Shantinath Indi

Icosahedral DNA Nanocapsules by Modular Assembly

Its a trap! DNA polyhedra formed through molecular self-assembly may function as nanocapsules for the targeted delivery of encapsulated entities. This functional aspect was demonstrated for the most complex DNA-based platonic solid: During the stepwise amalgamation of discrete polyhedra to form icosahedra, gold nanoparticles (GNPs) were encapsulated from solution (see illustration and TEM image of icosahedral cages containing GNPs).

Hydrophobic Pockets in a Nonpolymeric Aqueous Gel: Observation of such a Gelation Process by Color Change

Efficient gelation of aqueous fluids by a novel tripodal trischolamide generates chiral hydrophobic pockets in the gel network. A yellow to green color change (see absorbance spectra) in the presence of bromophenol blue (BPBH) indicates the formation of the gel. The bound (anionic) dye (BPB- ) shows induced circular dichroism, which is indicative of a chiral environment.

SPR and ITC determination of the kinetics and the thermodynamics of bivalent versus monovalent sugar ligand–lectin interactions

A kinetic study of the interaction of bivalent and monovalent sugar ligands with a lectin was undertaken with the aid of surface plasmon resonance (SPR) method. The study involved a series of bivalent α-d-mannopyranoside containing sugar ligands, with systematic variation in the distance between the sugar ligands. The detailed kinetic studies showed that bivalent ligands underwent a faster association (kon) and a slower dissociation (koff) of the ligand–lectin complexes, in comparison to the monovalent ligand–lectin complexes. The kinetic constants were complemented further by assessing the thermodynamic parameters with the aid of isothermal titration calorimetry (ITC). The initiation of cross-linking of ligand–lectin interactions emerge from the early stages of the complexation. The dynamic light scattering (DLS) and the transmission electron microscopy (TEM) techniques allowed judging the sizes and morphologies of the complex in the solution and solid states, respectively.

New evidences on Tau-DNA interactions and relevance to neurodegeneration §

Tau is mainly distributed in cytoplasm and also found to be localized in the nucleus. There is limited data on DNA binding potential of Tau. We provide novel evidence on nicking of DNA by Tau. Tau nicks the supercoiled DNA leading to open circular and linear forms. The metal ion magnesium (a co-factor for endonuclease) enhanced the Tau DNA nicking ability, while an endonuclease specific inhibitor, aurinetricarboxylic acid (ATA) inhibited the Tau DNA nicking ability. Further, we also evidenced that Tau induces B-C-A mixed conformational transition in DNA and also changes DNA stability. Tau-scDNA complex is more sensitive to DNAse I digestion indicating stability changes in DNA caused by Tau. These findings indicate that Tau alters DNA helicity and integrity and also nicks the DNA. The relevance of these novel intriguing findings regarding the role Tau in neuronal dysfunction is discussed.

Regulation of lipid biosynthesis, sliding motility, and biofilm formation by a membrane-anchored nucleoid-associated protein of Mycobacterium tuberculosis.

Bacteria use a number of small basic proteins for organization and compaction of their genomes. By their interaction with DNA, these nucleoid-associated proteins (NAPs) also influence gene expression. Rv3852, a NAP of Mycobacterium tuberculosis, is conserved among the pathogenic and slow-growing species of mycobacteria. Here, we show that the protein predominantly localizes in the cell membrane and that the carboxy-terminal region with the propensity to form a transmembrane helix is necessary for its membrane localization. The protein is involved in genome organization, and its ectopic expression in Mycobacterium smegmatis resulted in altered nucleoid morphology, defects in biofilm formation, sliding motility, and change in apolar lipid profile. We demonstrate its crucial role in regulating the expression of KasA, KasB, and GroEL1 proteins, which are in turn involved in controlling the surface phenotypes in mycobacteria.

In vitro Evidence that an Aqueous Extract of Centella asiatica Modulates α-Synuclein Aggregation Dynamics

α-Synuclein aggregation is one of the major etiological factors implicated in Parkinsons disease (PD). The prevention of aggregation of α-synuclein is a potential therapeutic intervention for preventing PD. The discovery of natural products as alternative drugs to treat PD and related disorders is a current trend. The aqueous extract of Centella asiatica (CA) is traditionally used as a brain tonic and CA is known to improve cognition and memory. There are limited data on the role of CA in modulating amyloid-β (Aβ) levels in the brain and in Aβ aggregation. Our study focuses on CA as a modulator of the α-synuclein aggregation pattern in vitro. Our investigation is focused on: (i) whether the CA leaf aqueous extract prevents the formation of aggregates from monomers (Phase I: α-synuclein + extract co-incubation); (ii) whether the CA aqueous extract prevents the formation of fibrils from oligomers (Phase II: extract added after oligomers formation); and (iii) whether the CA aqueous extract disintegrates the pre-formed fibrils (Phase III: extract added to mature fibrils and incubated for 9 days). The aggregation kinetics are studied using a thioflavin-T assay, circular dichroism, and transmission electron microscopy. The results showed that the CA aqueous extract completely inhibited the α-synuclein aggregation from monomers. Further, CA extract significantly inhibited the formation of oligomer to aggregates and favored the disintegration of the preformed fibrils. The study provides an insight in finding new natural products for future PD therapeutics.

Regulation of Growth, Cell Shape, Cell Division, and Gene Expression by Second Messengers (p)ppGpp and Cyclic Di-GMP in Mycobacterium smegmatis

UNLABELLED The alarmone (p)ppGpp regulates transcription, translation, replication, virulence, lipid synthesis, antibiotic sensitivity, biofilm formation, and other functions in bacteria. Signaling nucleotide cyclic di-GMP (c-di-GMP) regulates biofilm formation, motility, virulence, the cell cycle, and other functions. In Mycobacterium smegmatis, both (p)ppGpp and c-di-GMP are synthesized and degraded by bifunctional proteins Rel(Msm) and DcpA, encoded by rel(Msm) and dcpA genes, respectively. We have previously shown that the Δrel(Msm) and ΔdcpA knockout strains are antibiotic resistant and defective in biofilm formation, show altered cell surface properties, and have reduced levels of glycopeptidolipids and polar lipids in their cell wall (K. R. Gupta, S. Kasetty, and D. Chatterji, Appl Environ Microbiol 81:2571-2578, 2015,http://dx.doi.org/10.1128/AEM.03999-14). In this work, we have explored the phenotypes that are affected by both (p)ppGpp and c-di-GMP in mycobacteria. We have shown that both (p)ppGpp and c-di-GMP are needed to maintain the proper growth rate under stress conditions such as carbon deprivation and cold shock. Scanning electron microscopy showed that low levels of these second messengers result in elongated cells, while high levels reduce the cell length and embed the cells in a biofilm-like matrix. Fluorescence microscopy revealed that the elongated Δrel(Msm) and ΔdcpA cells are multinucleate, while transmission electron microscopy showed that the elongated cells are multiseptate. Gene expression analysis also showed that genes belonging to functional categories such as virulence, detoxification, lipid metabolism, and cell-wall-related processes were differentially expressed. Our results suggests that both (p)ppGpp and c-di-GMP affect some common phenotypes in M. smegmatis, thus raising a possibility of cross talk between these two second messengers in mycobacteria. IMPORTANCE Our work has expanded the horizon of (p)ppGpp and c-di-GMP signaling in Gram-positive bacteria. We have come across a novel observation that M. smegmatis needs (p)ppGpp and c-di-GMP for cold tolerance. We had previously shown that the Δrel(Msm) and ΔdcpA strains are defective in biofilm formation. In this work, the overproduction of (p)ppGpp and c-di-GMP encased M. smegmatis in a biofilm-like matrix, which shows that both (p)ppGpp and c-di-GMP are needed for biofilm formation. The regulation of cell length and cell division by (p)ppGpp was known in mycobacteria, but our work shows that c-di-GMP also affects the cell size and cell division in mycobacteria. This is perhaps the first report of c-di-GMP regulating cell division in mycobacteria.

In vitro polymerization of Mycobacterium leprae FtsZ OR Mycobacterium tuberculosis FtsZ is revived or abolished, respectively, by reciprocal mutation of a single residue

A single residue that dramatically influences polymerization of principal cell division protein FtsZ of Mycobacterium leprae (MlFtsZ) and Mycobacterium tuberculosis (MtFtsZ) has been identified. Soluble, recombinant MlFtsZ did not show polymerization in vitro, in contrast to MtFtsZ, which polymerised. Mutation of the lone non-conserved residue T172 in the N-terminal domain of MlFtsZ to A172, as it exists in MtFtsZ, showed dramatic polymerization of MlFtsZ-T172A in vitro. Reciprocal mutation of A172 in MtFtsZ to T172, as it exists in MlFtsZ, abolished polymerization of MtFtsZ-A172T in vitro. While T172A mutation enhanced weak GTPase activity of MlFtsZ, reciprocal A172T mutation marginally reduced GTPase activity of MtFtsZ in vitro. These observations demonstrate that the residue at position 172 plays critical role in the polymerization of MlFtsZ and MtFtsZ. A possible evolutionary correlation between the presence of polymerization-adversive or polymerization-favouring residue at position 172 in FtsZ and generation time of the respective bacterium are discussed.