Histamine H3 and H4 receptor affinity of branched 3-(1H-imidazol-4-yl)propyl N-alkylcarbamates

A series of imidazole-containing (non-)chiral carbamates were tested at human histamine H₃ receptor (H₃R). All compounds displayed K_i values below 100 nM. A trend for a stereoselectivity at human H₃R was observed for the chiral α-branched ligands. Selected compounds were also tested at human histamine H₄ receptor and showed moderate to weak affinities (118–1460 nM).     

A GDPase/UDPase bifunctional enzyme from Candida albicans: purification and biochemical characterization

The most frequently isolated human fungal pathogen is Candida albicans which is responsible for about 50% of all Candida infections. In healthy individuals, this organism resides as a part of the normal microbiota in equilibrium with the host. However, under certain conditions, particularly in immunocompromised patients, this opportunistic pathogen adheres to host cells causing serious systemic infections. Thus, much effort has been dedicated to the study of its physiology with emphasis on factors associated to pathogenicity. A representative analysis deals with the mechanisms of glycoprotein assembly as many cell surface antigens and other macromolecules that modulate the immune system fall within this chemical category. In this regard, studies of the terminal protein glycosylation stage which occurs in Golgi vesicles has led to the identification of nucleotidases that convert glycosyltransferase-generated dinucleotides into the corresponding mononucleotides, thus playing a double function: their activity prevent inhibition of further glycosyl transfer by the accumulation of dinucleotides and the resulting mononucleotides are exchanged by specific membrane transporters for equimolecular amounts of sugar donors from the cytosol. Here, using a simple protocol for protein separation we isolated a bifunctional nucleotidase from C. albicans active on GDP and UDP that was characterized in terms of its molecular mass, response to bivalent ions and other factors, substrate specificity and affinity. Results are discussed in terms of the similarities and differences of this nucleotidase with similar counterparts from other organisms thus contributing to the knowledge of a bifunctional diphosphatase not described before in C. albicans.

     

Cell size at S phase initiation: an emergent property of the G1/S network

The eukaryotic cell cycle is the repeated sequence of events that enable the division of a cell into two daughter cells. It is divided into four phases: G₁, S, G₂, and M. Passage through the cell cycle is strictly regulated by a molecular interaction network, which involves the periodic synthesis and destruction of cyclins that bind and activate cyclin-dependent kinases that are present in nonlimiting amounts. Cyclin-dependent kinase inhibitors contribute to cell cycle control. Budding yeast is an established model organism for cell cycle studies, and several mathematical models have been proposed for its cell cycle. An area of major relevance in cell cycle control is the G₁ to S transition. In any given growth condition, it is characterized by the requirement of a specific, critical cell size, P_S, to enter S phase. The molecular basis of this control is still under discussion. The authors report a mathematical model of the G₁ to S network that newly takes into account nucleo/cytoplasmic localization, the role of the cyclin-dependent kinase Sic1 in facilitating nuclear import of its cognate Cdk1-Clb5, Whi5 control, and carbon source regulation of Sic1 and Sic1-containing complexes. The model was implemented by a set of ordinary differential equations that describe the temporal change of the concentration of the involved proteins and protein complexes. The model was tested by simulation in several genetic and nutritional setups and was found to be neatly consistent with experimental data. To estimate P_S, the authors developed a hybrid model including a probabilistic component for firing of DNA replication origins. Sensitivity analysis of P_S provides a novel relevant conclusion: P_S is an emergent property of the G₁ to S network that strongly depends on growth rate.     

Combining data sharing with the master–worker paradigm in the common component architecture

Software component technologies are being accepted as an adequate solution for handling the complexity of applications. However, existing software component models tend to be specialized to some types of resource architectures (e.g. in-process, distributed environments, etc.) and/or do not provide a very high level of abstraction. This paper focuses on handling data sharing on operation invocations between components as a solution allowing applications to be efficiently executed on all kinds of resources. In particular, the data sharing pattern appears in master–worker applications, when workers need to access only a part of a large piece of data, either in read or write mode. This approach is applied to the Common Component Architecture model. Its benefits are discussed using an image rendering application.

An ultraviolet spectrophotometric assay for the screening of sn-2-specific lipases using 1,3-O-dioleoyl-2-O-α-eleostearoyl-sn-glycerol as substrate

In the present study, we propose a continuous assay for the screening of sn-2 lipases by using triacylglycerols (TAGs) from Aleurites fordii seed (tung oil) and a synthetic TAG containing the α-eleostearic acid at the sn-2 position and the oleic acid (OA) at the sn-1 and sn-3 positions [1,3-O-dioleoyl-2-O-α-eleostearoyl-sn-glycerol (sn-OEO)]. Each TAG was coated into a microplate well, and the lipase activity was measured by optical density increase at 272 nm due to transition of α-eleostearic acid from the adsorbed to the soluble state. The sn-1,3-regioselective lipases human pancreatic lipase (HPL), LIP2 lipase from Yarrowia lipolytica (YLLIP2), and a known sn-2 lipase, Candida antarctica lipase A (CALA) were used to validate this method. TLC analysis of lipolysis products showed that the lipases tested were able to hydrolyze the sn-OEO and the tung oil TAGs, but only CALA hydrolyzed the sn-2 position. The ratio of initial velocities on sn-OEO and tung oil TAGs was used to estimate the sn-2 preference of lipases. CALA was the enzyme with the highest ratio (0.22 ± 0.015), whereas HPL and YLLIP2 showed much lower ratios (0.072 ± 0.026 and 0.038 ± 0.016, respectively). This continuous sn-2 lipase assay is compatible with a high sample throughput and thus can be applied to the screening of sn-2 lipases.     

Fifty years of changes in UV Index and implications for skin cancer in Australia

Surface ultraviolet (UV) radiation plays an important role in human health. Increased exposure to UV radiation increases the risk of skin cancer. In Australia, public campaigns to prevent skin cancer include the promotion of daily UV forecasts. If all other atmospheric factors are equal, stratospheric ozone decreases result in UV increases. Given that Australia still has the highest skin cancer rates in the world, it is important to monitor Australia’s stratospheric ozone and UV radiation levels over time because of the effects cumulative exposure can have on humans. In this paper, two long-term ozone datasets derived from surface and satellite measurements, a radiation code and atmospheric meteorological fields are used to calculate clear-sky UV radiation over a 50-year period (1959–2009) for Australia. The deviations from 1970–1980 levels show that clear-sky UV is on the rise. After the 1990s, an overall annual increase from 2 to 6% above the 1970–1980 levels was observed at all latitudes. Examining the summer and winter deviations from 1970–1980 showed that the winter signal dominated the annual changes, with winter increases almost twice those in summer. With ozone levels not expected to recover to pre-depletion levels until the middle of this century, UV levels are expected to continue to rise. Combined with Australians favoring an outdoor life-style, when temperatures are warmer, under high levels of UV, the associated risk of skin cancer will increase.     

Lecithin:cholesterol acyltransferase deficiency protects against cholesterol-induced hepatic endoplasmic reticulum stress in mice

We recently reported that lecithin:cholesterol acyltransferase (LCAT) knock-out mice, particularly in the LDL receptor knock-out background, are hypersensitive to insulin and resistant to high fat diet-induced insulin resistance (IR) and obesity. We demonstrated that chow-fed Ldlr-/-xLcat+/+ mice have elevated hepatic endoplasmic reticulum (ER) stress, which promotes IR, compared with wild-type controls, and this effect is normalized in Ldlr-/-xLcat-/- mice. In the present study, we tested the hypothesis that hepatic ER cholesterol metabolism differentially regulates ER stress using these models. We observed that the Ldlr-/-xLcat+/+ mice accumulate excess hepatic total and ER cholesterol primarily attributed to increased reuptake of biliary cholesterol as we observed reduced biliary cholesterol in conjunction with decreased hepatic Abcg5/g8 mRNA, increased Npc1l1 mRNA, and decreased Hmgr mRNA and nuclear SREBP2 protein. Intestinal NPC1L1 protein was induced. Expression of these genes was reversed in the Ldlr-/-xLcat-/- mice, accounting for the normalization of total and ER cholesterol and ER stress. Upon feeding a 2% high cholesterol diet (HCD), Ldlr-/-xLcat-/- mice accumulated a similar amount of total hepatic cholesterol compared with the Ldlr-/-xLcat+/+ mice, but the hepatic ER cholesterol levels remained low in conjunction with being protected from HCD-induced ER stress and IR. Hepatic ER stress correlates strongly with hepatic ER free cholesterol but poorly with hepatic tissue free cholesterol. The unexpectedly low ER cholesterol seen in HCD-fed Ldlr-/-xLcat-/- mice was attributable to a coordinated marked up-regulation of ACAT2 and suppressed SREBP2 processing. Thus, factors influencing the accumulation of ER cholesterol may be important for the development of hepatic insulin resistance.