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Metabolic adaptations of Leishmania donovani in relation to differentiation, drug resistance, and drug pressure
No full textAntimonial (sodium stibogluconate, SSG) resistance
and differentiation have been shown to be closely
linked in Leishmania donovani, with SSG-resistant
strains showing an increased capacity to generate
infectious (metacyclic) forms. This is the first
untargeted LC-MS metabolomics study which integrated
both phenomena in one experimental design
and provided insights into metabolic differences
between three clinical L. donovani strains with a
similar genetic background but different SSGsusceptibilities.
We performed this analysis at different
stages during promastigote growth and in the
absence or presence of drug pressure. When comparing
SSG-resistant and SSG-sensitive strains, anumber of metabolic changes appeared to be constitutively
present in all growth stages, pointing towards
a clear link with SSG-resistance, whereas most metabolic
changes were only detected in the stationary
stage. These changes reflect the close intertwinement
between SSG-resistance and an increased metacyclogenesis
in resistant parasites. The metabolic
changes suggest that SSG-resistant parasites have (i)
an increased capacity for protection against oxidative
stress; (ii) a higher fluidity of the plasma membrane;
and (iii) a metabolic survival kit to better endure infection.
These changes were even more pronounced in a
resistant strain kept under SbIII drug pressur
Studies of Protein Conformation and Folding Using Biochemical and Biophysical Methods
No full textow a protein spontaneously folds into its biologically active folded structure is an extremely interesting and still an open question. In order to attain its final active structure, a protein needs to search an enormous number of possible conformations. This process may lead to some conformational defects and formation of partially folded intermediate states.These partially folded intermediates often contain exposed hydrophobic surface through which they can interact leading to aggregation. Protein aggregation has been implicated in several physiological disorders. In addition, aggregated proteins are often immunogenic and hence un –desirable in the formulation development of bio - therapeutics. An understanding of the conformational changes during protein folding is important to elucidate the mechanism of protein aggregation. It is also important to look for an appropriate agent or small molecule, which can prevent the protein aggregation. In this thesis, we have studied different aspects of protein folding, aggregation and explored the possible use of a chemical osmolyte to combat protein aggregation. We begin by investigating the thermodynamics of folding of a small, globular mycobacterial secretory protein, MPT63 using chemical and thermal denaturation. We observe significant similarities in the thermodynamic parameters obtained using different methods. We have also observed the presence of residual structures in the unfolded states. Our study demonstrates that key processes leading to protein folding can be understood through detailed structural studies using conventional biophysical methods (like fluorescence, circular dichroism etc) along with less conventional phosphorescence spectroscopy. Phosphorescence spectroscopy has been used to unravel valuable insights into the surroundings of individual tryptophan residues of this multi – tryptophan protein (containing four tryptophan residues). We have studied the conformational changes of MPT63 in different solution pH conditions and observed spatial rearrangement in two tryptophan environments. The other two tryptophan local environments are found unperturbed at low pH leading to protein aggregation. The study highlights the promise of phosphorescence spectroscopy for increasing our understanding of the link between partially folded intermediates and protein aggregation. Finally, we use arginine, as a small molecule stabilizer to inhibit protein aggregation and to increase refolding yield of the thermally unfolded proteins. We indicate that arginine inhibits the formation of partially folded intermediates during unfolding, which would otherwise lead to protein aggregation
Head and Neck Cancer: Role of DNA Repair Gene Polymorphisms and Mapping Altered Genomic Regions
No full textCancer is a disease of uncontrolled cell division that occurs in different parts of the body. The uncontrolled growth may sometime stop and became benign or may spread to other parts of the body by a process called metastasis. Over the years it became clear that an underlying genetic and epigenetic defect is responsible for such occasional abnormal growth of cells in an individual. At the gross level such genetic abnormality was first reflected as chromosomal aneuploidy (Lengauer, et al., 1997). More than 100 years ago, Theodor Heinrich Boveri, German biologist who postulated the association between abnormal growth of sea-urchin eggs and ‘wrong’ chromosomal complement in a cell (Boveri, 1902). Until now, although with the aid of hundreds of modern technologies, scientists have identified a large number of such abnormalities, more details about this complex disease is still unclear
Tricarbonyltechnetium(I) and tricarbonylrhenium(I) complexes of amino acids: crystal and molecular structure of a novel cyclic dimeric Re(CO)3-amino acid complex comprised of the OON donor atom set of the tridentate ligand
No full textRadiolabeled complexes of monoamino polycarboxylic, polyamino monocarboxylic and thiol containing
amino acid ligands were prepared from a fac-[99mTc(CO)3(H2O)3]+ precursor.The overall radiochemical
yield was 94–98%. The complexes exhibited substantial in vitro and in vivo stability. The corresponding Re(I)
complexes of the ligands DAPA, Asp and CysH were prepared and characterized by means of IR, NMR, and
MS spectroscopic studies, as well as X-ray crystallography (for those containing D,L-DAPA and D,L-Asp). The
rhenium complexes have been structurally correlated with the technetium complexes by means of HPLC
studies. The reaction of Re(CO)5Cl with D,L-Asp in presence of triethylamine led to the formation of a new
class of cyclic dimeric complexes formed by the OON donor atom set of the tridentate ligands. The amino
carboxylate ligand system formed well defined complexes with a fac-[M(CO)3(H2O)3]+ core and shows
good promise in 99mTc(CO)3 tracer developmen
Synthesis of Pyridine and Quinoline Based Novel Nheteroaromatics and Targeting Activity Against Macrophage-associated Disease
No full textThe work embodied in this thesis describes the synthesis of pyridine and quinoline based novel N-heteroaromatics and targeting activity against macrophageassociated disease. The work covers mainly four areas, (i) Development of novel methodology for the one pot synthesis of linear and angular fused quinazolinones. (ii) Synthesis of tetrahydropyrrolo[3',4':3,4] pyrrolo[2,1-a] isoquinoline-9,11-dione derivatives via a simple and convenient multi compartment reaction in aqueous miceller system. (iii) One pot synthesis of symmetrically 1, 4-disubstituted piperazine- 2, 5-diones. (iv) and Bioactivity of the above synthesized compounds as antileishmaniasis againsts ( Macrophage associated disease). The Chapter I begins with a review on strategic development toward the Pyridine and quinoline based Synthetic and Natural products against Leishmaniasis. Chapter II deals with the development of one step methodology using amino heterocycles and o-bromo benzyl/naphthyl bromides as reactants to produce Nheteroaromatic cationic intermediates, which upon base catalyzed nucleophilic aromatic substitution followed by in situ aerial oxidation at the benzylic position smoothly furnished the angular and linear quinazolinones . Chapter III presents the development of an efficient and environment-friendly novel approach for the synthesis of tetrahydropyrrolo[3',4':3,4] pyrrolo[2,1-a] isoquinoline-9,11-dione derivatives using isoquinolinium ylide (generated in situ from isoquinoline and phenacyl bromide in presence of a base) and an activated dienophile (aryl maleimide) in micellar solution at ambient temperature. Chapter IV describes a series of diketopiperazine derivatives synthesized by self-condensation of differently substituted α-chlorophenyl acetamides in a one-pot sequence in presence of sodium hydride under nitrogen atmosphere. Chapter V deals with the bio activity of the above synthesized compounds against lishmaniasis. The promising activities of the compounds are investigated further with in vivo study
Cholesterol lowering drug may infl uence cellular immune response by altering MHC II function
No full textMajor histocompatibility complex class II (MHC II)
expressed on the surface of antigen-presenting cells (APCs)
displays peptides to CD4 + T cells. Depletion of membrane
cholesterol from APCs by methyl � -cyclodextrin treatment
compromises peptide-MHC II complex formation coupled
with impaired binding of conformational antibody, which
binds close to the peptide binding groove of MHC II. Interestingly,the total cell surface of MHC II remains unaltered.These defects can be corrected by restoring membrane cholesterol.In silico docking studies with a three-dimensional model showed the presence of a cholesterol binding site in the transmembrane domain of MHC II (TM-MHC - II). From the binding studies it was clear that cholesterol, indeed, interacts with the TM-MHC - II and alters its conformation. Mutation of cholesterol binding residues (F240, L243, and
F246) in the TM-MHC - II decreased the affi nity for cholesterol.
Furthermore, transfection of CHO cells with full-length
mutant MHC II, but not wild-type MHC II, failed to activate
antigen-specifi c T cells coupled with decreased binding of conformation-
specifi c antibodies. Thus, cholesterol-induced
conformational change of TM-MHC - II may allosterically
modulate the peptide binding groove of MHC II leading to
T cell activation. —Roy, K., M. Ghosh, T. K. Pal, S. Chakrabarti,
and S. Roy. Cholesterol lowering drug may infl uence cellular
immune response by altering MHC I
Neuroprotective Potential of Silymarin against CNS Disorders:Insight into the Pathways and Molecular Mechanisms of Action
No full textSilymarin, a C25 containing flavonoid from the plant Silybum marianum, has been the gold standard drug to treat liver disorders associated with alcohol consumption, acute and chronic viral hepatitis, and toxin-induced hepatic failures since its discovery in 1960. Apart from the hepatoprotective nature, which is mainly due to its antioxidant and tissue regenerative properties, Silymarin has recently been reported to be a putative neuroprotective
agent against many neurologic diseases including Alzheimer’s and Parkinson’s diseases, and cerebral ischemia. Although the underlying neuroprotective mechanism of Silymarin is believed to be due to its capacity to inhibit oxidative stress in the brain, it also confers additional advantages by influencing pathways such as b-amyloid aggregation, inflammatory mechanisms, cellular apoptotic machinery, and estrogenic receptor mediation. In this review, we have elucidated the possible neuroprotective effects of Silymarin and the underlying
molecular events, and suggested future courses of action for its acceptance as a CNS drug for the treatment of neurodegenerative diseases
Effect of corchorusin-D, a saikosaponin like compound, on B16F10 melanoma cells (in vitro and in vivo)
No full textCorchorusin-D (COR-D), isolated from Corchorus acutangulus, was reported to induce
apoptosis in leukemic cells. However, no studies concerning its activity on melanoma
cells have been reported. We have evaluated its in vitro anti-cancer activity on
melanoma cells (B16F10, SK-MEL-28, and A375). The results demonstrate that CORD
showed maximum inhibition of B16F10 cells in vitro. COR-D induced
mitochondrial dysfunction and altered the Bax/Bcl-2 ratio with down regulation of
pro-caspases 9 and activation of caspase 3 in B16F10 cells, triggering intrinsic pathway
of apoptosis. Moreover, it inhibited the in vivo B16F10 tumor growth and increased the
survival rate of mice. Greater number of Annexin V-FITC and propidium iodide (PI)-
positive tumor cells signified that COR-D induced apoptosis in vivo also. The reduction
in tumor growth is well correlated with decreased microvascular density of the tumor
cells in treated mice. In conclusion, this study reveals that COR-D-induced
mitochondrial dysfunction is responsible for the induction of apoptotic cell deat
Binding of Isoquinoline Alkaloids Berberine, Palmatine and Coralyne to Hemoglobin: Sructural and Thermodynamic Characterization Studies
No full textBerberine, palmatine and coralyne, the isoquinoline alkaloids distributed in many botanical families, are extensively investigated due to their potential therapeutic actions and clinical utilities. In this work, their binding characteristics to hemoglobin (Hb) were studied by UV-vis absorption spectroscopy, fluorescence spectroscopy, circular dichroism spectroscopy, isothermal calorimetric titration and differential scanning calorimetric techniques. The results indicated that all the three alkaloids caused strong fluorescence quenching of Hb by the static quenching mechanism, but with differing quenching efficiencies. There was a single binding site on Hb for these alkaloids. According to the theory of Fo¨rster resonance energy transfer, the binding distances between b-Trp37 of Hb and berberine, palmatine and coralyne were evaluated to be 2.78 nm, 2.64 nm and 3.29 nm, respectively. The result of synchronous fluorescence, circular dichroism and 3D fluorescence revealed that the polarity around Trp residues experienced a significant increase in the presence of alkaloids. The binding was favoured by enthalpy and entropy changes. Results of circular dichroism, 3D and synchronous fluorescence studies confirmed that the binding of the alkaloids significantly changed the secondary structure of Hb. The studies revealed that berberine and palmatine bound to a site near to the a1b2 interface on Hb different than coralyne but the affinity of coralyne was one order higher than that of berberine and palmatine
