4-5FnIII. Removal of this Ca²⁺ ion causes a drastic structural change in the L4 loop, which presumably hinders the binding of the CLD to the ^4-5FnIII. The other bound Ca²⁺ ions (Ca1 and Ca3) have no significant effect on the structure of the CLD binding surface, and thus are not expected to affect the binding. Our results might also suggest that the role of Ca2 in maintaining the structure of the L4 loop is most important during the binding process. Once the complex is formed, the dependence of the complex on the structuring role of Ca2 is reduced. In response to tensile force, the CLD and ^4-5FnIII separate by breaking first the electrostatic interactions at the interface, followed by the hydrophobic ones. The sequence of the unbinding events and the maximum force required to separate the two proteins are independent of the presence of the Ca²⁺ ions, underlying the indirect role of Ca²⁺ in the binding.     

Dynamics of the extracellular gate and ion-substrate coupling in the glutamate transporter

Glutamate transporters (GluTs) are the primary regulators of extracellular concentration of the neurotransmitter glutamate in the central nervous system. In this study, we have investigated the dynamics and coupling of the substrate and Na⁺ binding sites, and the mechanism of cotransport of Na⁺ ions, using molecular dynamics simulations of a membrane-embedded model of GluT in its apo (empty form) and various Na⁺- and/or substrate-bound states. The results shed light on the mechanism of the extracellular gate and on the sequence of binding of the substrate and Na⁺ ions to GluT during the transport cycle. The results suggest that the helical hairpin HP2 plays the key role of the extracellular gate for the substrate binding site, and that the opening and closure of the gate is controlled by substrate binding. GluT adopts an open conformation in the absence of the substrate exposing the binding sites of the substrate and Na⁺ ions to the extracellular solution. Based on the calculated trajectories, we propose that Na1 is the first element to bind GluT, as it is found to be important for the completion of the substrate binding site. The subsequent binding of the substrate, in turn, is shown to result in an almost complete closure of the extracellular gate and the formation of the Na2 binding site. Finally, binding of Na2 locks the extracellular gate and completes the formation of the occluded state of GluT.     

The Cenomanian–Turonian boundary in Jordan: Ammonite biostratigraphy and faunal turnover

Well-exposed fossiliferous Upper Cenomanian–Lower Turonian marine sedimentary rocks are present in west-central Jordan. Ammonites serve as an important faunal marker for this interval and can be used to subdivide the Cenomanian–Turonian transition into two upper Cenomanian biozones (Neolobites vibrayeanus and Vascoceras cauvini) and two lower Turonian biozones (Vascoceras proprium and Choffaticeras segne). A revised stratigraphic range of the Vascoceras cauvini Zone in the study area is proposed, consisting of the Metoicoceras geslinianum and Neocardioceras juddii zones of the standard zonation. Based on intercontinental biostratigraphic correlation, a minor unconformity appears to be present around the Cenomanian–Turonian boundary, and a part of the lower Turonian is probably missing. In addition, a faunal turnover is recorded in the uppermost Cenomanian, marked by the disappearance of most of the Cenomanian taxa, including Costagyra olisiponensis (Sharpe), Ceratostreon flabellatum (Goldfuss), Ilymatogyra africana (Lamarck), Rhynchostreon suborbiculatum (Lamarck), Harpagodes nodosus (Sowerby), and Heterodiadema libycum (Desor). This bioevent is thought to be an effect of the Oceanic Anoxic Event OAE 2; the dramatic shifts in species richness and diversity spanning the Cenomanian–Turonian boundary in the study area occurred in response to the major paleoclimatic and paleoenvironmental perturbations prevailing at that time. The stratigraphic and paleontological patterns studied in Jordan are very similar to those recorded in Egypt in terms of litho- and biostratigraphy, event stratigraphy, and macroinvertebrate content, suggesting the presence of uniform triggering mechanisms and bio-sedimentary responses in the Upper Cretaceous basins of the Middle East and providing clues for a high-resolution correlation between the two areas.     

Area reduction and trace element pollution in Nile Delta wetland ecosystems

The three main Nile Delta wetland ecosystems, Manzala, Burullus and Edku lagoons, are among the most ecologically important and productive habitats in Egypt. We studied the area degradation and the human health risks associated with trace metal accumulation in Tilapia zillii harvested from these lakes. The area of Manzala lagoon has shrunken from about 3035 km² in 1800 to about 288 km² in 2015, the area of Edku has shrunken from about 336 km² in 1824 to about 18 km² in 2014, and the area of Burullus has shrunken from about 1116 km² in 1949 to about 546 km² in 2014. This area degradation is attributed to drying for housing, land reclamation and fish farming. As a result, the concentration of pollutants and nutrients has subsequently increased, and large parts of the lakes have been overgrown with aquatic vegetation, which increased the rate of degradation and land transformation. Metal pollution was detected in water, sediment and edible fish harvested from the lakes. The hazard index, an indicator of human health risks associated with fish consumption, showed adverse health effects of zinc and lead metals for habitual fish consumers. The impact of the high dam on the lakes was discussed.     

Electrostatic tuning of permeation and selectivity in aquaporin water channels

Water permeation and electrostatic interactions between water and channel are investigated in the Escherichia coli glycerol uptake facilitator GlpF, a member of the aquaporin water channel family, by molecular dynamics simulations. A tetrameric model of the channel embedded in a 16:0/18:1c9-palmitoyloleylphosphatidylethanolamine membrane was used for the simulations. During the simulations, water molecules pass through the channel in single file. The movement of the single file water molecules through the channel is concerted, and we show that it can be described by a continuous-time random-walk model. The integrity of the single file remains intact during the permeation, indicating that a disrupted water chain is unlikely to be the mechanism of proton exclusion in aquaporins. Specific hydrogen bonds between permeating water and protein at the channel center (at two conserved Asp-Pro-Ala "NPA" motifs), together with the protein electrostatic fields enforce a bipolar water configuration inside the channel with dipole inversion at the NPA motifs. At the NPA motifs water-protein electrostatic interactions facilitate this inversion. Furthermore, water-water electrostatic interactions are in all regions inside the channel stronger than water-protein interactions, except near a conserved, positively charged Arg residue. We find that variations of the protein electrostatic field through the channel, owing to preserved structural features, completely explain the bipolar orientation of water. This orientation persists despite water translocation in single file and blocks proton transport. Furthermore, we find that for permeation of a cation, ion-protein electrostatic interactions are more unfavorable at the conserved NPA motifs than at the conserved Arg, suggesting that the major barrier against proton transport in aquaporins is faced at the NPA motifs.     

Antitumor activities of iodoacetate and dimethylsulphoxide against solid Ehrlich carcinoma growth in mice

Treatment of tumor-bearing mice with LD12.5 values of iodoacetate; IAA (1.84 mg/100 g b.w.) and/or dimethylsulphoxide; DMSO (350 mg/100 g b.w.) significantly increased the cumulative mean survival time and percentage of survivors and reduced the mean tumor weight, compared to tumor-bearing controls, however, a more pronounced effect is recorded in the combined treatment. Also, an increase in the life span (ILS%) and tumor growth inhibition ratio (T/C%) are reported and amounted to 145.78 and 43.80%, 195.54 and 61.30% and 220.77 and 78.40% in IAA, DMSO and combined-treated groups, respectively. Results obtained from biochemical studies reveal that a single IAA treatment of tumor-bearing mice significantly increased the levels of plasma lactate dehydrogenase (LDH) activity, while it also significantly decreased the levels of plasma glucose and liver total protein, RNA and DNA, compared to normal controls. On the other hand, a single DMSO treatment significantly elevated the activities of blood antioxidant enzymes, i.e. glutathione peroxidase (GPx) and glucose-6-phosphate dehydrogenase (G6PDH) and decreased the liver RNA and DNA levels. Combined treatment increased significantly the levels of plasma LDH and erythrocytes G6PDH activities, as well as liver glycogen, and in contrast it decreased the levels of liver total protein, RNA and DNA, compared to normal controls.     

The polypeptide chain-releasing factor GSPT1/eRF3 is proteolytically processed into an IAP-binding protein

Smac/Diablo and HtrA2/Omi are inhibitors of apoptosis (IAP)-binding proteins released from the mitochondria of human cells during apoptosis and regulate apoptosis by liberating caspases from IAP inhibition. Here we describe the identification of a proteolytically processed isoform of the polypeptide chain-releasing factor GSPT1/eRF3 protein, which functions in translation, as a new IAP-binding protein. In common with other IAP-binding proteins, the processed GSPT1 protein harbors a conserved N-terminal IAP-binding motif (AKPF). Additionally, processed GSPT1 interacts biochemically with IAPs and could promote caspase activation, IAP ubiquitination and apoptosis. The IAP-binding motif of the processed GSPT1 is absolutely required for these activities. Our findings are consistent with a model whereby processing of GSPT1 into the IAP-binding isoform could potentiate apoptosis by liberating caspases from IAP inhibition, or target IAPs and the processed GSPT1 for proteasome-mediated degradation.