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1.
Joanna Strumillo Katarzyna E. Nowak Anita Krokosz Aleksandra Rodacka Mieczyslaw Puchala Grzegorz Bartosz 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(4):877-885
Background
Nitric oxide is a well-known gaseous signaling molecule and protein modifying agent. However, at higher concentrations or during oxidative stress nitric oxide may exert some deleterious effects on protein structure and function. Here we investigated the influence of nitric oxide and products of its oxidation on two glycolytic enzymes: GAPDH and LDH under in vitro nitrosative stress conditions. Secondly, we applied natural antioxidants: melatonin and resveratrol to examine their effects on the enzymes under studied conditions.Methods
By means of UV–VIS and fluorescence spectroscopy methods we compared nitric oxide mediated changes of enzyme activities, amount of free sulfhydryl groups (-SH) and bis-ANS probe binding. Finally, we predicted potential cysteine residues modified by nitric oxide in studied proteins using GPS-SNO software.Results
Our results indicated that nitric oxide reversibly inactivates GAPDH but does not affect the activity of LDH. Nitric oxide dependent GAPDH activity decline was accompanied by the reduction of the amount of free –SH groups and GAPDH-bound bis-ANS fluorescence. Reduction of the number of free –SH groups and protein-bound bis-ANS fluorescence was also observed in LDH treated with NO. Applied antioxidants increased inactivation of GAPDH and structural changes of GAPDH and LDH.Conclusions
Nitric oxide modifies function and structure of thiol-dependent enzyme such as GAPDH and structure of LDH which function do not rely on cysteine thiols. Both resveratrol and melatonin exerted prooxidative properties in studied conditions.General significance
Extensively studied antioxidants: resveratrol and melatonin may function as a prooxidative species under in vitro nitrosative stress conditions. 相似文献2.
Jiandong Zhang Zhimei Cui Honghong Chang Xiaojun Fan Qiuyong Zhao Wenlong Wei 《Biotechnology letters》2016,38(9):1559-1564
Objectives
To investigate the efficiency of a cofactor regeneration enzyme co-expressed with a glycerol dehydrogenase for the production of 1,3-dihydroxyacetone (DHA).Results
In vitro biotransformation of glycerol was achieved with the cell-free extracts containing recombinant GlyDH (glycerol dehydrogenase from Escherichia coli), LDH (lactate dehydrogenase form Bacillus subtilis) or LpNox1 (NADH oxidase from Lactobacillus pentosus), giving DHA at 1.3 g l?1 (GlyDH/LDH) and 2.2 g l?1 (GlyDH/LpNox1) with total turnover number (TTN) of NAD+ recycling of 6039 and 11100, respectively. Whole cells of E. coli (GlyDH–LpNox1) co-expressing both GlyDH and LpNox1 were constructed and converted 10 g glycerol l?1 to DHA at 0.2–0.5 g l?1 in the presence of zero to 2 mM exogenous NAD+. The cell free extract of E. coli (GlyDH–LpNox) converted glycerol (2–50 g l?1) to DHA from 0.5 to 4.0 g l?1 (8–25 % conversion) without exogenous NAD+.Conclusions
The disadvantage of the expensive consumption of NAD+ for the production of DHA has been overcome.3.
Yue Liu James Clement Ross Grant Perminder Sachdev Nady Braidy 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(12):2527-2532
Background
Nicotinamide adenine dinucleotide (NAD+) is an essential pyridine nucleotide that is currently investigated as an important target to extend lifespan and health span. Age-related NAD+ depletion due to the accumulation of oxidative stress is associated with reduced energy production, impaired DNA repair and genomic instability.Scope of review
NAD+ levels can be elevated therapeutically using NAD+ precursors or through lifestyle modifications including exercise and caloric restriction. However, high amounts of NAD+ may be detrimental in cancer progression and may have deleterious immunogenic roles.Major conclusions
Standardized quantitation of NAD+ and related metabolites may therefore represent an important component of NAD+ therapy.General significance
Quantitation of NAD+ may serve dual roles not only as an ageing biomarker, but also as a diagnostic tool for the prevention of malignant disorders. 相似文献4.
Wataru Yoshida Mizuki Terasaka Saowalak Laddachote Isao Karube 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(9):1933-1937
Background
DNA methylation at the 5-position of cytosine is an epigenetic modification of CpG dinucleotides. In addition to CpG methylation, the G-quadruplex (G4) structure has been reported as a regulator of gene expression. The identification of G4 forming sequences in CpG islands suggests an involvement of CpG-methylated G4 structures in biological processes; however, few reports have addressed the effects of CpG methylation on G4 structure.Methods
The thermostability of a methylated, 21-mer G4 structure located on the vascular endothelial growth factor (VEGF) gene promoter containing four CpG sites (C1, C6, C11, and C17) were investigated using circular dichroism (CD) spectral analysis.Results
CD melting analysis revealed that VEGF G4 was stabilized by a single CpG methylation on C11 in the presence of Na+ and Mg2+. However, either C1 or C11 methylation enhanced VEGF G4 thermal stability in the presence of K+.Conclusions
Single CpG methylation appears to enhance VEGF G4 thermostability in a manner dependent on both the CpG methylation site and cation type.General significance
These results are expected to contribute to the elucidation of the roles of CpG methylation-stabilized G4 structures in biological processes. 相似文献5.
Xuejing Jia Yeer Liang Chao Zhang Kai Wang Yanbei Tu Meiwan Chen Peng Li Jian-Bo Wan Chengwei He 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(8):1751-1759
Background
Polysaccharides, one of the active ingredients in herbal medicine, are proved to enhance innate immunity against infections. The aim of this study is to explore the immunoregulatory ability of polysaccharides from Rhynchosia minima root in vitro and in vivo.Methods
Polysaccharide fractions of R. minima root were obtained by chromatographic column. The content of NO was measured by spectrophotometry. The levels of cytokines (tumor necrosis factor-α, TNF-α; interleukin-6, IL-6; and monocyte chemoattractant protein-1, MCP-1) were determined by enzyme-linked immuno-sorbent assay (ELISA) kits. The translocation of p65 into the nucleus was imaged by confocal microscopy. The mRNA expression of TNF-α, IL-6, and MCP-1 was determined by quantitative real-time PCR. T-lymphocyte subgroups of spleen from immunosuppressive mouse were evaluated by flow cytometry.Results
PRM3 remarkably enhanced the phagocytic ability of macrophages and promoted the release of NO and the secretion of cytokines (TNF-α, IL-6, and MCP-1) from macrophages. Simultaneously, PRM3 potently activated NF-κB signaling pathway via Toll-like receptor 4 (TLR4). In addition, PRM3 obviously increased the levels of serum cytokines, markedly up-regulated the percentages of CD3+ and CD4+ T lymphocytes and the CD4+/CD8+ ratio of splenocytes, and effectively attenuated cyclophosphamide induced immunosuppression in mice.Conclusions
PRM3 profoundly enhanced the immune function in vitro and in vivo through TLR4-NF-κB pathway and is a promising candidate of immunopotentiator which could be applied in functional foods or drugs.General significance
This study reported a polysaccharide PRM3 from R. minima root exhibited potent immunoenhancing activity and significantly alleviated cyclophosphamide-induced immunosuppression through TLR4-NF-κB pathway. 相似文献6.
M.A. Carrascal M. Silva J.A. Ferreira R. Azevedo D. Ferreira A.M.N. Silva D. Ligeiro L.L. Santos R. Sackstein P.A. Videira 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(9):2069-2080
Background
The glycan moieties sialyl-Lewis-X and/or -A (sLeX/A) are the primary ligands for E-selectin, regulating subsequent tumor cell extravasation into distant organs. However, the nature of the glycoprotein scaffolds displaying these glycans in breast cancer remains unclear and constitutes the focus of the present investigation.Methods
We isolated glycoproteins that bind E-selectin from the CF1_T breast cancer cell line, derived from a patient with ductal carcinoma. Proteins were identified using bottom-up proteomics approach by nanoLC-orbitrap LTQ-MS/MS. Data were curated using bioinformatics tools to highlight clinically relevant glycoproteins, which were validated by flow cytometry, Western blot, immunohistochemistry and in-situ proximity ligation assays in clinical samples.Results
We observed that the CF1_T cell line expressed sLeX, but not sLeA and the E-selectin reactivity was mainly on N-glycans. MS and bioinformatics analysis of the targeted glycoproteins, when narrowed down to the most clinically relevant species in breast cancer, identified CD44 glycoprotein (HCELL) and CD13 as key E-selectin ligands. Additionally, the co-expression of sLeX-CD44 and sLeX-CD13 was confirmed in clinical breast cancer tissue samples.Conclusions
Both CD44 and CD13 glycoforms display sLeX in breast cancer and bind E-selectin, suggesting a key role in metastasis development. Such observations provide a novel molecular rationale for developing targeted therapeutics.General significance
While HCELL expression in breast cancer has been previously reported, this is the first study indicating that CD13 functions as an E-selectin ligand in breast cancer. This observation supports previous associations of CD13 with metastasis and draws attention to this glycoprotein as an anti-cancer target. 相似文献7.
8.
Huaidong Zhang Qin Li Lina Wang Yan Chen 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(5):1199-1208
Background
Alcohol dehydrogenases (ADHs) catalyze the reversible oxidation of alcohol using NAD+ or NADP+ as cofactor. Three ADH homologues have been identified in Komagataella phaffii GS115 (also named Pichia pastoris GS115), ADH1, ADH2 and ADH3, among which adh3 is the only gene responsible for consumption of ethanol in Komagataella phaffii GS115. However, the relationship between structure and function of mitochondrial alcohol dehydrogenase isozyme III from Komagataella phaffii GS115 (KpADH3) is still not clear yet.Methods
KpADH3 was purified, identified and characterized by multiple biophysical techniques (Nano LC-MS/MS, Enzymatic activity assay, X-ray crystallography).Results
The crystal structure of KpADH3, which was the first ADH structure from Komagataella phaffii GS115, was solved at 1.745?Å resolution. Structural analysis indicated that KpADH3 was the sole dimeric ADH structure with face-to-face orientation quaternary structure from yeast. The major structural different conformations located on residues 100–114 (the structural zinc binding loop) and residues 337–344 (the loop between α12 and β15 which covered the catalytic domain). In addition, three channels were observed in KpADH3 crystal structure, channel 2 and channel 3 may be essential for substrate specific recognition, ingress and egress, channel 1 may be the pass-through for cofactor.Conclusions
KpADH3 plays an important role in the metabolism of alcohols in Komagataella phaffii GS115, and its crystal structure is the only dimeric medium-chain ADH from yeast described so far.General significance
Knowledge of the relationship between structure and function of KpADH3 is crucial for understanding the role of KpADH3 in Komagataella phaffii GS115 mitochondrial metabolism. 相似文献9.
Bin Zhang Ronne Wee Yeh Yeo Ruenn Chai Lai Eugene Wei Kian Sim Keh Chuang Chin Sai Kiang Lim 《Cytotherapy》2018,20(5):687-696
Background aims
The immunomodulatory property of mesenchymal stromal cell (MSC) exosomes is well documented. On the basis of our previous report that MSC exosomes increased regulatory T-cell (Treg) production in mice with allogenic skin graft but not in ungrafted mice, we hypothesize that an activated immune system is key to exosome-mediated Treg production.Methods
To test our hypothesis, MSC exosomes were incubated with mouse spleen CD4+ T cells that were activated with either anti-CD3/CD28 mAbs or allogenic antigen-presenting cell (APC)-enriched spleen CD11c+ cells to determine whether production of mouse CD4+CD25+ T cells or CD4+CD25+Foxp3+ Tregs could be induced. MSC exosomes were also administered to the lethal chimeric human-SCID mouse model of graft-versus-host disease (GVHD) in which human peripheral blood mononuclear cells were infused into irradiated NSG mice to induce GVHD.Results
We report here that MSC exosome–induced production of CD4+CD25+ T cells or CD4+CD25+Foxp3+ Tregs from CD4+ T cells activated by allogeneic APC-enriched CD11C+ cells but not those activated by anti-CD3/CD28 mAbs. This induction was exosome- and APC dose–dependent. In the mouse GVHD model in which GVHD was induced by transplanted human APC-stimulated human anti-mouse CD4+ T cell effectors, MSC exosome alleviated GVHD symptoms and increased survival. Surviving exosome-treated mice had a significantly higher level of human CD4+CD25+CD127low/– Tregs than surviving mice treated with Etanercept, a tumor necrosis factor inhibitor.Conclusions
MSC exosome enhanced Treg production in vitro and in vivo through an APC-mediated pathway. 相似文献10.
Cristina Pérez-Arnaiz Natalia Busto Javier Santolaya José M. Leal Giampaolo Barone Begoña García 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(3):522-531
Background
Stabilization of G-quadruplex helices by small ligands has attracted growing attention because they inhibit the activity of the enzyme telomerase, which is overexpressed in > 80% cancer cells. TMPyP4, one of the most studied G-quadruplex ligands, is used as a model to show that the ligands can exhibit different binding features with different conformations of a human telomeric specific sequence.Methods
UV–Vis, FRET melting Assay, Isothermal Titration Calorimetry, Time-resolved Fluorescence lifetime, T-Jump and Molecular Dynamics.Results
TMPyP4 yields two different complexes with two Tel22 telomeric conformations in the presence of Na+ or K+. T-Jump kinetic experiments show that the rates of formation and dissociation of these complexes in the ms time scale differ by one order of magnitude. MD simulations reveal that, in K+ buffer, “hybrid 1” conformation yields kinetic constants on interaction with TMPyP4 one order lower than “hybrid 2”. The binding involves π–π stacking with external loop bases.Conclusions
For the first time we show that for a particular buffer TMPyP4 interacts in a kinetically different way with the two Tel22 conformations even if the complexes formed are thermodynamically indistinguishable.General significance
G-quadruplexes, endowed with technological applications and potential impact on regulation mechanisms, define a new research field. The possibility of building different conformations from same sequence is a complex issue that confers G-quadruplexes very interesting features. The obtaining of reliable kinetic data constitutes an efficient tool to determine reaction mechanisms between conformations and small molecules. 相似文献11.
Sonia Bustamante Tharusha Jayasena Dulama Richani Robert Bruce Gilchrist Lindsay E. Wu David A. Sinclair Perminder Singh Sachdev Nady Braidy 《Metabolomics : Official journal of the Metabolomic Society》2018,14(1):15
Introduction
Nicotinamide adenine dinucleotide (NAD+) is an essential pyridine nucleotide that serves as a key hydride transfer coenzyme for several oxidoreductases. It is also the substrate for intracellular secondary messenger signalling by CD38 glycohydrolases, DNA repair by poly(adenosine diphosphate ribose) polymerase, and epigenetic regulation of gene expression by a class of histone deacetylase enzymes known as sirtuins. The measurement of NAD+ and its related metabolites (hereafter, the NAD+ metabolome) represents an important indicator of cellular function.Objectives
A study was performed to develop a sensitive, selective, robust, reproducible, and rapid method for the concurrent quantitative determination of intracellular levels of the NAD+ metabolome in glial and oocyte cell extracts using liquid chromatography coupled to mass spectrometry (LC/MS/MS).Methods
The metabolites were separated on a versatile amino column using a dual HILIC-RP gradient with heated electrospray (HESI) tandem mass spectrometry detection in mixed polarity multiple reaction monitoring mode.Results
Quantification of 17 metabolites in the NAD+ metabolome in U251 human astroglioma cells could be achieved. Changes in NAD+ metabolism in U251 cell line, and murine oocytes under different culture conditions were also investigated.Conclusion
This method can be used as a sensitive profiling tool, tailoring chromatography for metabolites that express significant pathophysiological changes in several disease conditions and is indispensable for targeted analysis.12.
Background
Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is a key enzyme of the glycolytic pathway, reversibly catalyzing the sixth step of glycolysis and concurrently reducing the coenzyme NAD+ to NADH. In photosynthetic organisms a GAPDH paralog (Gap2 in Cyanobacteria, GapA in most photosynthetic eukaryotes) functions in the Calvin cycle, performing the reverse of the glycolytic reaction and using the coenzyme NADPH preferentially. In a number of photosynthetic eukaryotes that acquired their plastid by the secondary endosymbiosis of a eukaryotic red alga (Alveolates, haptophytes, cryptomonads and stramenopiles) GapA has been apparently replaced with a paralog of the host’s own cytosolic GAPDH (GapC1). Plastid GapC1 and GapA therefore represent two independent cases of functional divergence and adaptations to the Calvin cycle entailing a shift in subcellular targeting and a shift in binding preference from NAD+ to NADPH.Methods
We used the programs FunDi, GroupSim, and Difference Evolutionary-Trace to detect sites involved in the functional divergence of these two groups of GAPDH sequences and to identify potential cases of convergent evolution in the Calvin-cycle adapted GapA and GapC1 families. Sites identified as being functionally divergent by all or some of these programs were then investigated with respect to their possible roles in the structure and function of both glycolytic and plastid-targeted GAPDH isoforms.Conclusions
In this work we found substantial evidence for convergent evolution in GapA/B and GapC1. In many cases sites in GAPDHs of these groups converged on identical amino acid residues in specific positions of the protein known to play a role in the function and regulation of plastid-functioning enzymes relative to their cytosolic counterparts. In addition, we demonstrate that bioinformatic software like FunDi are important tools for the generation of meaningful biological hypotheses that can then be tested with direct experimental techniques. 相似文献13.
Jizhong Wang Chengli Yang Xing Chen Bingxin Bao Xuan Zhang Dali Li Xingfan Du Ruofu Shi Junfang Yang Ronghui Zhu 《Biotechnology letters》2016,38(8):1315-1320
Objectives
To find an efficient and cheap system for NAD+ regenerationResults
A NADH-ferricyanide dehydrogenase was obtained from an isolate of Escherichia coli. Optimal activity of the NADH dehydrogenase was at 45 °C and pH 7.5, with a K m value for NADH of 10 μM. By combining the NADH dehydrogenase, potassium ferricyanide and laccase, a bi-enzyme system for NAD+ regeneration was established. The system is attractive in that the O2 consumed by laccase is from air and the sole byproduct of the reaction is water. During the reaction process, 10 mM NAD+ was transformed from NADH in less than 2 h under the condition of 0.5 U NADH dehydrogenase, 0.5 U laccase, 0.1 mM potassium ferricyanide at pH 5.6, 30 °CConclusion
The bi-enzyme system employed the NADH-ferricyanide dehydrogenase and laccase as catalysts, and potassium ferricyanide as redox mediator, is a promising alternative for NAD+ regeneration.14.
Objectives
To use permeabilized cells of the fission yeast, Schizosaccharomyces pombe, that expresses human UDP-glucose 6-dehydrogenase (UGDH, EC 1.1.1.22), for the production of UDP-glucuronic acid from UDP-glucose.Results
In cell extracts no activity was detected. Therefore, cells were permeabilized with 0.3 % (v/v) Triton X-100. After washing away all low molecular weight metabolites, the permeabilized cells were directly used as whole cell biocatalyst. Substrates were 5 mM UDP-glucose and 10 mM NAD+. Divalent cations were not added to the reaction medium as they promoted UDP-glucose hydrolysis. With this reaction system 5 mM UDP-glucose were converted into 5 mM UDP-glucuronic acid within 3 h.Conclusions
Recombinant permeabilized cells of S. pombe can be used to synthesize UDP-glucuronic acid with 100 % yield and selectivity.15.
Yejin Lee Byungho Lim Seon Woo Lee Woo Rin Lee Yong-In Kim Minhyeok Kim Hyoungseok Ju Mi Young Kim Suk-Jo Kang Ji-Joon Song J. Eugene Lee Changwon Kang 《生物化学与生物物理学报:疾病的分子基础》2018,1864(10):3145-3153
Background
Human ANKRD9 (ankyrin repeat domain 9) expression is altered in some cancers.Methods
We tested genetic association of ANKRD9 with gastric cancer susceptibility and examined functional association of ANKRD9 with altered proliferation of MKN45 gastric cancer cells. We then identified ANKRD9-binding partners in HEK 293 embryonic kidney cells using quantitative proteomics, western blotting and complex reconstitution assays. We finally demonstrated ANKRD9's role of recognizing substrates for ubiquitination using in vitro ubiquitylation assay.Results
ANKRD9 is associated with cancer susceptibility in a comparison of single-nucleotide polymorphisms between 1092 gastric cancer patients and 1206 healthy controls. ANKRD9 depletion accelerates tumor progression by increasing cellular proliferation, piling up, and anchorage-independent growth of MKN45 cells. We discovered that ANKRD9 is a ubiquitin ligase substrate receptor subunit and has an anti-proliferative activity. ANKRD9 associates with CUL5 (not CUL2), ELOB, ELOC, and presumably RNF7 subunits, which together assemble into a cullin-RING superfamily E3 ligase complex. ANKRD9 belongs to the ASB family of proteins, which are characterized by the presence of ankyrin repeats and a SOCS box. In addition to its interactions with the other E3 ligase subunits, ANKRD9 interacts with two isoforms of inosine monophosphate dehydrogenase (IMPDH). These IMPDH isoforms are cognate substrates of the ANKRD9-containing E3 enzyme, which ubiquitinates them for proteasomal degradation. Their ubiquitination and turnover require the presence of ANKRD9.Conclusion
ANKRD9, a previously unidentified E3 substrate receptor subunit, functions in tumor suppression by recognizing the oncoprotein IMPDH isoforms for E3 ubiquitination and proteasomal degradation. 相似文献16.
Dunja Urbančič Anita Kotar Alenka Šmid Marko Jukič Stanislav Gobec Lars-Göran Mårtensson Janez Plavec Irena Mlinarič-Raščan 《Biochimica et Biophysica Acta (BBA)/General Subjects》2019,1863(1):182-190
Background
Methylation driven by thiopurine S-methylatransferase (TPMT) is crucial for deactivation of cytostatic and immunosuppressant thiopurines. Despite its remarkable integration into clinical practice, the endogenous function of TPMT is unknown.Methods
To address the role of TPMT in methylation of selenium compounds, we established the research on saturation transfer difference (STD) and 77Se NMR spectroscopy, fluorescence measurements, as well as computational molecular docking simulations.Results
Using STD NMR spectroscopy and fluorescence measurements of tryptophan residues in TPMT, we determined the binding of selenocysteine (Sec) to human recombinant TPMT. By comparing binding characteristics of Sec in the absence and in the presence of methyl donor, we confirmed S-adenosylmethionine (SAM)-induced conformational changes in TPMT. Molecular docking analysis positioned Sec into the active site of TPMT with orientation relevant for methylation reaction. Se-methylselenocysteine (MeSec), produced in the enzymatic reaction, was detected by 77Se NMR spectroscopy. A direct interaction between Sec and SAM in the active site of rTPMT and the formation of both products, MeSec and S-adenosylhomocysteine, was demonstrated using NMR spectroscopy.Conclusions
The present study provides evidence on in vitro methylation of Sec by rTPMT in a SAM-dependant manner.General significance
Our results suggest novel role of TPMT and demonstrate new insights into enzymatic modifications of the 21st amino acid. 相似文献17.
Viacheslav V. Senichkin Gelina S. Kopeina Eugeniia A. Prokhorova Alexey V. Zamaraev Inna N. Lavrik Boris Zhivotovsky 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(3):557-566
Background
The development of approaches that increase therapeutic effects of anti-cancer drugs is one of the most important tasks of oncology. Caloric restriction in vivo or serum deprivation (SD) in vitro has been shown to be an effective tool for sensitizing cancer cells to chemotherapeutic drugs. However, the detailed mechanisms underlying the enhancement of apoptosis in cancer cells by SD remain to be elucidated.Methods
Flow cytometry, caspase activity assay and western blotting were used for cell death rate evaluation. Western blotting, gel-filtration, siRNA approach and qRT-PCR were used to elucidate the mechanism underlying cell death potentiation upon SD.Results
We demonstrated that SD sensitizes cancer cells to treatment with chemotherapeutic agent cisplatin. This effect is independent on activation of caspases-2 and -8, apical caspases triggering apoptosis in response to genotoxic stress. SD potentiates cell death via downregulation of the anti-apoptotic protein Mcl-1. In fact, SD reduces the Mcl-1 mRNA level, which consequently decreases the Mcl-1 protein level and renders cells more susceptible to apoptosis induction via the formation of apoptosome.Conclusions
Mcl-1 protein is an important regulator of sensitivity of cancer cells to apoptotic stimuli upon SD.General significance
This study identifies Mcl-1 as a new target for the sensitization of human cancer cells to cell death by SD, which is of great significance for the development of efficient anti-cancer therapies. 相似文献18.
Xu Lu Dongmei Zhang Hayato Shoji Chengwei Duan Guowei Zhang Tomoya Isaji Yuqin Wang Tomohiko Fukuda Jianguo Gu 《Biochimica et Biophysica Acta (BBA)/General Subjects》2019,1863(3):598-608
Background
α1,6-Fucosyltransferase-deficient (Fut8?/?) mice displayed increased locomotion and schizophrenia-like behaviors. Since neuroinflammation is a common pathological change in most brain diseases, this study was focused on investigating the effects of Fut8 in microglia and astrocytes.Methods
Brain tissues were analyzed using immunohistochemical staining. Core fucosylation and protein expression were analyzed using lectin blot and western blot, respectively. Fut8-knockout (KO) cells were established by the CRISPR/Cas9 system.Results
The number of Iba-1 positive cells and GFAP positive cells were significantly increased in both untreated and lipopolysaccharide stimulated inflammatory conditional Fut8?/? mice by comparison with both wild-type (Fut8+/+) and hetero (Fut8+/?) mice. Stimulation with pro-inflammatory factors, such as IFN-γ and IL-6, induced expression levels of fucosylation in primary microglia and astrocytes, as well as in glial cell lines. Cell motility and iNOS expression were easily induced by IFN-γ in Fut8-KO BV-2 cells compared with wild-type (WT) cells. In a similar manner, both Fut8-KO C6 cells and primary astrocytes treated with 2-fluoro-L-fucose, a specific inhibitor for fucosylation, showed a higher response to IL-6-stimulated phospho-STAT3 signaling, compared with WT cells.Conclusions
Core fucosylation negatively regulates the states of neuroinflammation by modulating the sensitivity of microglia and astrocytes to inflammatory mediators. The disorders of Fut8?/? mice are caused not only by neurons but also by glial cell dysfunction.General significance
Core fucose is a novel regulator for neuroinflammation in the central nervous system. 相似文献19.
Patricia Santofimia-Castaño Bruno Rizzuti Olga Abián Adrián Velázquez-Campoy Juan L. Iovanna José L. Neira 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(6):1283-1295
Background
NUPR1 is a multifunctional intrinsically disordered protein (IDP) involved, among other functions, in chromatin remodelling, and development of pancreatic ductal adenocarcinoma (PDAC). It interacts with several biomolecules through hydrophobic patches around residues Ala33 and Thr68. The drug trifluoperazine (TFP), which hampers PDAC development in xenografted mice, also binds to those regions. Because of the large size of the hot-spot interface of NUPR1, small molecules could not be adequate to modulate its functions.Methods
We explored how amphipathic helical-designed peptides were capable of interacting with wild-type NUPR1 and the Thr68Gln mutant, inhibiting the interaction with NUPR1 protein partners. We used in vitro biophysical techniques (fluorescence, circular dichroism (CD), nuclear magnetic resonance (NMR) and isothermal titration calorimetry (ITC)), in silico studies (docking and molecular dynamics (MD)), and in cellulo protein ligation assays (PLAs) to study the interaction.Results
Peptide dissociation constants towards wild-type NUPR1 were ~ 3?μM, whereas no interaction was observed with the Thr68Gln mutant. Peptides interacted with wild-type NUPR1 residues around Ala33 and residues at the C terminus, as shown by NMR. The computational results clarified the main determinants of the interactions, providing a mechanism for the ligand-capture that explains why peptide binding was not observed for Thr68Gln mutant. Finally, the in cellulo assays indicated that two out of four peptides inhibited the interaction of NUPR1 with the C-terminal region of the Polycomb RING protein 1 (C-RING1B).Conclusions
Designed peptides can be used as lead compounds to inhibit NUPR1 interactions.General significance
Peptides may be exploited as drugs to target IDPs. 相似文献20.
Shruti Chakraborty Sayak Ganguli Aritra Chowdhury Michael Ibba Rajat Banerjee 《Biochimica et Biophysica Acta (BBA)/General Subjects》2018,1862(8):1801-1809