Suvroma Gupta

The structural characterization and biological activity of sulfamethoxazolyl-azo-p-cresol, its copper(II) complex and their theoretical studies

(E)-4-((2-Hydroxy-5-methylphenyl)diazenyl)-N-(5-methylisoxazole-3-yl)benzene sulfonamide (SMX-NN-C6H3(p-Me)-OH, 1) and its Cu(II) complex, [Cu(SMX-NN-C6H3(p-Me)-O)2]n (2), have been characterized by spectroscopic data and single crystal X-ray diffraction studies. The supramolecular 1D chain of 1 is constituted by inter- and intra-molecular hydrogen bonds and also by π⋯π interactions of aromatic rings. Complex 2 has a tetragonally distorted octahedral structure in which ligand 1 serves as an N,O chelator and forms a planar Cu(N,O)2 motif; two axial positions are occupied by the oxazolyl-N of neighbouring units and a 3D structure is formed. The biological activities of these compounds have been evaluated against Gram positive (B. subtilis; IC50: 105 μg ml−1 (1) and 105 μg ml−1 (2)) and Gram negative bacteria (E. coli; IC50: 66.36 μg ml−1 (1) and 62.2 μg ml−1 (2)) and the complexes exhibit better efficiency. Interactions of DNA with 1 and 2 have been examined and the binding constants are Kb(1), 5.920 × 104 M−1 and Kb(2), 4.445 × 104 M−1. The in silico test is used to predict the most favoured binding mode of 1 and 2 with the active site residues of DHPS (dihydropteroate synthetase) of E. coli and of DNA. The Cu(II) complex (2) binds more efficiently (CDOCKER energy, 2, −61.35 a.u.) in the DHPS cavity than ligand 1 (CDOCKER energy, 1, −43.90 a.u.). The electronic structures and spectral properties of 1 and 2 have been investigated by DFT and TD-DFT computation of optimized geometries of the compounds.

Optimization of Solid State Fermentation Conditions and Characterization of Thermostable Alpha Amylase from Bacillus subtilis (ATCC 6633)

Alpha amylase production using microbial source and solid state fermentation has been conducted for past few years in search of thermostable enzyme. Owing to the prolific use of thermostable alpha amylase in various industries like paper, food, detergent, brewing and starch liquefaction process, the production of alpha amylase is still going on. In the present work, Bacillus subtilis (ATCC 6633) has been utilized for generation of alpha amylase followed by optimization of the fermentation media. Change in fermentation conditions like fermentation hour, temperature, inoculums size, nitrogen and sugar sources have pivotal role to enhance alpha amylase yield. The thermal, pH and detergent stability of partially purified alpha amylase have been tested and compared with purified porcine pancreatic amylase. The result is encouraging with approximate 80% retention of alpha amylase activity comparable to purified porcine pancreatic amylase in presence of drastic condition of temperature (60°C), pH (6-11) and detergents. This makes it apt for use in various industries like detergent, food and paper industries.

Production and characterization of thermostable alkaline protease of Bacillus subtilis (ATCC 6633) from optimized solid‐state fermentation

Proteases are the most important group of enzymes utilized commercially in various arenas of industries, such as food, detergent, leather, dairy, pharmaceutical, diagnostics, and waste management, accounting for nearly 20% of the world enzyme market. Microorganisms of specially Bacillus genera serve as a vast repository of diverse set of industrially important enzymes and utilized for the large‐scale enzyme production using a fermentation technology. Approximately 30%–40% of the cost of industrial enzymes originates from the cost of the growth medium. This study is attempted to produce protease from Bacillus subtilis (ATCC 6633) after optimization of various process parameters with the aid of solid‐state fermentation using a cheap nutrient source such as wheat bran. B. subtilis (ATCC 6633) produces proteases of molecular weight 36 and 20 kDa, respectively, in the fermented medium as evident from SDS zymogram. Alkaline protease activity has been detected with optimum temperature at 50 °C and is insensitive to ethylenediaminetetraacetic acid. This thermostable alkaline protease exhibits dual pH optimum at 7 and 10 with moderate pH stability at alkaline pH range. It preserves its activity in the presence of detergent such as SDS, Tween 20, and Triton X‐100 and may be considered as an effective additive to detergent formulation with some industrial importance.

Optimization of Amylase Production from B. amyloliquefaciens (MTCC 1270) Using Solid State Fermentation

Demand for microbial amylase production persists because of its immense importance in wide spectrum industries. The present work has been initiated with a goal of optimization of solid state fermentation condition for amylase using agroindustrial waste and microbial strain like B. amyloliquefaciens (MTCC 1270). In an aim to improve the productivity of amylase, fermentation has been carried out in the presence of calcium (Ca+2), Nitrate (NO3 −), and chloride ions (Cl−) as well as in the presence of D-inositol and mannitol. Amylase needs calcium ion for the preservation of its structure, activity and stability that proves beneficial also for amylase production using solid state fermentation. The inclusion of ions and sugars in the SSF media is promising which can be explained by the protection offered by them against thermal decay of amylase at various incubation periods at 37°C.

MAP2c prevents arachidonic acid-induced fibril formation of tau: Role of chaperone activity and phosphorylation

Tau has long been associated with Alzheimers disease, where it forms neurofibrillary tangles. Here we show for the first time by electron microscopy that MAP2c prevents arachidonic acid-induced in vitro aggregation of tau. However, phosphorylated MAP2c failed to prevent the same. Previously we reported that MAP2c possesses chaperone-like activity while tau does not (Sarkar et al., 2004, Eur J Biochem., 271(8), 1488-96). Here we demonstrate that phosphorylation severely impaired the chaperone activity of MAP2c, implying a crucial role of chaperone in preventing tau fibrillation. Additionally, the ability of MAP2c to induce microtubule polymerization was abolished completely upon phosphorylation. As tau and MAP2c possess highly homologous C-termini, we speculated that the N-terminus of MAP2c might account for its chaperone activity. Nevertheless, experiments showed that N-terminus of MAP2c alone is inactive as a chaperone. Our preliminary findings suggest that MAP2c/MAP2 could be one of the regulators maintaining tau homeostasis in the cell.

Cytotoxic biphenyl-4-carboxylic acid targets the tubulin–microtubule system and inhibits cellular migration in HeLa cells

Abstract Two structurally similar biphenyl compounds, biphenyl-2-carboxylic acid (B2C) and biphenyl-4-carboxylic acid (B4C), were selected to assess their cellular cytotoxic and antimitotic behaviour in the quest of a potent anticancer compound. The HeLa and MCF-7 cell lines were used to determine the cytotoxic effect of the two biphenyl compounds using the MTT assay. Confocal microscopy was performed to analyze the degree of nuclear condensation and fragmentation associated with depolymerized microtubules that result from inhibiting in vitro tubulin polymerization. Circular dichroism spectroscopy along with DTNB kinetics were conducted to predict alterations in the tubulin secondary structure and global conformational changes in the tertiary structure of the protein, respectively. Finally, a wound healing assay was employed to assess whether cellular migration was inhibited in the treated HeLa cells. B4C imparted more cellular cytotoxicity in the HeLa and MCF-7 cell lines (IC50 ∼ 4 μM). B4C inhibited in vitro tubulin polymerization into microtubules with an IC50 of 50 μM. Confocal microscopy of treated cell indicated presumptive apoptosis, exhibiting fragmented nuclei with significant microtubular disruption, which was confirmed by Western blot analysis. Circular dichroism revealed a significant reduction in the α-helix content of treated tubulin. DTNB kinetics showed that approximately six –SH groups were buried in the structure. The wound healing assay revealed that B4C prevented cellular invasion. B4C had greater cytotoxic and antimitotic effects against HeLa cells than B2C. To delineate the specific mechanism of action of B4C and its derivatives, further research is warranted.