The carbohydrases of Bacillus stearothermophilus NCIB 11412 and NCIB 10814

Summary Two strains (NCIB 11412 and NCIB 10814) of the thermophilic organism Bacillus stearothermophilus were found to produce complex carbohydrase systems. The enzyme activities in each system include -amylase as the major component, maltase, pullulanase, a minor amylase and cyclodextrinase. The latter three activities are produced in low yield in both strains. A crude enzyme preparation from each strain possessed maltogenic properties on hydrolysis of soluble starch. Following rigorous purification procedures, the purified major -amylase from either strain did not produce maltose as a major end-product of starch hydrolysis. However, a partially purified mixture of pullulanase, minor amylase and cyclodextrinase activities from NCIB 11412 and NCIB 10814 produced 56.4% and 62.0% maltose, respectively, from soluble starch.     

Production and properties of polygalacturonate lyase by an alkalophilic microorganism Bacillus sp. RK9.

Bacillus sp. RK9 was isolated from soil and produced a constitutive polygalacturonate lyase. Production of the enzyme required the presence of complex nitrogen (peptone and yeast extract). Highest activity was obtained with an initial pH of 9.7. The organism was alkalophilic. No growth occurred below pH 7.5. The enzyme was purified by salt precipitation and diethylaminoethyl (DEAE) cellulose ion-exchange chromatography. The pH optimum for activity was 10.0 in 0.01 M glycine-NaOH buffer. Calcium alone, of divalent cations, activated the enzyme by 2.9-fold. Complete inhibition of enzyme activity was achieved by 1 mM ethylenediaminetetraacetic acid (EDTA). Hydrolysis of substrate occurred in a random fashion and the enzyme was 50% more active towards acid soluble pectic acid (ASPA) than towards sodium polypectate.     

The role of snail aestivation in transmission of schistosomiasis in changing climatic conditions

Schistosomiasis vector snails are subjected to extreme seasonal changes, particularly in ephemeral rivers and lentic waterbodies. In the tropics, aestivation is one of the adaptive strategies for survival and is used by snails in times of extremely high temperatures and desiccation. Aestivation therefore plays an important role in maintaining the transmission of schistosomiasis. This review assesses the possible impacts of climate change on the temporal and spatial distribution of schistosomiasis-transmitting snails with special emphasis on aestivation, and discusses the effect of schistosome infection on aestivation ability. The impacts of parasite development on snails, as well as physiological changes, are discussed with reference to schistosomiasis transmission. This review shows that schistosome-infected snails have lower survival rates during aestivation, and that those that survive manage to get rid of the infection. In general, snail aestivation ability is poor and survival chances diminish with time. Longer dry periods result in fewer, as well as uninfected, snails. However, the ability of the surviving snails to repopulate the habitats is high.     

The Effects of Sub-Regional Climate Velocity on the Distribution and Spatial Extent of Marine Species Assemblages

Many studies illustrate variable patterns in individual species distribution shifts in response to changing temperature. However, an assemblage, a group of species that shares a common environmental niche, will likely exhibit similar responses to climate changes, and these community-level responses may have significant implications for ecosystem function. Therefore, we examine the relationship between observed shifts of species in assemblages and regional climate velocity (i.e., the rate and direction of change of temperature isotherms). The assemblages are defined in two sub-regions of the U.S. Northeast Shelf that have heterogeneous oceanography and bathymetry using four decades of bottom trawl survey data and we explore temporal changes in distribution, spatial range extent, thermal habitat area, and biomass, within assemblages. These sub-regional analyses allow the dissection of the relative roles of regional climate velocity and local physiography in shaping observed distribution shifts. We find that assemblages of species associated with shallower, warmer waters tend to shift west-southwest and to shallower waters over time, possibly towards cooler temperatures in the semi-enclosed Gulf of Maine, while species assemblages associated with relatively cooler and deeper waters shift deeper, but with little latitudinal change. Conversely, species assemblages associated with warmer and shallower water on the broad, shallow continental shelf from the Mid-Atlantic Bight to Georges Bank shift strongly northeast along latitudinal gradients with little change in depth. Shifts in depth among the southern species associated with deeper and cooler waters are more variable, although predominantly shifts are toward deeper waters. In addition, spatial expansion and contraction of species assemblages in each region corresponds to the area of suitable thermal habitat, but is inversely related to assemblage biomass. This suggests that assemblage distribution shifts in conjunction with expansion or contraction of thermal habitat acts to compress or stretch marine species assemblages, which may respectively amplify or dilute species interactions to an extent that is rarely considered. Overall, regional differences in climate change effects on the movement and extent of species assemblages hold important implications for management, mitigation, and adaptation on the U.S. Northeast Shelf.     

Attenuation of pattern recognition receptor signaling is mediated by a MAP kinase kinase kinase

Pattern recognition receptors (PRRs) play a key role in plant and animal innate immunity. PRR binding of their cognate ligand triggers a signaling network and activates an immune response. Activation of PRR signaling must be controlled prior to ligand binding to prevent spurious signaling and immune activation. Flagellin perception in Arabidopsis through FLAGELLIN‐SENSITIVE 2 (FLS2) induces the activation of mitogen‐activated protein kinases (MAPKs) and immunity. However, the precise molecular mechanism that connects activated FLS2 to downstream MAPK cascades remains unknown. Here, we report the identification of a differentially phosphorylated MAP kinase kinase kinase that also interacts with FLS2. Using targeted proteomics and functional analysis, we show that MKKK7 negatively regulates flagellin‐triggered signaling and basal immunity and this requires phosphorylation of MKKK7 on specific serine residues. MKKK7 attenuates MPK6 activity and defense gene expression. Moreover, MKKK7 suppresses the reactive oxygen species burst downstream of FLS2, suggesting that MKKK7‐mediated attenuation of FLS2 signaling occurs through direct modulation of the FLS2 complex.     

A multi‐resolution model to capture both global fluctuations of an enzyme and molecular recognition in the ligand‐binding site

In multi‐resolution simulations, different system components are simultaneously modeled at different levels of resolution, these being smoothly coupled together. In the case of enzyme systems, computationally expensive atomistic detail is needed in the active site to capture the chemistry of ligand binding. Global properties of the rest of the protein also play an essential role, determining the structure and fluctuations of the binding site; however, these can be modeled on a coarser level. Similarly, in the most computationally efficient scheme only the solvent hydrating the active site requires atomistic detail. We present a methodology to couple atomistic and coarse‐grained protein models, while solvating the atomistic part of the protein in atomistic water. This allows a free choice of which protein and solvent degrees of freedom to include atomistically. This multi‐resolution methodology can successfully model stable ligand binding, and we further confirm its validity by exploring the reproduction of system properties relevant to enzymatic function. In addition to a computational speedup, such an approach can allow the identification of the essential degrees of freedom playing a role in a given process, potentially yielding new insights into biomolecular function. Proteins 2016; 84:1902–1913. © 2016 Wiley Periodicals, Inc.     

The amylopullulanase of Bacillus sp. DSM 405

The amylopullulanse produced by Bacillus sp. DSM 405 was purified to homogeneity. It exhibited dual activity, cleaving the α1-4 bonds in starch, releasing a range of malto-oligosaccharides, and also cleaving the α1-6 bonds in pullulan, releasing maltotriose as the sole end-product. The enzyme was a glycoprotein and had a relative molecular mass of 126 000 and an isoelectric point of 4.3. While the enzyme was optimally active on starch at pH 6.5 and at pH 6.0 on pullulan, activity on both substrates was maximal at 70 °C. Kinetic analyses of the enzyme in a system that contained both starch and pullulan as two competing substrates demonstrated the dual specificity of the enzyme. Chemical modification of the carboxyl groups within the active centre of the protein showed that one active site was responsible for hydrolysis of the α1-4 and α1-6 bonds in starch and pullulan respectively. This is the first comprehensive investigation of an amylopullulanse produced by an aerobic bacterium, showing a single active site responsible for both activities. Received: 3 August 1998 / Received revision: 13 October 1998 / Accepted: 16 October 1998     

Unconventional myosin Myo1c promotes membrane fusion in a regulated exocytic pathway

Glucose homeostasis is controlled in part by regulation of glucose uptake into muscle and adipose tissue. Intracellular membrane vesicles containing the GLUT4 glucose transporter move towards the cell cortex in response to insulin and then fuse with the plasma membrane. Here we show that the fusion step is retarded by the inhibition of phosphatidylinositol (PI) 3-kinase. Treatment of insulin-stimulated 3T3-L1 adipocytes with the PI 3-kinase inhibitor LY294002 causes the accumulation of GLUT4-containing vesicles just beneath the cell surface. This accumulation of GLUT4-containing vesicles near the plasma membrane prior to fusion requires an intact cytoskeletal network and the unconventional myosin motor Myo1c. Remarkably, enhanced Myo1c expression under these conditions causes extensive membrane ruffling and overrides the block in membrane fusion caused by LY294002, restoring the display of GLUT4 on the cell exterior. Ultrafast microscopic analysis revealed that insulin treatment leads to the mobilization of GLUT4-containing vesicles to these regions of Myo1c-induced membrane ruffles. Thus, localized membrane remodeling driven by the Myo1c motor appears to facilitate the fusion of exocytic GLUT4-containing vesicles with the adipocyte plasma membrane.