Regulation of the xylan-degrading apparatus of Cellvibrio japonicus by a novel two-component system

The microbial degradation of lignocellulose biomass is not only an important biological process but is of increasing industrial significance in the bioenergy sector. The mechanism by which the plant cell wall, an insoluble composite structure, activates the extensive repertoire of microbial hydrolytic enzymes required to catalyze its degradation is poorly understood. Here we have used a transposon mutagenesis strategy to identify a genetic locus, consisting of two genes that modulate the expression of xylan side chain-degrading enzymes in the saprophytic bacterium Cellvibrio japonicus. Significantly, the locus encodes a two-component signaling system, designated AbfS (sensor histidine kinase) and AbfR (response regulator). The AbfR/S two-component system is required to activate the expression of the suite of enzymes that remove the numerous side chains from xylan, but not the xylanases that hydrolyze the beta1,4-linked xylose polymeric backbone of this polysaccharide. Studies on the recombinant sensor domain of AbfS (AbfS(SD)) showed that it bound to decorated xylans and arabinoxylo-oligosaccharides, but not to undecorated xylo-oligosaccharides or other plant structural polysaccharides/oligosaccharides. The crystal structure of AbfS(SD) was determined to a resolution of 2.6A(.) The overall fold of AbfS(SD) is that of a classical Per Arndt Sim domain with a central antiparallel four-stranded beta-sheet flanked by alpha-helices. Our data expand the number of molecules known to bind to the sensor domain of two-component histidine kinases to include complex carbohydrates. The biological rationale for a regulatory system that induces enzymes that remove the side chains of xylan, but not the hydrolases that cleave the backbone of the polysaccharide, is discussed.     

Proteomics reveals that redox regulation is disrupted in patients with ethylmalonic encephalopathy

Deficiency of the sulfide metabolizing protein ETHE1 is the cause of ethylmalonic encephalopathy (EE), an inherited and severe metabolic disorder. To study the molecular effects of EE, we performed a proteomics study on mitochondria from cultured patient fibroblast cells. Samples from six patients were analyzed and revealed seven differentially regulated proteins compared with healthy controls. Two proteins involved in pathways of detoxification and oxidative/reductive stress were underrepresented in EE patient samples: mitochondrial superoxide dismutase (SOD2) and aldehyde dehydrogenase X (ALDH1B). Sulfide:quinone oxidoreductase (SQRDL), which takes part in the same sulfide pathway as ETHE1, was also underrepresented in EE patients. The other differentially regulated proteins were apoptosis inducing factor (AIFM1), lactate dehydrogenase (LDHB), chloride intracellular channel (CLIC4) and dimethylarginine dimethylaminohydrolase 1 (DDAH1). These proteins have been reported to be involved in encephalopathy, energy metabolism, ion transport, and nitric oxide regulation, respectively. Interestingly, oxidoreductase activity was overrepresented among the regulated proteins indicating that redox perturbation plays an important role in the molecular mechanism of EE. This observation may explain the wide range of symptoms associated with the disease, and highlights the potency of the novel gaseous mediator sulfide.     

A whole-genome shotgun approach for assembling and anchoring the hexaploid bread wheat genome

Polyploid species have long been thought to be recalcitrant to whole-genome assembly. By combining high-throughput sequencing, recent developments in parallel computing, and genetic mapping, we derive, de novo, a sequence assembly representing 9.1 Gbp of the highly repetitive 16 Gbp genome of hexaploid wheat, Triticum aestivum, and assign 7.1 Gb of this assembly to chromosomal locations. The genome representation and accuracy of our assembly is comparable or even exceeds that of a chromosome-by-chromosome shotgun assembly. Our assembly and mapping strategy uses only short read sequencing technology and is applicable to any species where it is possible to construct a mapping population.

Electronic supplementary material

The online version of this article (doi:10.1186/s13059-015-0582-8) contains supplementary material, which is available to authorized users.     

Age-associated differences in motor output variability and coordination during the simultaneous dorsiflexion of both feet

Older adults are more variable than young adults on tasks that demand the simultaneous control of more than one effector, and the difference between age groups may be related to their different capacity of coordinating the force output of the involved effectors. The goal of this study was to determine whether age-associated differences in motor output variability during tasks involving the simultaneous dorsiflexion of two feet can be partially explained by differences in coordination and possibly attenuated by physical training. Ten young and 22 old adults (10 trained and 12 untrained old adults) volunteered to participate in the study. Trained older adults had experience in a high-intensity mixed modality training (MMT) regime for a minimum of 1?year. Volunteers performed successive trials of a constant force task and a goal-directed task, with and without visual feedback. Within- and between-trial variability were calculated and coordination was quantified using the uncontrolled manifold (UCM) approach (i.e., co-variation of the force outputs of both feet were used to quantify a motor synergy index). Older adults exhibited greater variability and lower synergy (p?p?相似文献   

Crystallization,crystal structure analysis and molecular model of the third domain of Japanese quail ovomucoid,a Kazal type inhibitor

The third domain of Japanese quail ovomucoid, a Kazal type inhibitor, has been crystallized and its crystal structure determined at 2.5 Å resolution using multiple isomorphous replacement techniques. The asymmetric unit contains four molecules. In the crystal the molecules are arranged in two slightly different octamers with approximate D4 symmetry. The molecules are held together mainly by interactions of the N-terminal residues, which form a novel secondary structural element, a β-channel.The molecule is globular with approximate dimensions 35 Å × 27 Å × 19 Å. The secondary structural elements are a double-stranded anti-parallel β-sheet of residues Pro22 to Gly32 and an α-helix from Asn33 to Ser44. The reactive site Lys18-Asp19 is located in an exposed loop. It is close to Asn33 at the N terminus of the helical segment. The polypeptide chain folding of ovomucoid bears some resemblance to other inhibitors in the existence of an anti-parallel double strand following the reactive site loop.     

Comparison of mesophilic and thermophilic feruloyl esterases: characterization of their substrate specificity for methyl phenylalkanoates

The active sites of feruloyl esterases from mesophilic and thermophilic sources were probed using methyl esters of phenylalkanoic acids. Only 13 out of 26 substrates tested were significant substrates for all the enzymes. Lengthening or shortening the aliphatic side chain while maintaining the same aromatic substitutions completely abolished activity for both enzymes, which demonstrates the importance of the correct distance between the aromatic group and the ester bond. Maintaining the phenylpropanoate structure but altering the substitutions of the aromatic ring demonstrated that the type-A esterase from the mesophilic fungus Fusarium oxysporum (FoFaeA) showed a preference for methoxylated substrates, in contrast to the type-B esterase from the same source (FoFaeB) and the thermophilic type-B (StFaeB) and type-C (StFaeC) from Sporotrichum thermophile, which preferred hydroxylated substrates. All four esterases hydrolyzed short chain aliphatic acid (C2-C4) esters of p-nitrophenol, but not the C12 ester of laurate. All the feruloyl esterases were able to release ferulic acid from the plant cell wall material in conjunction with a xylanase, but only the type-A esterase FoFaeA was effective in releasing the 5,5' form of diferulic acid. The thermophilic type-B esterase had a lower catalytic efficiency than its mesophilic counterpart, but released more ferulic acid from plant cell walls.     

Something from nothing: bridging the gap between constraint-based and kinetic modelling

Two divergent modelling methodologies have been adopted to increase our understanding of metabolism and its regulation. Constraint-based modelling highlights the optimal path through a stoichiometric network within certain physicochemical constraints. Such an approach requires minimal biological data to make quantitative inferences about network behaviour; however, constraint-based modelling is unable to give an insight into cellular substrate concentrations. In contrast, kinetic modelling aims to characterize fully the mechanics of each enzymatic reaction. This approach suffers because parameterizing mechanistic models is both costly and time-consuming. In this paper, we outline a method for developing a kinetic model for a metabolic network, based solely on the knowledge of reaction stoichiometries. Fluxes through the system, estimated by flux balance analysis, are allowed to vary dynamically according to linlog kinetics. Elasticities are estimated from stoichiometric considerations. When compared to a popular branched model of yeast glycolysis, we observe an excellent agreement between the real and approximate models, despite the absence of (and indeed the requirement for) experimental data for kinetic constants. Moreover, using this particular methodology affords us analytical forms for steady state determination, stability analyses and studies of dynamical behaviour.     

Diel patterns of leaf C export and of main shoot growth for Flaveria linearis with altered leaf sucrose-starch partitioning

Diel C export from source leaves of two Flaveria linearis lines [85-1: high cytosolic fructose-1,6-bisphosphatase (cytFBPase) and 84-9: low cytFBPase] were estimated using three methods, including leaf steady-state (14)CO(2) labelling, leaf metabolite analysis, and leaf dry mass analysis in conjunction with leaf CO(2) exchange measurements. Synthesis and accumulation of starch during the daytime were much higher in 84-9. Relative (14)C-export (export as a % of photosynthesis) in the light was 36% higher in 85-1. The diel export patterns from (14)C-analyses correlated with those based on metabolite or dry weight/gas exchange analyses during the daytime, but not during the night. Night-time export estimated from (14)C-disappearance was 3.6 times lower than those estimated using the other methods. Even though the starch degradation at night was greater for 84-9, night-time export in 84-9 was similar to 85-1, since 84-9 showed both higher respiration and accumulation of soluble sugars (i.e. glucose) at night. Patterns of (14)C allocation to sink organs were also different in the two lines. Main stem growth was less in 84-9, being reduced most in the light when leaf export was lower relative to 85-1. Supplementation with sucrose for 1 h daily via the roots at a time when leaf export in 84-9 was low relative to 85-1 increased the stem growth rate of 84-9 to a level similar with that of 85-1. This study provides evidence that diel C availability predicted by source strength (e.g. C-export rate) influences main stem extension growth and the pattern of sink development in F. linearis.