泛素蛋白磷酸化修饰的功能与解析

泛素化是存在于真核生物中一种重要的翻译后修饰过程,参与调控包括蛋白质降解在内的多种生命活动。实现这一调控过程需要将一个由76个氨基酸组成的泛素蛋白共价连接到底物蛋白上。同时,泛素本身也存在多种翻译后修饰,包括泛素化、磷酸化、乙酰化等,进一步丰富了泛素的修饰类型,决定了底物蛋白不同的命运。近年来,伴随着第65位丝氨酸磷酸化泛素蛋白参与调控线粒体自噬这一突破性进展,泛素蛋白其余磷酸化位点的功能研究也获得越来越多的关注。本文根据目前已有的国内外研究和报道,总结了泛素蛋白已知的磷酸化修饰位点,梳理了泛素蛋白第12位和66位苏氨酸、第57位和65位丝氨酸等位点的磷酸化修饰对其生物物理特性带来的改变,并对相应修饰位点所涉及的生物学功能调控进行了综述。     

Effect of Axial Ligand Length on Biological and Anticancer Properties of Axially Disubstituted Silicon Phthalocyanines

In this study, three new axially disubstituted silicon phthalocyanines ( SiPc1–3 ) and their quaternized phthalocyanine derivatives ( QSiPc1–3 ) were prepared and characterized. The biological properties (antioxidant, antimicrobial, antibiofilm, and microbial cell viability activities) of the water-soluble silicon phthalocyanines were examined, as well. A 1 % DMSO diluted with pure water was used as a solvent in biological activity studies. All the compounds exhibited high antioxidant activity. They displayed efficient antimicrobial and antimicrobial photodynamic therapeutic properties against various microorganisms, especially Gram (+) bacteria. Additionally, they demonstrated high antibiofilm activities against S. aureus and P. aeruginosa. In addition, 100 % bacterial reduction was obtained for all the studied phthalocyanines against E. coli viable cells. Besides, the DNA cleavage and binding features of compounds ( QSiPc1–3 ) were studied using pBR322 DNA and CT-DNA, respectively. Furthermore, the human topoisomerase I enzyme inhibition activities of compounds QSiPc1 – 3 were studied. Anticancer properties of the water-soluble compounds were investigated using cell proliferation MTT assay. They exhibited anticarcinogenic activity against the human colon cancer cell line (DLD-1). Compounds QSiPc1 and QSiPc3 displayed a high anticarcinogenic effect on the DLD-1 cell line. The obtained results indicated that all the studied compounds may be effective biological agents and anticancer drugs after further investigations.     

Structure-Activity Relationship of Methyl 4-Aminobenzoate Derivatives as Being Drug Candidate Targeting Glutathione Related Enzymes: in Vitro and in Silico Approaches

A thiol compound, glutathione, is essential for healthy cell defence against xenobiotics and oxidative stress. Glutathione reductase (GR) and glutathione S-transferase (GST) are two glutathione-related enzymes that function in the antioxidant and the detoxification systems. In this study, potential inhibitory effects of methyl 4-aminobenzoate derivatives on GR and GST were examined in vitro. GR and GST were isolated from human erythrocytes with 7.63 EU/mg protein and 5.66 EU/mg protein specific activity, respectively. It was found that compound 1 (methyl 4-amino-3-bromo-5-fluorobenzoate with K_i value of 0.325±0.012 μM) and compound 5 (methyl 4-amino-2-nitrobenzoate with K_i value of 92.41±22.26 μM) inhibited GR and GST stronger than other derivatives. Furthermore, a computer-aided method was used to predict the binding affinities of derivatives, ADME characteristics, and toxicities. Derivatives 4 (methyl 4-amino-2-bromobenzoate) and 6 (methyl 4-amino-2-chlorobenzoate) were estimated to have the lowest binding energies into GR and GST receptors, respectively according to results of in silico studies.     

In Silico Design of an Oseltamivir Derivative with Increased Affinity against Wild-Type and Mutant Variants of Neuraminidase and Hemagglutinin of Influenza A H1N1 Virus

Antiviral resistance has turned into a world concern nowadays. Influenza A H1N1 emerged as a problem at the world level due to the neuraminidase (NA) mutations. The NA mutants conferred resistance to oseltamivir and zanamivir. Several efforts were conducted to develop better anti-influenza A H1N1 drugs. Our research group combined in silico methods to create a compound derived from oseltamivir to be tested in vitro against influenza A H1N1. Here we show the results of a new compound derived from oseltamivir but with specific chemical modifications, with significant affinity either on NA (in silico and in vitro assays) or HA (in silico) from influenza A H1N1 strain. We include docking and molecular dynamics (MD) simulations of the oseltamivir derivative at the binding site onto NA and HA of influenza A H1N1. Additionally, the biological experimental results show that oseltamivir derivative decreases the lytic-plaque formation on viral susceptibility assays, and it does not show cytotoxicity. Finally, oseltamivir derivative assayed on viral NA showed a concentration-dependent inhibition behavior at nM, depicting a high affinity of the compound for the enzyme, corroborated with the MD simulations results, placing our designed oseltamivir derivative as a potential antiviral against influenza A H1N1.     

HPLC-DAD Analysis and Versatile Bioactivities of Turkish Sunflower Honeys Using Chemometric Approaches

Sunflower honey (SH) is bright yellow, fragrant, pollen-flavoured, slightly herbaceous and has a unique taste. The present research aims to examine the enzyme inhibitory, antioxidant, anti-inflammatory, antimicrobial and anti-quorum sensing activities and phenolic compositions of 30 sunflower honeys (SHs) produced from several regions of Turkey with chemometric study. SAH from Samsun exhibited the best antioxidant activity in β-carotene linoleic acid (IC₅₀: 7.33±0.17 mg/mL) and CUPRAC (A_0.50: 4.94±0.13 mg/mL) assays, anti-urease activity (60.63±0.87 %) and anti-inflammatory activity against COX-1 (73.94±1.08 %) and COX-2 (44.96±0.85 %). SHs exhibited mild antimicrobial activity against the test microorganisms while they showed high quorum sensing inhibition zones measured in the range of 42–52 mm against the CV026 strain. The phenolic composition was determined by high performance liquid chromatography with diode array detection (HPLC-DAD) system and levulinic, gallic, p-hydroxybenzoic, vanillic and p-coumaric acids were identified in all studied SHs. The classification of SHs was performed the using PCA and HCA. This study revealed that phenolic compounds and biological properties are effective in classification of SHs according to their geographical origin. The results suggest that studied SHs could be valued as potential agents with versatile bioactivities in oxidative stress-related disease, microbial infections, inflammation, melanoma, and peptic ulcer.     

Energy Metabolism During Late Gestation and Lactation in Muciparous Sows in Relation to Backfat Thickness and the Interval from Weaning to First Oestrus

Ten crossbred, fourth or fifth parity sows were divided into 2 groups - high (H) and low (L) - according to their backfat thickness 9 days before parturition. Body weight, backfat thickness and litter weight were recorded repeatedly during a 5 week lactation period. The length of the interval from weaning to first oestrus was also noted. All sows were fed a commercial diet (11.9 MJ/kg, 14.5% crude protein). During gestation, daily food intake was 2.2 kg/sow, while during lactation it was 3.0 kg/sow plus 0.4 kg/piglet. Blood samples were drawn on day 9 before parturition and on days 2,7,14 and 21 of lactation. The samples were analysed to determine concentrations of glucose, urea nitrogen, creatinine, triglycerides, free fatty acids and beta-hydroxybutyric acid. In both groups, concentrations of free fatty acids and urea nitrogen were low on day 9 before parturition while those of triglycerides were high, indicating anabolism regardless of backfat thickness. During the first week of lactation, concentrations of free fatty acids increased in the H-group but not in the L-group, and concentrations of urea nitrogen were higher in the H-group. These differences, together with the greater loss of weight observed in the H-group, indicate that catabolism of maternal fat and protein depots was more pronounced in the Η-group than in the L-group during this time. On day 14 of lactation, both groups showed equally low concentrations of free fatty acids, decreasing creatinine concentrations and stable triglyceride and urea nitrogen concentrations. Furthermore, weight loss during the second and third weeks of lactation was low in both groups. These facts, taken together, indicate that the catabolic rate was decreasing in both groups during this period. No differences in return to oestrus interval were noted between the groups. The present study indicates that under a restricted feeding regime the catabolic rate during the first week of lactation is higher in sows with higher backfat thickness in late gestation. As lactation progresses, a more balanced metabolism is achieved regardless of backfat thickness, which may tend to reduce differences in return to oestrous interval.     

Nutraceutical Activity of Anthocyanins from the Edible Berries of Rhamnus pompana

We present the inhibitory properties of the R. pompana anthocyanin fraction (RPAF) and its major constituents on alpha-glucosidase (AG), pancreatic lipase (PL), HMG-CoA reductase, and ornithine decarboxylase (ODC). The effect of RPAF was also evaluated in ICR male mice subjected to oral glucose tolerance test (OGTT) and hypercaloric/atherogenic diet for 30 days. RP-HPLC/MS profiling revealed that RPAF contained five major anthocyanins and induced slight inhibition on PL and HMG-CoA reductase (IC₅₀, 245–338 μg mL⁻¹) whereas strong activity on AG and ODC (IC₅₀, 130–133 μg mL⁻¹) was observed. Kinetic studies and molecular docking with pelargonidin-3-O-rutinoside (P3R) on ODC, revealed changes in K_m (0.9514–0.9746 mM) and V_max (1.96–2.32 μmol mg⁻¹ min⁻¹) suggesting mixed inhibition and molecular interaction with two active sites of ODC. P3R showed antiproliferative activity (IC_50, 46.5 μM) and decreased polyamine accumulation in DLD-1 cells. The results of OGTT confirmed that RPAF regulates postprandial glucose levels in diabetic animals which experienced a significant glucose depletion (30 %; p<0.001) from 30 to 120 min post-treatment. Prolonged supplementation of RPAF caused significant decrease (p<0.001) in plasma glucose, total cholesterol, LDL-c and triglycerides as well as significant increase (p<0.001) of HDL-c compared with normoglycemic untreated animals.     

A chromosome-level genome assembly enables the identification of the follicule stimulating hormone receptor as the master sex-determining gene in the flatfish Solea senegalensis

Sex determination (SD) shows huge variation among fish and a high evolutionary rate, as illustrated by the Pleuronectiformes (flatfishes). This order is characterized by its adaptation to demersal life, compact genomes and diversity of SD mechanisms. Here, we assembled the Solea senegalensis genome, a flatfish of great commercial value, into 82 contigs (614 Mb) combining long- and short-read sequencing, which were next scaffolded using a highly dense genetic map (28,838 markers, 21 linkage groups), representing 98.9% of the assembly. Further, we established the correspondence between the assembly and the 21 chromosomes by using BAC-FISH. Whole genome resequencing of six males and six females enabled the identification of 41 single nucleotide polymorphism variants in the follicle stimulating hormone receptor (fshr) consistent with an XX/XY SD system. The observed sex association was validated in a broader independent sample, providing a novel molecular sexing tool. The fshr gene displayed differential expression between male and female gonads from 86 days post-fertilization, when the gonad is still an undifferentiated primordium, concomitant with the activation of amh and cyp19a1a, testis and ovary marker genes, respectively, in males and females. The Y-linked fshr allele, which included 24 nonsynonymous variants and showed a highly divergent 3D protein structure, was overexpressed in males compared to the X-linked allele at all stages of gonadal differentiation. We hypothesize a mechanism hampering the action of the follicle stimulating hormone driving the undifferentiated gonad toward testis.     

中国东北地区三种蜜环菌的生物学特性及奥氏蜜环菌人工培养

蜜环菌属Armillaria真菌具有较高的食药用价值。由于蜜环菌的生长发育过程较复杂,还未完全实现商业化栽培,野生资源的供应受到季节性和地域性的影响。本研究以采自东北地区蜜环菌属的3个菌株为研究对象,通过培养物的形态特征及分子标记确定菌株JG19016为奥氏蜜环菌A. ostoyae,菌株JG19017为高卢蜜环菌A. gallica,菌株JG19018为中国蜜环菌生物种C。奥氏蜜环菌JG19016最适生长温度为25 ℃,高卢蜜环菌JG19017的最适生长温度为22 ℃,中国蜜环菌生物种C JG19018则在22-25 ℃时菌丝生长速度最快;3个菌株最适pH为5-6。奥氏蜜环菌JG19016对葡萄糖和蔗糖利用率较好,高卢蜜环菌JG19017对葡萄糖利用率较好,中国蜜环菌生物种C JG19018对葡萄糖和淀粉利用率较好;蛋白胨对3个菌株促进作用最强,为最适氮源。培养基中加入VB₁,对3个菌株的菌丝生长均有明显的促进作用。奥氏蜜环菌JG19016菌丝生长的最优培养基配方为：葡萄糖20 g,蛋白胨3 g,磷酸二氢钾2 g,硫酸镁1.5 g,VB₁ 10 mg,琼脂20 g,水1 L。在木屑基质中培养,其配方的最优碳氮比为38:1,最佳木屑粗细比为3:1以上。出菇条件探索结果显示,菌丝及菌索长满菌袋(17 mm×33 mm×5 mm丝聚乙烯袋)需要50-60 d,之后在18 ℃、60%湿度和12 h散射光的环境中,10 d左右可观察到原基产生。增加菇房湿度到90%-95%,2-3 d可观察到1-3 cm的幼子实体,7 d左右菌柄和菌盖完全分化,10 d左右观察到菌盖展开。     

Genome-wide RAD sequencing data suggest predominant role of vicariance in Sino-Japanese disjunction of the monotypic genus Conandron (Gesneriaceae)

Disjunct distribution is a key issue in biogeography and ecology, but it is often difficult to determine the relative roles of dispersal vs. vicariance in disjunctions. We studied the phylogeographic pattern of the monotypic Conandron ramondioides (Gesneriaceae), which shows Sino-Japanese disjunctions, with ddRAD sequencing based on a comprehensive sampling of 11 populations from mainland China, Taiwan Island, and Japan. We found a very high degree of genetic differentiation among these three regions, with very limited gene flow and a clear Isolation by Distance pattern. Mainland China and Japan clades diverged first from a widespread ancestral population in the middle Miocene, followed by a later divergence between mainland China and Taiwan Island clades in the early Pliocene. Three current groups have survived in various glacial refugia during the Last Glacial Maximum, and experienced contraction and/or bottlenecks since their divergence during Quaternary glacial cycles, with strong niche divergence between mainland China + Japan and Taiwan Island ranges. Thus, we verified a predominant role of vicariance in the current disjunction of the monotypic genus Conandron. The sharp phylogenetic separation, ecological niche divergence among these three groups, and the great number of private alleles in all populations sampled indicated a considerable time of independent evolution, and suggests the need for a taxonomic survey to detect potentially overlooked taxa.