Evaluation of a positive conditioning technique for influencing big sagebrush (Artemisia tridentata subspp. wyomingensis) consumption by goats

Diets of angora goats (Capra hircus) positively conditioned to eat big sagebrush (Artemisia tridentata subspp. wyomingensis) were contrasted with control groups to assess the effects of positive conditioning. Goats were conditioned by including ever-increasing amounts of sagebrush in the daily ration, to a maximum of 25% by weight as fed. Conditioning effects were evaluated by comparing relative consumption of big sagebrush in a rangeland setting. Field trials were conducted at the Squaw Butte Experimental Range, a shrub steppe rangeland in eastern Oregon. Our results indicate that neither conditioned does nor kids had significantly different intake of sagebrush when compared with control animals. Young animals consumed shrub species sooner than adults and ate significantly more shrubs throughout all seasons until the second summer when diets did not differ between age groups.     

Predivision in human oocytes at meiosis I: a mechanism for trisomy formation in man

Summary Cytogenetic preparations from oocytes remaining unfertilised after in vitro fertilisation revealed single chromatids (as opposed to whole chromosomes) in 4 out of 38 meiosis II metaphases. In one oocyte, a single chromatid was present in addition to the normal 23,X complement, and in three oocytes, two identical but separate chromatids replaced one whole chromosome within the complement. The data indicate that the chromatids have arisen as a result of premature division of the centromeres at meiosis I (predivision). None of the oocytes were hyperhaploid with an extra whole chromosome. These findings are at variance with the general belief that trisomy in man is largely a consequence of nondisjunction of whole bivalents at meiosis I and they suggest that predivision resulting in separate chromatids may be a significant mechanism for human trisomy.     

Mapping fungal ion channel locations

Ion channel mapping techniques are described and the results for two fungal organisms, Saprolegnia ferax and Neurospora crassa, are presented. In these species, two channel types have been characterized, stretch-activated channels exhibiting significant calcium permeability and spontaneous channels having significant potassium permeability. Two distinct analyses of patch clamp data, analysis of channel self-clustering and association between different channel types, and localization along the hyphae, reveal significant differences between the two organisms. S. ferax maintains a tip-high gradient of both channel types which is lost after disruption of the actin cytoskeleton. There is significant self-clustering of the channels, as well as interactions between channel types. N. crassa on the other hand does not maintain tip-high gradients, and clustered distributions are observed only for the stretch-activated channels. In terms of physiological roles, evidence is quite strong that the stretch-activated channels function as a growth sensor in S. ferax, but have an unknown function in N. crassa. In both organisms, the potassium permeable channels presumably function in potassium uptake. The differences between these two organisms may be due, in part, to differences in their normal environment: aquatic versus terrestrial. Copyright 1998 Academic Press.     

Protein unfolding, and the "tuning in" of reversible intermediate states, in protic ionic liquid media

Protic ionic liquids (PILs) are currently being shown to be as interesting and valuable to chemical manipulations as the well-known aprotic ionic liquids (APIL). PILs have the additional advantage that the proton activity (PA) can be adjusted by the choice of Bronsted base and Bronsted acid used in their formation. In the absence of solvent, the PA plays the role of pH in ordinary solutions. Previously, we have shown that solution of proteins in ionic-liquid-rich solutions conveys surprising stabilization against hydrolysis and aggregation, permitting multiple unfold/refold cycles without loss to aggregation. Here, we show that the denaturing temperatures of both hen egg white lysozyme and ribonuclease A are sensitive to the PA of the PIL as much as they are to pH in aqueous solutions. A maximum stability for more basic solutions is found, and the unfolding process is well described by the two-state (cooperative) model. Finally, we show that, by PA tuning, the PILs can select folding pathways featuring the postulated intermediates so that they are fully populated during the unfolding process. The intermediates are themselves capable of multiple unfold/refold cycles with little loss per cycle to aggregation process.     

Biphenyl amide p38 kinase inhibitors 4: DFG-in and DFG-out binding modes

The biphenyl amides (BPAs) are a series of p38α MAP kinase inhibitors. Compounds are able to bind to the kinase in either the DFG-in or DFG-out conformation, depending on substituents. X-ray, binding, kinetic and cellular data are shown, providing the most detailed comparison to date between potent compounds from the same chemical series that bind to different p38α conformations. DFG-out-binding compounds could be made more potent than DFG-in-binding compounds by increasing their size. Unexpectedly, compounds that bound to the DGF-out conformation showed diminished selectivity. The kinetics of binding to the isolated enzyme and the effects of compounds on cells were largely unaffected by the kinase conformation bound.     

Biphenyl amide p38 kinase inhibitors 1: Discovery and binding mode

The biphenyl amides (BPAs) are a novel series of p38 MAP kinase inhibitors. The discovery of the series through structure-based focused screening is described, and the binding mode of the compounds is explained with reference to X-ray crystal structures.     

Toward a human blood serum proteome: analysis by multidimensional separation coupled with mass spectrometry

Blood serum is a complex body fluid that contains various proteins ranging in concentration over at least 9 orders of magnitude. Using a combination of mass spectrometry technologies with improvements in sample preparation, we have performed a proteomic analysis with submilliliter quantities of serum and increased the measurable concentration range for proteins in blood serum beyond previous reports. We have detected 490 proteins in serum by on-line reversed-phase microcapillary liquid chromatography coupled with ion trap mass spectrometry. To perform this analysis, immunoglobulins were removed from serum using protein A/G, and the remaining proteins were digested with trypsin. Resulting peptides were separated by strong cation exchange chromatography into distinct fractions prior to analysis. This separation resulted in a 3-5-fold increase in the number of proteins detected in an individual serum sample. With this increase in the number of proteins identified we have detected some lower abundance serum proteins (ng/ml range) including human growth hormone, interleukin-12, and prostate-specific antigen. We also used SEQUEST to compare different protein databases with and without filtering. This comparison is plotted to allow for a quick visual assessment of different databases as a subjective measure of analytical quality. With this study, we have performed the most extensive analysis of serum proteins to date and laid the foundation for future refinements in the identification of novel protein biomarkers of disease.     

Hyperquenching and cold equilibration strategies for the study of liquid--liquid and protein folding transitions

In this paper we consider the extension of the recent quantitative studies of hyperquenched glassformers to include (1). systems that exhibit first order liquid-liquid phase transitions, and (2). systems that contain molecules, which, during normal cooling, undergo internal structural changes above the glass temperature. The general aim of these studies is to trap-in a high enthalpy, high entropy, state of the system and then observe it evolving in time at low temperatures during a controlled annealing procedure. In this manner events that normally occur during change of temperature may be observed occurring during passage of time, at much lower temperatures. At such low temperatures the smearing effects of vibrations are greatly reduced. While the case of most interest in the second class is the refolding of thermally denatured protein molecules, any reconstructive molecular or chemical exchange process is a potential subject for investigation. Processes that occur in stages can be studied in greater detail, and any stage of interest can be frozen when desired, by drop of temperature, for more detailed spectroscopic examination. We review an electrospray method for hyperquenching liquids at approximately 10(5) K/s, and discuss some results of such experiments in order to illustrate a calorimetric approach to exploiting the hyperquenching-and- cold-equilibration strategy. To apply the idea to the study of proteins, the following protein solvent requirements must be met: (1). the solvents must not crystallize ice on cooling or heating, yet must not denature the proteins; (2). the solvents must support thermally denatured molecules without permitting aggregation. We describe two solvent systems, the first of which meets the first requirement, but the second only partially. The second solvent system apparently meets both. Preliminary results, only at the proof of concept stage, are reported for cold refolding of lysozyme, which, it seems, can be trapped in our solvent in the unfolded but refoldable state, with only moderate (approx. 120 K/s) quenching rates.     

Differential expression of specific sulphate transporters underlies seasonal and spatial patterns of sulphate allocation in trees

Sulphate uptake and its distribution within plants depend on the activity of different sulphate transporters (SULTR). In long‐living deciduous plants such as trees, seasonal changes of spatial patterns add another layer of complexity to the question of how the interplay of different transporters adjusts S distribution within the plant to environmental changes. Poplar is an excellent model to address this question because its S metabolism is already well characterized. In the present study, the importance of SULTRs for seasonal sulphate storage and mobilization was examined in the wood of poplar (Populus tremula × P. alba) by analysing their gene expression in relation to sulphate contents in wood and xylem sap. According to these results, possible functions of the respective SULTRs for seasonal sulphate storage and mobilization in the wood are suggested. Together, the present results complement the previously published model for seasonal sulphate circulation between leaves and bark and provide information for future mechanistic modelling of whole tree sulphate fluxes.