Engineering co-culture system for production of apigetrin in <Emphasis Type="Italic">Escherichia coli</Emphasis>

Microbial cells have extensively been utilized to produce value-added bioactive compounds. Based on advancement in protein engineering, DNA recombinant technology, genome engineering, and metabolic remodeling, the microbes can be re-engineered to produce industrially and medicinally important platform chemicals. The emergence of co-culture system which reduces the metabolic burden and allows parallel optimization of the engineered pathway in a modular fashion restricting the formation of undesired byproducts has become an alternative way to synthesize and produce bioactive compounds. In this study, we present genetically engineered E. coli-based co-culture system to the de novo synthesis of apigetrin (APG), an apigenin-7-O-β-d-glucopyranoside of apigenin. The culture system consists of an upstream module including 4-coumarate: CoA ligase (4CL), chalcone synthase, chalcone flavanone isomerase (CHS, CHI), and flavone synthase I (FNSI) to synthesize apigenin (API) from p-coumaric acid (PCA). Whereas, the downstream system contains a metabolizing module to enhance the production of UDP-glucose and expression of glycosyltransferase (PaGT3) to convert API into APG. To accomplish this improvement in titer, the initial inoculum ratio of strains for making the co-culture system, temperature, and media component was optimized. Following large-scale production, a yield of 38.5 µM (16.6 mg/L) of APG was achieved. In overall, this study provided an efficient tool to synthesize bioactive compounds in microbial cells.     

Aboveground,belowground biomass and nutrients pool in <Emphasis Type="Italic">Salicornia brachiata</Emphasis> at coastal area of India: interactive effects of soil characteristics

The present study determined the plant biomass (aboveground and belowground) of Salicornia brachiata from six different salt marshes distributed in Indian coastal area over one growing season (September 2014–May 2015). The nutrients concentration and their pools were estimated in plant as well as soil. Belowground biomass in S. brachiata was usually lower than the aboveground biomass. Averaged over different locations, highest biomass was observed in the month of March (2.1 t ha^?1) followed by May (1.64 t ha^?1), February (1.60 t ha^?1), November (0.82 t ha^?1) and September (0.05 t ha^?1). The averaged aboveground to belowground ratio was 12.0. Aboveground and belowground biomass were negatively correlated with pH of soil, while positively with soil electrical conductivity. Further, there were positive relationships between organic carbon and belowground biomass; and available sodium and aboveground biomass. The nutrient pools in aboveground were always higher than to belowground biomass. Aboveground pools of carbon (543 kg ha^?1), nitrogen (48 kg ha^?1), phosphorus (4 kg ha^?1), sodium (334 kg ha^?1) and potassium (37 kg ha^?1) were maximum in the month of March 2015. Bioaccumulation and translocation factors for sodium of S. brachiata were more than one showing tolerance to salinity and capability of phytoremediation for the saline soil.     

Novel KV7 ion channel openers for the treatment of epilepsy and implications for detrusor tissue contraction

Neuronal voltage-gated potassium channels, K_V7s, are the molecular mediators of the M current and regulate membrane excitability in the central and peripheral neuronal systems. Herein, we report novel small molecule K_V7 openers that demonstrate anti-seizure activities in electroshock and pentylenetetrazol-induced seizure models without influencing Rotarod readouts in mice. The anti-seizure activity was determined to be proportional to the unbound concentration in the brain. K_V7 channels are also expressed in the bladder smooth muscle (detrusor) and activation of these channels may cause localized undesired effects. Therefore, the impact of individual K_V7 isoforms was investigated in human detrusor tissue using a panel of K_V7 openers with distinct activity profiles among K_V7 isoforms. KCNQ4 and KCNQ5 mRNA were highly expressed in detrusor tissue, yet a compound that has significantly reduced activity on homomeric K_V7.4 did not reduce detrusor contraction. This may suggest that the homomeric K_V7.4 channel plays a less significant role in bladder contraction and further investigation is needed.     

Biotic and Abiotic Drivers of Peatland Growth and Microtopography: A Model Demonstration

Peatlands are important carbon reserves in terrestrial ecosystems. The microtopography of a peatland area has a strong influence on its carbon balance, determining carbon fluxes at a range of spatial scales. These patterned surfaces are very sensitive to changing climatic conditions. There are open research questions concerning the stability, behaviour and transformation of these microstructures, and the implications of these changes for the long-term accumulation of organic matter in peatlands. A simple two-dimensional peat microtopographical model was developed, which accounts for the effects of microtopographical variations and a dynamic water table on competitive interactions between peat-forming plants. In a case study of a subarctic mire in northern Sweden, we examined the consequences of such interactions on peat accumulation patterns and the transformation of microtopographical structure. The simulations demonstrate plausible interactions between peatland growth, water table position and microtopography, consistent with many observational studies, including an observed peat age profile from the study area. Our model also suggests that peatlands could exhibit alternative compositional and structural dynamics depending on the initial topographical and climatic conditions, and plant characteristics. Our model approach represents a step towards improved representation of peatland vegetation dynamics and net carbon balance in Earth system models, allowing their potentially important implications for regional and global carbon balances and biogeochemical and biophysical feedbacks to the atmosphere to be explored and quantified.     

Earliness index deters <Emphasis Type="Italic">Pectinophora gossypiella</Emphasis> incidence on advanced cultivars of <Emphasis Type="Italic">Bt</Emphasis> cotton

Pink bollworm (Pectinophora gossypiella) is recognized as an important pest of cotton and can damage flowers and bolls of both Bt and non-Bt cultivars. Cry-1Ac in Bt cultivars is considered very effective in controlling lepidopterous larvae; therefore, the present study was carried out to investigate the impact of Cry1-Ac and the earliness index on the natural incidence of P. gossypiella at the Cotton Research Institute, Faisalabad. During 2015–2016, ten cultivars were used to determine the incidence of pink bollworm infestation. The experiment was repeated for 2 years. During the next year, Cry1-Ac and earliness traits of selected cultivars were also observed to determine their impact on pink bollworm. Correlation coefficient results regarding days to first flower (r value = 0.66) as well as the earliness index (r value = ? 0.62) exhibited a strong association with pink bollworm, but Cry1-Ac had a weak association (r value = ? 0.058) with pink bollworm. The coefficient of determination (R ²) explained that variability of pink bollworm due to Cry1-Ac, the earliness index, and days to first flower was 18.0, 38.5, and 43.5%, respectively. Principal component analysis results showed that the first two PCs expressed 87% of the total variability. Clusters made on the basis of the studied parameters revealed that clusters 2 and 3 comprised the cotton cultivars possessing earliness traits compared with cluster 1. Therefore, it can be concluded that the earliness index in cotton is an important component for the sustainable management of pink bollworm infestation, the need for which is endless to evade the pink bollworm problem in the era of climate change.     

Biocatalytic polyester synthesis: analysis of the evolution of molecular weight and end group functionality

The enzymatic synthesis of polyesters from activated diesters and diols has been investigated. Differences between enzymatic synthesis and traditional chemical condensation processes are discussed. The disappearance of monomers during the initial phase of reaction indicates that enzyme has a higher specificity for transesterification of ester-terminated oligomers. During the intermediate phase, enzymatic polymerization involves a competition between diol and enzyme-bound water for the nucleophilic attack of the acyl enzyme intermediate. Competition between enzymatic transesterification and hydrolysis at different stages of polymerization in nonaqueous media is responsible for termination of polyesters with acid end-groups and also for limiting the polymer molecular weight. The resulting oligoester consists of chains that are either terminated with - OH groups and/or - COOH groups. We have used Matrix Assisted Laser Desorption/Ionization - Time of Flight Mass spectroscopy (MALDI-TOF) along with colorimetric titration techniques to determine the acidity of enzyme-synthesized polyesters. This paper addresses how the enzymatic polymerization proceeds, and compares our results to the growing literature in this field. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 227-239, 1997.     

Identification and activation of mitogen-activated protein (MAP) kinase in normal human osteoblastic and bone marrow stromal cells: Attenuation of MAP kinase activation by cAMP,parathyroid hormone and forskolin

The mitogen-activated protein (MAP) kinases (p44^mapk and p42^mapk), also known as extracellular signal-regulated kinases 1 and 2 (ERK1 and ERK2), are activated in response to a variety of extracellular signals, including growth factors, hormones and, neurotransmitters. We have investigated MAP kinase signal transduction pathways in normal human osteoblastic cells. Normal human bone marrow stromal (HBMS), osteoblastic (HOB), and human (TE85, MG-63, SaOS-2), rat (ROS 17/2.8, UMR-106) and mouse (MC3T3-E1) osteoblastic cell lines contained immunodetectable p44^mapk/ERK1 and p42^mapk/ERK2. MAP kinase activity was measured by 'in-gel' assay myelin basic protein as the substrate. Mainly ERK2 was rapidly activated (within 10 min) by bFGF, IGF-I and PDGF-BB in normal HOB, HBMS and human osteosarcoma cells, whereas both ERK1 and ERK2 were activated by growth factors in rat osteoblast-like cell lines, ROS 17/2.8 and UMR-106. The ERK1 activation was greater than the ERK2 in ROS 17/2.8 cells. Furthermore, ERK2 was also activated by bFGF and PDGF-BB in the mouse osteoblastic cell line, MC3T3-E1. This is the first demonstration of inter-species differences in the activation of MAP kinases in osteoblastic cells. Cyclic AMP derivatives or cAMP generating agents such as PTH and forskolin inhibited ERK2 activation by bFGF and PDGF-BB suggesting a 'cross-talk' between the two different signalling pathways activated by receptor tyrosine kinases and cAMP-dependent protein kinase. The accumulated results also suggest that the MAP kinases may be involved in mediating mitogenic and other biological actions of bFGF, IGF-I and PDGF-BB in normal human osteoblastic and bone marrow stromal cells.     

Use of Polyelectrolytes and Electron Microscopy for Detection of Viruses from Stool

Insoluble polyelectrolytes (PE60) were used for the concentration of viruses from stool specimens, confirming the results of Wallis et al. (1969). Ten percent suspensions inoculated with poliovirus type 3 were used in these experiments. A small number of stool specimens from patients naturally infected with enteroviruses were also tested. Preferential adsorption of viruses to PE60 was maximum at a pH range of 4.5 to 6.0. The elution of the adsorbed viruses was optimal at pH 8.5. Other parameters were also investigated. Electron microscopy was used successfully to detect the eluted viruses.