Purification,physicochemical and regulatory properties of serine hydroxymethyltransferase from sheep liver

Serine hydroxymethyltransferase (EC 2.1.2.1) was purified from the cytosolic fraction of sheep liver by ammonium sulphate fractionation, CM-Sephadex chromatography, gel filtration using Ultrogel ACA 34 and Blue Sepharose affinity chromatography. The homogeneity of the enzyme was rigorously established by Polyacrylamide gel and sodium dodecyl sulphate-polyacrylamide gel electrophoresis, isoelectrofocusing, ultracentrifugation, immunodiffusion and Immunoelectrophoresis. The enzyme was a homotetramer with a molecular weight of 210,000 ±5000. The enzyme showed homotropic cooperative interactions with tetrahydrofolate (n_H =2.8) and a hyperbolic saturation pattern with L-serine. At the lowest concentration of tetrahydrofolate used (0.2 mM), only 5% of the added folate was oxidized during preincubation and assay. TheⁿH value was independent of the time of preincubation. Preincubation of the enzyme with serine resulted in a partial loss of the cooperative interactions (n_H =1.6) with tetrahydrofolate. The enzyme was regulated allosterically by interaction with nicotinamide nucleotides; NADH was a positive effector while NAD+ was a negative allosteric effector. The subunit interactions were retained even at the temperature optimum of 60‡C unlike in the case of the monkey liver enzyme, where these interactions were absent at higher temperatures. D-Cycloserine, a structural analogue of serine caused a sigmoid pattern of inhibition, in contrast with the observations on the monkey liver enzyme. Cibacron blue F3GA completely inhibited the enzyme and this inhibition could be reversed by tetrahydrofolate. Unlike in the monkey liver enzyme, NAD+ and NADH gave considerable protection against this inhibition. The sheep liver enzyme differs significantly in its kinetic and regulatory properties from the serine hydroxymethyltransferases isolated from other sources.     

Cholinesterase inhibitory activity of highly functionalized fluorinated spiropyrrolidine heterocyclic hybrids

Two series of dimethoxyindanone imbedded novel fluorinated spiropyrrolidine heterocyclic hybrids were synthesized employing two different less explored azomethine ylides and were measured for their efficiency as inhibitors for Alzheimer’s disease. Among the spiropyrrolidine heterocyclic hybrids, the indole based fluorinated compound with a methoxy substituent at the meta- position of the aryl ring exhibited the utmost potent AChE and BChE inhibitory activities with an IC₅₀ of 1.97 ± 0.19 µM and 7.08 ± 0.20 µM respectively. The plausible mechanism of inhibition on ChE receptors was unveiled via molecular docking studies.     

Analysis of sequential accumulation of individual pigeonpea protease inhibitors during seed development

Seeds of pigeonpea are known to accumulate protease inhibitors (PIs), belonging to the Bowman-Burk inhibitor family. PIs are important for natural defense against phytophagous insect pests. Most insects attack crops at the early stages of seed development. Accumulation patterns of individual PIs and their relationship with each other were studied in developing seeds of 76 pigeonpea lines derived from BDN2 cultivar by ethyl methane sulfonate induced chemical mutagenesis. PIs extracted from these lines, collected between 10 and 40 days after flowering (DAF) and from mature seeds of BDN2 cultivar, were detected by using gel X-ray film contact print method. A total of 9 trypsin–chymotrypsin inhibitors were detected in mature seeds. All the nine PIs were capable of inhibiting proteases. Appearance of detectable levels of individual PIs started around 10 DAF. The PI-3 appeared early and was the most stable. It was accumulated in all the studied lines and was also detected in 84 % of the samples collected 10 DAF. In particular, 14 DAF, the individual PIs were accumulated and accumulation sequence was observed (PI-3, PI-2, PI-5, PI-7, PI-6, PI-4, PI-8, PI-9 and PI-1). Accumulation continued up to 40 DAF when seeds started hardening. Chemical mutagenesis could not produce any variation in the profile of individual PIs in the 76 studied lines in mature seeds. The process of accumulation of inhibitors is sturdy and mutagenesis fails to alter it. The robust mechanism is responsible for early appearance of PI-3. Early accumulated PIs in this study need further exploration for strengthening natural defense of pigeonpea against pests.     

Expression and immunological characterization of cardamom mosaic virus coat protein displaying HIV gp41 epitopes

The coat protein of cardamom mosaic virus (CdMV), a member of the genus Macluravirus, assembles into virus‐like particles when expressed in an Escherichia coli expression system. The N and C‐termini of the coat protein were engineered with the Kennedy peptide and the 2F5 and 4E10 epitopes of gp41 of HIV. The chimeric proteins reacted with sera from HIV positive persons and also stimulated secretion of cytokines by peripheral blood mononuclear cells from these persons. Thus, a system based on the coat protein of CdMV can be used to display HIV‐1 antigens.     

Synthesis,antimalarial activity and cytotoxic potential of new monocarbonyl analogues of curcumin

A series of novel monocarbonyl analogues of curcumin have been designed, synthesized and tested for their activity against Molt4, HeLa, PC3, DU145 and KB cancer cell lines. Six of the analogues showed potent cytotoxicity towards these cell lines with IC₅₀ values below 1 μM, which is better than doxorubicin, a US FDA approved drug. Several analogues were also found to be active against both CQ-resistant (W2 clone) and CQ-sensitive (D6) strains of Plasmodium falciparum in an in-vitro antimalarial screening. This level of activity warrants further investigation of the compounds for development as anticancer and antimalarial agents.     

DnaJs,the critical drivers of Hsp70s: genome-wide screening,characterization and expression of DnaJ family genes in Sorghum bicolor

Molecular Biology Reports - The DnaJ/Hsp40s, are important components in the chaperone machine, and play pivotal roles in plant growth, development and stress tolerance. Sorghum, the semi-arid...     

Effects of macro elements and nitrogen source on biomass accumulation and bacoside A production from adventitious shoot cultures of <Emphasis Type="Italic">Bacopa monnieri</Emphasis> (L.)

The present work deals with optimization of adventitious shoot culture of Bacopa monnieri for the production of biomass and bacoside A and has investigated the effects of macro elements (NH₄NO_3, KNO_3, CaCl_2, MgSO₄ and KH₂PO₄) and nitrogen source [NH₄ ⁺/NO₃ ⁻] of Murashige and Skoog (Physiol Plant 15:473–497, 1962) medium (MS) on accumulation of biomass and bacoside A content. Optimum number of adventitious shoots (99.33 shoots explant⁻¹), fresh weight (1.841 g) and dry weight (0.150 g) were obtained in the medium with 2.0× strength of NH₄NO₃. The highest production of bacoside A content was also recorded in the medium of 2.0× NH₄NO₃, which produced 17.935 mg g⁻¹ DW. The number of adventitious shoot biomass and bacoside A content were optimum when the NO₃ ⁻ concentration was higher than that of NH₄ ⁺. Maximum number of shoots (70.00 shoots explant⁻¹), biomass (fresh weight 1.137 g and dry weight 0.080 g) and also bacoside A content (27.106 mg g⁻¹ DW) were obtained at NH₄ ⁺/NO₃ ⁻ ratio of 14.38/37.60 mM. Overall, MS medium supplemented with 2.0× NH₄NO₃ is recommended for most efficient bacoside A production.