Human Insulin (HI) is a well-characterized natural hormone which regulates glycose levels into the blood-stream and is widely used for diabetes treatment. Numerous studies have manifested that despite significant efforts devoted to structural characterization of this molecule and its complexes with organic compounds (ligands), there is still a rich diagram of phase transitions and novel crystalline forms to be discovered. Towards the improvement of drug delivery, identification of new insulin polymorphs from polycrystalline samples, simulating the commercially available drugs, is feasible today via macromolecular X- ray powder diffraction (XRPD). This approach has been developed and is considered as a respectable method, which can be employed in biosciences for various purposes such as observing phase transitions and characterizing bulk pharmaceuticals. An overview of structural studies on human insulin complexes performed over the past decade employing both synchrotron and laboratory sources for XRPD measurements, is reported herein. This review aims to assemble all recent advances in diabetes treatment field in terms of drug formulation, verifying in parallel the efficiency and applicability of protein XRPD for quick and accurate preliminary structural characterization in large scale.

This review focuses on the regulation and modulation of human DNA polymerase δ (Pol δ). The emphasis is on mechanisms that regulate the activity and properties of Pol δ in DNA repair and replication. The areas covered are the degradation of the p12 subunit of Pol δ, which converts it from a heterotetramer (Pol δ4) to a heterotrimer (Pol δ3), in response to DNA damage and also during the cell cycle. The biochemical mechanisms that lead to degradation of p12 are reviewed, as well as the properties of Pol δ4 and Pol δ3 that provide insights into their functions in DNA replication and repair. The second focus of the review involves the functions of two Pol δ binding proteins, PDIP46 and PDIP38, both of which are multi-functional proteins. PDIP46 is a novel activator of Pol δ4, and the impact of this function is discussed in relation to its potential roles in DNA replication. Several new models for the roles of Pol δ3 and Pol δ4 in leading and lagging strand DNA synthesis that integrate a role for PDIP46 are presented. PDIP38 has multiple cellular localizations including the mitochondria, the splicesosomes and the nucleus. It has been implicated in a number of cellular functions, including the regulation of specialized DNA polymerases, mitosis, the DNA damage response, Mdm2 alternative splicing and the regulation of the Nox4 NADPH oxidase.

Foods are mixed with saliva in the oral cavity and swallowed. During staying in the stomach, saliva is contentiously provided to mix with the ingested foods. Because a salivary component nitrite is protonated to produce active nitrous acid at acidic pH, the redox reactions of nitrous acid with phenolic compounds in foods become possible in the stomach. In the reactions, nitrous acid is reduced to nitric oxide (•NO), producing various products from phenolic compounds. In the products, stable hydroxybezoyl benzofuranone derivatives, which are produced from quercetin and its 7-O-glucoside, are included. Caffeic acid, chlorogenic acid, and rutin are oxidized to quinones and the quinones can react with thiocyanic acid derived from saliva producing stable oxathiolone derivatives. 6,8-Dinitrosocatechis are produced from catechins by the redox reaction, and the dinitrocatechins are oxidized further by nitrous acid producing the quinones, which can make charge transfer complexes with the dinitrosocatechin and can react with thiocyanic acid producing the stable thiocyanate conjugates. In this way, various products can be produced by the reactions of salivary nitrite with dietary phenolic compounds, and reactive and toxic quinones formed by the reactions are postulated to be removed in the stomach by thiocyanic acid derived from saliva.

Proteins are the workhorses of the cell that perform biological functions by interacting with other proteins. Many statistical methods for protein-protein interaction (PPI) have been studied without considering time-dependent changes in networks and the functionalities. I introduced a novel method that models PPI networks as being dynamic in nature and evolving time-varying multivariate distribution with Conditional Random Fields (CRF). This research is directed towards implementing this new combinatorial algorithm on massively parallel architectures such as Graphics Processing Units (GPUs) for efficient computations for large scale bioinformatics datasets. I compared Conditional Random Fields (CRF) and the proposed novel method using CRF combined with the Block Coordinate Descent algorithm for human protein-protein interaction data set. Both are implemented on GPU-Accelerated Computing Architecture and the proposed novel method showed the advantages in predicting protein-protein interaction sites. I also show that the proposed approach is more efficient in 6.13% than standalone CRF++ in predicting protein-protein interaction sites.

Gliomas are one of the most aggressive and treatment-resistant types of human cancer. One of the most promising field in gliomas cancer therapy is identification and evaluation of anticancer properties of compounds found in plants i.a. naringenin (N) and 8-prenylnaringenin (8PN). The prenyl group seem to be crucial to the anticancer activity of flavones, which may lead to enhanced cell membrane targeting and thus increased intracellular activity. Unfortunately, 8PN content in hop cones is from 10 to 100 times lower compared to other flavonoids i.e. xanthohumol. In this study we used a simple method for the synthesis of 8PN from isoxanthohumol, via O-demethylation with high, 97% of the isolated yield. Cellular accumulation and cytotoxicity of naringenin and 8-prenylnaringenin in normal (BJ) and cancer cells (U-118 MG) were also examined. Obtained data indicated that 8-prenylnaringenin exhibited higher toxicity against used cell lines than naringenin and both flavones inhibited stronger glioblastoma U-118 MG cells than normal fibroblasts. The anticancer properties of 8PN correlated with its significantly greater (37%), accumulation in glioblastoma cells than in normal fibroblasts. Additionally, naringenin indicated higher selectivity for glioblastoma as it was over 6 times more toxic for cancer than normal cells. Our results provide evidence that examined prenylated and non-prenylated flavanones have different biological activity against normal and cancer cell lines and this phenomenon may be useful in clinical practice to construct new, anticancer drugs for glioblastoma.

The development of new diabetes drugs continues to be explored. Loureirin B, a flavonoid, extracted from Dracaena cochinchinensis, has been confirmed to increase insulin secretion and decrease blood glucose levels. For understanding the mechanism, a series of experiments had been employed based on computational methods and cell experiments. The insulin secretion significantly increased with the incubation of 0.01μM loureirin B for 4 hours. The viability of Ins-1 cells showed no significant difference with the treatment of loureirin B. Through computational methods, we hypothesized that loureirin B could interacts with K_ATP channels to promote insulin secretion. In cell experiments, it could be found that the current of K_ATP channel of Ins-1 cells was inhibited by the effect of loureirin B. After then, the voltage-dependent calcium channels were activated, the increase of Cx43 protein expression might mediate the Ca²⁺ to the intracellular. In summary, it could be concluded that loureirin B promoted insulin secretion mainly through inhibiting K_ATP current, the influx of Ca²⁺ to the Intracellular and the expression of Cx43.

Cholangiocarcinoma is an epithelial malignancy arising in the region between the intrahepatic bile ducts and the ampulla of Vater at the distal end of the common bile duct. The effect of current chemotherapy regimens against cholangiocarcinoma is limited, and the prognosis of patients with cholangiocarcinoma is poor. Aberrant DNA methylation and histone modification induce silencing of tumor suppressor genes and chromosomal instability during carcinogenesis. Studies have shown that the tumor suppressor genes and microRNAs (miRNAs) including MLH1, p14, p16, DAPK, miR-370 and miR-376c are frequently methylated in cholangiocarcinoma. Silencing of these tumor suppressor genes and miRNAs plays critical roles in the initiation and progression of cholangiocarcinoma. In addition, recent studies have demonstrated that DNA methylation inhibitors induce expression of endogenous retroviruses and exert the anti-tumor effect of via an anti-viral immune response. Aberrant DNA methylation of tumor suppressor genes and miRNAs could be a powerful biomarker for diagnosis and treatment of cholangiocarcinoma. Epigenetic therapy with DNA methylation inhibitors hold considerable promise for the treatment of cholangiocarcinoma through re-activation of tumor suppressor genes and miRNAs as well as induction of an anti-viral immune response.

The string of bacteria, Vibrio. sp. QD-5 utilizing alginate, was separated from rotten kelp. The results of genome sequencing showed that the strain QD-5 contained four alginate lyase genes.One of the alginate genes Aly-IV was cloned and linked to the plasmid pET-22b (+). The heterologous expressed alginate lyase aly-IVwas characterized，which possessed the theoretical molecular mass of 62 kDa, and theoretical isoelectric point (pI) of 5.12. - The enzyme aly-IV was purified and the activity reached 1256.78 U/mg, with optimal temperature of 35 ^oC and pH value of 8.9. Nurtured in the temperature below 25 ^oC for 30 minutes, the activity was almost stable. The result also suggested that the activity of enzyme was strongly affected by - NaCl whose optimal concentration was 15 mM in a lab environment The TLC and ESI-TOF-MS analysis suggested that the enzyme aly-IV could degrade sodium alginate and polyG in endo-lytic type, producing monomer, dimer and trimmer. So, aly-IV can also be widely applied to make large scale preparation of alginate oligosaccharides with low degree of polymerization (DP).

Vitamin A and E are important during pregnancy, the neonatal period, and childhood. The objective of this study was to assess whether maternal RRR-a-tocopherol supplementation affects serum and breast milk retinol. Serum was collected at baseline and twenty days later, and breast milk, at baseline, and on days 1, 7, and 20 after delivery. After the baseline serum collection, the supplemented group (n=16) received a single 400 IU of RRR-α-tocopherol. The control group (n=18) was only performed collections. Retinol and alpha tocopherol levels were determined by liquid chromatography. Serum retinol and alpha tocopherol at baseline and 20 days after delivery indicated proper vitamin A (> 20 µg/dL) and E (> 516 μg/dL) statuses in the control and supplemented groups (p > 0.05). Colostrum retinol levels on days 1 and 7 after delivery were significantly higher in the supplemented group (p = 0.018 and p = 0.012, respectively). Maternal vitamin E supplementation increased colostrum retinol by 52.23% and 111.2%, 24 hours and 7 days, respectively. However, retinol in mature milk did not differ between the groups (p > 0.05). In conclusion, the supplementation with 400 IU of RRR-α-tocopherol improved vitamin A bioavailability in breast milk.

G protein-coupled receptors (GPCRs) comprise the largest family of membrane receptors that control many cellular processes and consequently often serve as drug targets. These receptors undergo a strict regulation by mechanisms such as internalization and desensitization, which are strongly influenced by posttranslational modifications. Ubiquitination is a posttranslational modification with a broad range of functions that is currently gaining increased appreciation as a regulator of GPCR activity. The role of ubiquitination in directing GPCRs for lysosomal degradation has already been well-established. Furthermore, this modification can also play a role in targeting membrane and endoplasmic reticulum-associated receptors to the proteasome. Most recently, ubiquitination was also shown to be involved in GPCR signaling. In this review, we present current knowledge on the molecular basis of GPCR regulation by ubiquitination, and highlight the importance of E3 ubiquitin ligases, deubiquitinating enzymes and β-arrestins. Finally, we discuss classical and newly-discovered functions of ubiquitination in controlling GPCR activity.

Green Tea Catechins (GTCs) are a family of chemically related compounds usually classified as antioxidant molecules. Epidemiological evidences, supported by interventional studies, highlighted a more than promising role for GTCs in human Prostate Cancer (PCa) chemoprevention.In the last decades many efforts have been made to gain new insights into the mechanism of action of GTCs. Now it is clear that GTCs anticancer action can no longer be simplistically limited to their direct antioxidant/pro-oxidant properties. Recent contributions to the advancement of knowledge in this field have shown that GTCs specifically interact with cellular targets including, cell surface receptors, lipid rafts and endoplasmic reticulum, modulate gene expression through direct effect on transcription factors or indirect epigenetic mechanisms, interfere with intracellular proteostasis at various levels. Many of the effects observed in vitro are dose and cell context dependent and take place at concentration that cannot be achieved in vivo.Poor intestinal absorption together with an extensive systemic and enteric metabolism influence GTCs bioavailability through still poor understood mechanisms. Recent efforts to develop delivery systems that increase GTCs overall bioavailability, by mean of biopolymeric nanoparticles, represent the main way to translate preclinical results in a real clinical scenario for PCa chemoprevention.

The aim of this study was to examine the oxidation of human serum albumin (HSA) caused by oxidative stress after an exhaustive exercise such as ultra-marathon race. Thus, blood samples from 12 adult runners who underwent a 103 km mountain ultra-marathon race were collected pre- and 24, 48 and 72 h post race. HSA was partially purified using affinity chromatography and then was subjected to Western blot analysis for disulfide dimers determination, indicating oxidation. The results were correlated with those from a previous study in which the same samples were analyzed using different oxidative stress markers and a good correlation with protein carbonyls (PC) at all time points was observed. Moreover there was a significant correlation with static oxidation-reduction potential (sORP) at 24 h, and a negative correlation with capacity oxidation-reduction potential (cORP) at 24 and 48 h. In addition, an individual analysis of albumin dimers exhibited great inter-individual differences. This inter-individual variability in the oxidation of HSA may suggest different interventions (e.g. through diet) in order to confront the effects on athletes’ organism after a strenuous exercise. In conclusion, this study supported the importance of the assessment of albumin dimers as a predictive marker for exercise-induced oxidative stress.

The objective of this study is to investigate the aquatic-toxic effects of glyoxal-containing cellulose ether with four different glyoxal concentrations (0, 1.4, 2.3 and 6.3%) in response to global chemical regulations, e.g., European Union Classification, Labeling and Packaging (EU CLP). Tests of the aquatic-toxic effects of glyoxal-containing cellulose ether on 11 freeze-dried microbial strains, Microcystis aeruginosa, Daphnia magna and zebrafish embryos were designed as an initial stage for toxicity screening, and were performed with the standardized toxicity test guidelines. Glyoxal-containing cellulose ether showed no significant toxic effects in the toxicity tests for the 11 freeze-dried microbial strains, Daphnia magna and zebrafish embryos. Alternatively, 6.3% glyoxal-containing cellulose ether led to more than a 60% reduction of Microcystis aeruginosa growth after 7 days of exposure. Approximately 10% developmental abnormalities (e.g., bent spine) in zebrafish embryos were also observed in the group exposed to 6.3% glyoxal-containing cellulose ether after 6 days of exposure. These results imply that <6.3% glyoxal-containing cellulose ether results in non-toxic effects on the acute toxicity of aquatic organisms. However, ≥6.3% glyoxal-containing cellulose ether may affect the health of aquatic organisms with long-term exposure. In order to better evaluate the eco-safety of cellulosic products contained in glyoxal, further studies regarding the toxic effects of glyoxal-containing cellulose ether with long-term exposure are required. The results from this study allow us to evaluate the auatic-toxic effects of glyoxal-containing cellulosic products, under EU chemical regulations, on the health of aquatic organisms.

Prompt removal of misfolded membrane proteins and misassembled membrane protein complexes is essential for membrane homeostasis. However, the elimination of these toxic proteins from the hydrophobic membrane environment has high energetic barriers. Transmembrane FtsH is the only known ATP-dependent protease responsible for this task, unlike other well-studied soluble ATP-dependent proteases. The mechanisms by which FtsH recognizes, unfolds, translocates, and proteolyzes its substrates remain unclear. Here, we report the crystal structures of the Thermotoga maritima FtsH periplasmic domain (PD) in an associative trimeric state at a 1.5-1.95 Å resolution. We also describe the pH-dependent oligomerization states of the isolated PD using dynamic light scattering. These observations help us understand how FtsH recognizes membrane-anchored misfolded proteins.

The present study aimed at investigating the in vitro and in vivo susceptibility of malaria parasites to crude extracts and fractions from Polyalthia suaveolens, Uvaria angolensis, and Monodora tenuifolia. The ethanolic extracts were prepared by maceration, and were further partitioned using water, dichloromethane, hexane, and methanol. The most promising fraction was subjected to column chromatography and the sub-fractions tested for activity in vitro. The antiplasmodial effect of extracts and fractions was tested against the Chloroquine resistant (PfK1) strain in 96 wells microtiter plate format using SYBR green florescence assay. The promising fraction was further assessed for cytotoxicity and acute toxicity in Swiss albino mice and subsequently against the rodent malaria parasite, P. berghei. Qualitative phytochemical screening was also performed on the promising fraction. The methanol fractions exerted the overall better effect with that of the twigs of P. suaveolens (PStw(Ace)) showing the highest potency with a IC50 value of 3.24 µg/mL followed by the fractions of leaf of M. tenuifolia (MoTel(Ace), IC50= 3.84 µg/ml) and stem bark of P. suaveolens (IC50= 4.90 µg/ml). The phytochemical screening showed the presence of alkaloids, lactones, and phenols in the more active fraction of P. suaveolens (PStw(Ace)). The chromatographic fractionation of this fraction led to 12 sub-fractions with PS8 sub-fraction being the most active (IC50= 4.42 µg/mL). In vivo, oral administration of 2000 mg/kg b.w of fraction PStw(Ace) in mice showed no signs of toxicity. Also, fraction PStw(Ace) at 400 mg/kg b.w exerted the highest suppressive effect against P. berghei strain B throughout the 4 days experiment (% parasitaemia below 5.2%). Overall, the results achieved supported the use of the three plants in the traditional treatment of malaria in Cameroon. More interestingly, the methanolic fraction of the twigs extract from P. suaveolens might be of interest in future development of an antimalarial phytodrug.

Aquaporin-11 (AQP11) is an intracellular water channel expressed at the endoplasmic reticulum (ER) of the kidney proximal tubule. Its gene disruption in mice leads to intracellular vacuole formation at one week old and the subsequent development of polycystic kidneys at three week old. As the damaged proximal tubular cells with intracellular vacuoles later form cysts, autophagy may play a role in their survival. We examined the autophagy activity before and after the development of cysts in AQP11(-/-) kidneys. We first observed an enhanced expression of LC3 gene (Map1lc3b) as well as other autophagy-related genes in AQP11(-/-) mice by quantitative PCR analysis. We then examined the formation of autophagosomes visualized by a green fluorescent fusion protein, GFP–LC3 in its transgenic mice. The expression of GFP-LC3 puncta was increased in the proximal tubule of AQP11(-/-) mice before the cyst formation. Interestingly, they were also observed in the cyst-lining epithelial cell. Further PCR analyses revealed the enhanced expression of apoptosis- and ER stress-related caspase genes before and after the cyst formation suggesting that ER stress may have enhanced autophagy. We conclude that autophagy will play an important role in the development and the survival of the kidney cysts in AQP11(-/-) mice.

Environmental contamination has exposed humans to various metal agents, including mercury. It has been determined that mercury is not only harmful to the health of vulnerable populations such as pregnant women and children, but is also toxic to ordinary adults in various ways. For many years, mercury was used in a wide variety of human activities. Nowadays, the exposure to this metal from both natural and artificial sources is significantly increasing. Recent studies suggest that chronic exposure, even to low concentration levels of mercury, can cause cardiovascular, reproductive and developmental toxicity, neurotoxicity, nephrotoxicity, immunotoxicity, and carcinogenicity. Possible biological effects of mercury, including the relationship between mercury toxicity and diseases of the cardiovascular system, such as hypertension, coronary heart disease and myocardial infarction, are being studied. As heart rhythm and function are under autonomic nervous system control, it has been hypothesized that the neurotoxic effects of mercury might also impact cardiac autonomic function. Mercury exposure could have a long-lasting effect on cardiac parasympathetic activity and some evidence show that mercury exposure might affect heart rate variability, particularly early exposures in children. The mechanism by which mercury produces toxic effects on the cardiovascular system is not fully elucidated, but this mechanism is believed to involve an increase in oxidative stress. The exposure to mercury increases the production of free radicals, potentially because of the role of mercury in the Fenton reaction and a reduction in the activity of antioxidant enzymes, such as glutathione peroxidase. In this review we report an overview on the toxicity of mercury and focus our attention on the toxic effects on the cardiovascular system.

β-Lactams are pharmacologically important compounds because of their various biological uses, including antibiotic and so on. β-Lactams were synthesized from benzylidene-inden derivatives and acetoxyacetyl chloride. The inhibitory effect of these compounds was also examined for human carbonic anhydrase I and II (hCA I, and II) and acetylcholinesterase (AChE). The results reveal that β-lactams are inhibitors of hCA I, II and AChE. The K_i values of β-lactams (2a-k) were 0.44-6.29 nM against hCA I, 0.93-8.34 nM against hCA II, and 0.25-1.13 nM against AChE. Our findings indicate that β-lactams (2a-k) inhibit both CA isoenzymes and AChE at low nanomolar concentrations.