Characterization of peptide components in the venom of the scorpion Liocheles australasiae (Hemiscorpiidae).

Scorpion venoms are composed of a number of neurotoxic peptides. A variety of toxins have been isolated from the venoms of scorpions of the family Buthidae, however, little interest has been paid to non-Buthidae scorpions. In this study, we examined the toxicity of the venom of Liocheles australasiae (Hemiscorpiidae) to mice and crickets, and characterized the peptide components by HPLC and mass spectrometry. Over 200 components were detected in the L. australasiae venom by LC/MS analysis, with components of molecular masses ranging from 500 to 5000 Da being particularly abundant. A number of peptides contained two to four disulfide bridges, which was estimated based on the mass difference after derivatization of Cys residues. A peptide having a monoisotopic molecular mass of 7781.6 Da and four disulfide bridges was isolated from the venom. The peptide has a primary structure similar in terms of the position of eight Cys residues to those observed in several peptides found from scorpions, ticks and insects, although biological roles of these peptides are unknown.     

Erythrocyte reduced/oxidized glutathione and serum thiol/disulfide homeostasis in patients with rheumatoid arthritis

BackgroundChronic inflammation and oxidative stress are the most known mechanisms in Rheumatoid Arthritis (RA) pathophysiology, which is still not fully elucidated. In this study, we evaluated oxidative status by determining intracellular reduced/oxidized glutathione (GSH/GSSG) homeostasis and serum thiol/disulfide (SH/SS) homeostasis in RA patients.MethodsA total of 152 RA patient and 89 healthy controls were included in the study. RA patients were subdivided according to disease activity score-28 (DAS-28) as active RA and remission RA. Intracellular GSH/GSSG and serum SH/SS homeostasis parameters were analyzed.ResultsMedian (1st–3rd quartile values) SS/SH and GSSG/GSH percent ratio levels were significantly higher in RA patients (6.94 (6.02–8.54) and 69.8 (44.05–85.29); respectively) compared to controls (4.62 (4.15–5.46) and 34.9 (22.43–62.2); respectively) (p < 0.05 for all). SS/SH and GSSG/GSH percent ratio levels were significantly higher in active RA patients when compared to remission RA patients and controls (p < 0.05 for all). SS/SH and GSSG/GSH percent ratios were significantly increased in remission RA group compared to controls (p < 0.05 for all). DAS28 scores were positively correlated with SS/SH and GSSG/GSH percent ratios (rho = 0.259 and 0.296; respectively).ConclusionsThese findings suggest that active intracellular and extracellular thiol group oxidation process might play a role in RA pathogenesis and further work in these areas may be warranted to show potential value of evaluating intracellular GSSG/GSH and serum SH/SS balances together in disease monitoring.     

Immunoregulation through membrane proteins modified by reducing conditions induced by immune reactions

Selected disulfide bonds in membrane proteins are labile and are thus susceptible to changes in redox potential and/or the presence of thiol isomerase enzymes. Modification of these disulfide bonds can lead to conformational changes of the protein that in turn may alter protein activity and function. This occurs in the entry of several enveloped viruses into their host cells, e.g. HIV, hepatitis C virus and Newcastle disease virus. Labile disulfide bonds are also important in platelet activation, cytokine signalling and in a variety of diseases including cancer and arthritis. In this review we will concentrate on recent advances in understanding the conditions that lead to disulfide bond reduction in membrane proteins and their effects in regulating immune function.     

Disulfide connectivity prediction based on structural information without a prior knowledge of the bonding state of cysteines

Previous studies predicted the disulfide bonding patterns of cysteines using a prior knowledge of their bonding states. In this study, we propose a method that is based on the ensemble support vector machine (SVM), with the structural features of cysteines extracted without any prior knowledge of their bonding states. This method is useful for improving the predictive performance of disulfide bonding patterns. For comparison, the proposed method was tested with the same dataset SPX that was adopted in previous studies. The experimental results demonstrate that bridge classification and disulfide connectivity predictions achieve 96.5% and 89.2% accuracy, respectively, using the ensemble SVM model, which outperforms the traditional method (51.5% and 51.0%, respectively) and the model that is based on a single-kernel SVM classifier (94.6% and 84.4%, respectively). For protein chain and residue classifications, the sensitivity, specificity, and accuracy of ensemble and single-kernel SVM approaches are better than those of the traditional methods. The predictive performances of the ensemble SVM and single-kernel models are identical, indicating that the ensemble model can converge to the single-kernel model for some applications.     

Engineering of the Dsb (disulfide bond) proteins – contribution towards understanding their mechanism of action and their applications in biotechnology and medicine

Abstract

The Dsb protein family in prokaryotes catalyzes the generation of disulfide bonds between thiol groups of cysteine residues in nascent proteins, ensuring their proper three-dimensional structure; these bonds are crucial for protein stability and function. The first Dsb protein, Escherichia coli DsbA, was described in 1991. Since then, many details of the bond-formation process have been described through microbiological, biochemical, biophysical and bioinformatics strategies. Research with the model microorganism E. coli and many other bacterial species revealed an enormous diversity of bond-formation mechanisms. Research using Dsb protein engineering has significantly helped to reveal details of the disulfide bond formation. The first part of this review presents the research that led to understanding the mechanism of action of DsbA proteins, which directly transfer their own disulfide into target proteins. The second part concentrates on the mechanism of electron transport through the cell cytoplasmic membrane. Third and lastly, the review discusses the contribution of this research towards new antibacterial agents.     

Hypoxia augments MHC class I antigen presentation via facilitation of ERO1‐α‐mediated oxidative folding in murine tumor cells

To establish an effective cancer immunotherapy, it is crucial that cancer cells present a cancer‐specific antigen in a hypoxic area, a hallmark of the tumor microenvironment. Here, we show the impact of hypoxia on MHC class I antigen presentation in vitro and in vivo in murine tumors. Activation of antigen‐specific CTLs by tumor cells that had been pre‐incubated under a condition of hypoxia was enhanced compared with that by tumor cells pre‐incubated under a condition of normoxia. Cell surface expression of MHC class I‐peptide complex on the tumor cells was increased under a condition of hypoxia, thereby leading to higher susceptibility to specific CTLs. We show that the hypoxia‐inducible ER‐resident oxidase ERO1‐α plays an important role in the hypoxia‐induced augmentation of MHC class I‐peptide complex expression. ERO1‐α facilitated oxidative folding of MHC class I heavy chains, thereby resulting in the augmentation of cell surface expression of MHC class I‐peptide complex under hypoxic conditions. These results suggest that since the expression of MHC class I‐peptide complex is augmented in a hypoxic tumor microenvironment, strategies for inhibiting the function of regulatory T cells and myeloid‐derived suppressor cells and/or immunotherapy with immune checkpoint inhibitors are promising for improving cancer immunotherapy.     

A winged-helix protein from Sulfolobus turreted icosahedral virus points toward stabilizing disulfide bonds in the intracellular proteins of a hyperthermophilic virus

Sulfolobus turreted icosahedral virus (STIV) was the first non-tailed icosahedral virus to be isolated from an archaeal host. Like other archaeal viruses, its 37 open reading frames generally lack sequence similarity to genes with known function. The roles of the gene products in this and other archaeal viruses are thus largely unknown. However, a protein's three-dimensional structure may provide functional and evolutionary insight in cases of minimal sequence similarity. In this vein, the structure of STIV F93 reveals a homodimer with strong similarity to the winged-helix family of DNA-binding proteins. Importantly, an interchain disulfide bond is found at the dimer interface, prompting analysis of the cysteine distribution in the putative intracellular proteins of the viral proteome. The analysis suggests that intracellular disulfide bonds are common in cellular STIV proteins, where they enhance the thermostability of the viral proteome.     

A remarkable point for evaluating the severity of burns: Thiol–disulfide profile

AimThe objective of this study was to examine the thiol–disulfide profile tests in patients suffering from burn injuries.MethodsThis case–control study comprised 48 patients with thermal burn injuries and 61 healthy individuals. Thiol–disulfide tests were conducted in both groups, and also, the changes of thiol–disulfide parameters were analyzed at zero time and on days 3, 7, 15, and 30 of the admission in patients with burn injuries.ResultsThe patients had significantly decreased native and total thiol levels and native thiol/total thiol ratios, and significantly increased disulfide/native thiol and disulfide/total thiol ratios compared to control individuals (p < 0.001 for all). The variations of native thiol levels, total thiol levels, and disulfide/native thiol ratios were significantly different over time in patients with burn injuries (p < 0.001, p < 0.001, p < 0.05, respectively). There were strong associations with the clinical parameters and thiol–disulfide profile tests (p < 0.05 for all).ConclusionThere was a metabolic disturbance of the thiol–disulfide system among patients with burn injuries. The courses of thiol–disulfide variables in time overlapped with the burn mechanism. Strong associations provide that thiol–disulfide homeostasis might be a notable key for evaluating the severity of burns and predicting the survival.     

以二硫键连接尿嘧啶核苷C-5位肽缀合物的合成

为了方便快捷地制备嘧啶核苷-肽缀合物,我们发展了一种一釜合成的方法。从尿嘧啶核苷出发,经过4步制备了关键中间体5-乙酰巯亚甲基-2′,3′-二-O-异亚丙基尿苷（4）。在酸性条件下,脱除化合物4的乙酰基,同时游离的巯基与PySS-R（8,12,15,Py=2-吡啶基,R=氨基酸或肽）反应形成新的二硫键,即形成了尿苷与氨基酸或肽以二硫键连接的缀合物（9,13,2）。