Tree growth and water-use in hyper-arid Acacia occurs during the hottest and driest season

Oecologia - Drought-induced tree mortality has been recently increasing and is expected to increase further under warming climate. Conversely, tree species that survive under arid conditions might...     

Structure of a flavonoid glucosyltransferase reveals the basis for plant natural product modification

Glycosylation is a key mechanism for orchestrating the bioactivity, metabolism and location of small molecules in living cells. In plants, a large multigene family of glycosyltransferases is involved in these processes, conjugating hormones, secondary metabolites, biotic and abiotic environmental toxins, to impact directly on cellular homeostasis. The red grape enzyme UDP-glucose:flavonoid 3-O-glycosyltransferase (VvGT1) is responsible for the formation of anthocyanins, the health-promoting compounds which, in planta, function as colourants determining flower and fruit colour and are precursors for the formation of pigmented polymers in red wine. We show that VvGT1 is active, in vitro, on a range of flavonoids. VvGT1 is somewhat promiscuous with respect to donor sugar specificity as dissected through full kinetics on a panel of nine sugar donors. The three-dimensional structure of VvGT1 has also been determined, both in its 'Michaelis' complex with a UDP-glucose-derived donor and the acceptor kaempferol and in complex with UDP and quercetin. These structures, in tandem with kinetic dissection of activity, provide the foundation for understanding the mechanism of these enzymes in small molecule homeostasis.     

On the dynamic nature of the transition state for protein-protein association as determined by double-mutant cycle analysis and simulation

The process of protein-protein association starts with their random collision, which may develop into an encounter complex followed by a transition state and final complex formation. Here we aim to experimentally characterize the nature of the transition state of protein-protein association for three different protein-protein interactions; Barnase-Barstar, TEM1-BLIP and IFNalpha2-IFNAR2, and use the data to model the transition state structures. To model the transition state, we determined inter-protein distance-constraints of the activated complex by using double mutant cycles (DMC) assuming that interacting residues are spatially close. Significant DeltaDeltaG(double dagger)(int) values were obtained only between residues on Barnase and Barstar. However, introducing specific mutations that optimize the charge complementarity between BLIP and TEM1 resulted in the introduction of significant DeltaDeltaG(double dagger)(int) values also between residues of these two proteins. While electrostatic interactions make major contributions towards stabilizing the transition state, we show two examples where steric hindrance exerts an effect on the transition state as well. To model the transition-state structures from the experimental DeltaDeltaG(double dagger)(int) values, we introduced a method for structure perturbation, searching for those inter-protein orientations that best support the experimental DeltaDeltaG(double dagger)(int) values. Two types of transition states were found, specific as observed for Barnase-Barstar and the electrostatically optimized TEM1-BLIP mutants, and diffusive as shown for wild-type TEM1-BLIP and IFNalpha2-IFNAR2. The specific transition states are characterized by defined inter-protein orientations, which cannot be modeled for the diffusive transition states. Mutations introduced through rational design can change the transition state from diffusive to specific. Together, these data provide a structural view of the mechanism allowing rates of association to differ by five orders of magnitude between different protein complexes.     

Karyotype and C-banding inTrigonella sectionFoenumgraecum (Fabaceae)

The six species of the sectionFoenum-graecum ofTrigonella have the same chromosome number, 2n = 16.T. gladiata andT. cariensis have fairly symmetrical karyotypes, while those ofT. foenum-graecum, T. berythea, T. macrorrhyncha andT. cassia are asymmetrical. C-bands are present in all six species but the number of bands and their positive vary considerably among the species. The karyotype evidence suggests that none of the available species of theFoenum-graecum section can be considered as the wild progenitor of fenugreek.     

Gene expression of Caenorhabditis elegans neurons carries information on their synaptic connectivity

The claim that genetic properties of neurons significantly influence their synaptic network structure is a common notion in neuroscience. The nematode Caenorhabditis elegans provides an exciting opportunity to approach this question in a large-scale quantitative manner. Its synaptic connectivity network has been identified, and, combined with cellular studies, we currently have characteristic connectivity and gene expression signatures for most of its neurons. By using two complementary analysis assays we show that the expression signature of a neuron carries significant information about its synaptic connectivity signature, and identify a list of putative genes predicting neural connectivity. The current study rigorously quantifies the relation between gene expression and synaptic connectivity signatures in the C. elegans nervous system and identifies subsets of neurons where this relation is highly marked. The results presented and the genes identified provide a promising starting point for further, more detailed computational and experimental investigations.     

Enantioselective metabolism of ifosfamide by the kidney

Ifosfamide (IF), a potent chemotherapeutic agent for solid tumors, is known to cause high rates of nephrotoxicity, which is most likely due to the renal production of the metabolite chloroacetaldehyde. Enantioselective oxidation of IF has been shown in the liver but has never been reported in the kidney. Using porcine and human kidney samples, as well as the renal porcine cell line LLCPK-1, we document enantioselective metabolism of IF with prevalent production of the N-dechloroethylifosfamide (DCEIF) metabolites from the (S)-IF enantiomer compared to the amount of N-DCEIF metabolites produced from the (R)-IF enantiomers. Since IF enantiomers appear to be equally effective in chemotherapy, these results suggest that replacing the clinically standard racemic mixture of IF with (R)-IF may decrease renal metabolism of the drug and hence may decrease nephrotoxicity.     

Studies of the Extracellular Glycocalyx of the Anaerobic Cellulolytic Bacterium Ruminococcus albus 7

Anaerobic cellulolytic bacteria are thought to adhere to cellulose via several mechanisms, including production of a glycocalyx containing extracellular polymeric substances (EPS). As the compositions and structures of these glycocalyces have not been elucidated, variable-pressure scanning electron microscopy (VP-SEM) and chemical analysis were used to characterize the glycocalyx of the ruminal bacterium Ruminococcus albus strain 7. VP-SEM revealed that growth of this strain was accompanied by the formation of thin cellular extensions that allowed the bacterium to adhere to cellulose, followed by formation of a ramifying network that interconnected individual cells to one another and to the unraveling cellulose microfibrils. Extraction of 48-h-old whole-culture pellets (bacterial cells plus glycocalyx [G] plus residual cellulose [C]) with 0.1 N NaOH released carbohydrate and protein in a ratio of 1:5. Boiling of the cellulose fermentation residue in a neutral detergent solution removed almost all of the adherent cells and protein while retaining a residual network of adhering noncellular material. Trifluoroacetic acid hydrolysis of this residue (G plus C) released primarily glucose, along with substantial amounts of xylose and mannose, but only traces of galactose, the most abundant sugar in most characterized bacterial exopolysaccharides. Linkage analysis and characterization by nuclear magnetic resonance suggested that most of the glucosyl units were not present as partially degraded cellulose. Calculations suggested that the energy demand for synthesis of the nonprotein fraction of EPS by this organism represents only a small fraction (<4%) of the anabolic ATP expenditure of the bacterium.