Design,synthesis and anticancer activity of piperazine hydroxamates and their histone deacetylase (HDAC) inhibitory activity

Six compounds were synthesized with piperazine in linker region and hydroxamate as Zinc Binding Group (ZBG). They were screened against three cancer cell-lines (NCIH460; HCT116; U251). Compounds 5c and 5f with GI₅₀ value of 9.33 ± 1.3 μM and 12.03 ± 4 μM, respectively, were tested for their inhibitory potential on hHDAC8. Compound 5c had IC₅₀ of 33.67 μM. Compounds were also screened for their anticancer activity against HL60 human promyelocytic leukemia cell line due to the presence of pharmacophoric features of RR inhibitors in them. Compound 5c had IC₅₀ of 0.6 μM at 48 h.     

Chemical scaffolds with structural similarities to siderophores of nonribosomal peptide-polyketide origin as novel antimicrobials against Mycobacterium tuberculosis and Yersinia pestis

Mycobacterium tuberculosis (Mtb) and Yersinia pestis (Yp) produce siderophores with scaffolds of nonribosomal peptide-polyketide origin. Compounds with structural similarities to these siderophores were synthesized and evaluated as antimicrobials against Mtb and Yp under iron-limiting conditions mimicking the iron scarcity these pathogens encounter in the host and under standard iron-rich conditions. Several new antimicrobials were identified, including some with increased potency in the iron-limiting condition. Our study illustrates the possibility of screening compound libraries in both iron-rich and iron-limiting conditions to identify antimicrobials that may selectively target iron scarcity-adapted bacteria and highlights the usefulness of building combinatorial libraries of compounds having scaffolds with structural similarities to siderophores to feed into antimicrobial screening programs.     

Raptor-rictor axis in TGFbeta-induced protein synthesis

Transforming growth factor-beta (TGFbeta) stimulates pathological renal cell hypertrophy for which increased protein synthesis is critical. The mechanism of TGFbeta-induced protein synthesis is not known, but PI 3 kinase-dependent Akt kinase activity is necessary. We investigated the contribution of downstream effectors of Akt in TGFbeta-stimulated protein synthesis. TGFbeta increased inactivating phosphorylation of Akt substrate tuberin in a PI 3 kinase/Akt dependent manner, resulting in activation of mTOR kinase. mTOR activity increased phosphorylation of S6 kinase and the translation repressor 4EBP-1, which were sensitive to inhibition of both PI 3 kinase and Akt. mTOR inhibitor rapamycin and a dominant negative mutant of mTOR suppressed TGFbeta-induced phosphorylation of S6 kinase and 4EBP-1. PI 3 kinase/Akt and mTOR regulated dissociation of 4EBP-1 from eIF4E to make the latter available for binding to eIF4G. mTOR and 4EBP-1 modulated TGFbeta-induced protein synthesis. mTOR is present in two multi protein complexes, mTORC1 and mTORC2. Raptor and rictor are part of mTORC1 and mTORC2, respectively. shRNA-mediated downregulation of raptor inhibited TGFbeta-stimulated mTOR kinase activity, resulting in inhibition of phosphorylation of S6 kinase and 4EBP-1. Raptor shRNA also prevented protein synthesis in response to TGFbeta. Downregulation of rictor inhibited serine 473 phosphorylation of Akt without any effect on phosphorylation of its substrate, tuberin. Furthermore, rictor shRNA increased phosphorylation of S6 kinase and 4EBP-1 in TGFbeta-independent manner, resulting in increased protein synthesis. Thus mTORC1 function is essential for TGFbeta-induced protein synthesis. Our data also provide novel evidence that rictor negatively regulates TORC1 activity to control basal protein synthesis, thus conferring tight control on cellular hypertrophy.     

Effects of nonionic surfactant on hydrolysis and fermentation of protein rich tannery solid waste

The untanned proteinaceous tannery solid waste, the animal fleshing (ANFL), was used as substrate in the treatment process (hydrolysis and fermentation) involving Synergistes sp. The nonionic surfactant (Tween 80) was evaluated for its ability to influence on microbial growth and enzyme activity in the hydrolysis and fermentation of ANFL. The addition of Tween 80 in the process significantly increased the activities of hydrolytic and fermentative enzymes like protease (338-360 Um l(-1)) and deaminase (187-206 Um l(-1)) compared to that of control (protease 195-220 Um l(-1) and deaminase 70-83 Um l(-1)). The total viable bacterial count was increased more than twofold, compared to the control in the presence of 0.15% Tween 80. The ANFL fermentation and formation of other metabolites were evidenced by Gas Chromatography and Mass Spectroscopy (GC-MS), Proton Nuclear Magnetic Resonance spectroscopy ((1)H NMR) and Fourier transform infra red spectroscopy (FT-IR). The breakdown of fibrillar proteins in ANFL was confirmed by the scanning electron microscopy (SEM) and the transmission electron microscopy (TEM).     

Toxicity analysis of N- and C-terminus-deleted vegetative insecticidal protein from Bacillus thuringiensis.

A vegetative insecticidal protein (VIP)-encoding gene from a local isolate of Bacillus thuringiensis has been cloned, sequenced, and expressed in Escherichia coli. The expressed protein shows insecticidal activity against several lepidopteran pests but is ineffective against Agrotis ipsilon. Comparison of the amino acid sequence with those of reported VIPs revealed a few differences. Analysis of insecticidal activity with N- and C-terminus deletion mutants suggests a differential mode of action of VIP against different pests.     

Investigation on interaction of tannic acid with type I collagen and its effect on thermal,enzymatic, and conformational stability for tissue engineering applications

Collagen is an essential component of tissues, which is the most abundant component in extracellular matrix and highly conserved across the animal kingdom. It can assemble into fiber and play an essential role in cell adhesion and growth and could be extremely useful in tissue engineering. In this study, the effect of tannic acid (TA) on the thermal, enzymatic and conformational stability of type I collagen has been investigated for the development of collagen‐based biomaterials. Interaction of TA with collagen demonstrates the role of hydrogen bonding and hydrophobic interaction in providing the thermal and enzymatic stability. Thermal analysis studies reveal that, hydrothermal stability of collagen increases as well as inhibits the breakdown of collagenase by formation of hydrogen bonds and hydrophobic interactions. TA binds to the collagen with high affinity because the structural flexibility of the collagen compensates for the structural rigidity of the phenolics. Increase in concentration of TA induces significant change in the conformation of triple helix. The free binding energy of TA with collagen‐like peptide was determined to be in the range of ?9.4 to ?11.2 kcal mol^?1, which was calculated by using Autodock Vina software and showed numerous hydrophobic and hydrogen bond interactions. We anticipate that these collagen‐based biomaterials hold great potential for biomedical applications. © 2013 Wiley Periodicals, Inc. Biopolymers 101: 471–483, 2014.     

The gag gene products of human immunodeficiency virus type 1: alignment within the gag open reading frame, identification of posttranslational modifications, and evidence for alternative gag precursors.

Seven human immunodeficiency virus gag polypeptides were identified in the purified virus and in infected CD4+ lymphocytes by peptide mapping and limited amino acid sequencing of immune-purified proteins. Two gag polyproteins of 55,000 (p55) and 41,000 (p41) daltons were rapidly labeled and readily processed into the major internal gag proteins that were aligned within the gag open reading frame (ORF) as NH2-p16 (MA)-p24 (CA)-p9 (NC)-p7-COOH. The myristoylated p16 (matrix, MA) protein was processed from the myristoylated p55 gag precursor protein. The immunoreactivity of the p16 (MA) protein with region-specific gag antisera and the conservation of the N-terminal myristyl group of the p55 precursor protein in p16 (MA) confirmed its position as the N-terminal-most protein. The p9 (nucleocapsid, NC) protein was localized to residue 378 of the gag ORF, next to the C terminus of the p24/p25 (core antigen, CA) protein. The p9 protein had a repeating Cys residue containing motif which is found in the nucleic acid-binding Cys residue-containing proteins of retroviruses. The p24 (CA) protein, which was localized to residue 133 of the gag ORF, was apparently derived by C-terminal processing of an intermediate polypeptide, p25. Both the mature p24 (CA) and p16 (MA) proteins were phosphorylated at Ser residue(s). We also identified two forms of gag p41 species, one resulting from the C-terminal processing of p55 and the other originating either from N-terminal processing of p55 or from de novo synthesis.     

Distinct mechanisms of agonist-induced endocytosis for human chemokine receptors CCR5 and CXCR4

Desensitization of the chemokine receptors, a large class of G protein-coupled receptors, is mediated in part by agonist-driven receptor endocytosis. However, the exact pathways have not been fully defined. Here we demonstrate that the rate of ligand-induced endocytosis of CCR5 in leukocytes and expression systems is significantly slower than that of CXCR4 and requires prolonged agonist treatment, suggesting that these two receptors use distinct mechanisms. We show that the C-terminal domain of CCR5 is the determinant of its slow endocytosis phenotype. When the C-tail of CXCR4 was exchanged for that of CCR5, the resulting CXCR4-CCR5 (X4-R5) chimera displayed a CCR5-like trafficking phenotype. We found that the palmitoylated cysteine residues in this domain anchor CCR5 to plasma membrane rafts. CXCR4 and a C-terminally truncated CCR5 mutant (CCR5-KRFX) lacking these cysteines are not raft associated and are endocytosed by a clathrin-dependent pathway. Genetic inhibition of clathrin-mediated endocytosis demonstrated that a significant fraction of ligand-occupied CCR5 trafficked by clathrin-independent routes into caveolin-containing vesicular structures. Thus, the palmitoylated C-tail of CCR5 is the major determinant of its raft association and endocytic itineraries, differentiating it from CXCR4 and other chemokine receptors. This novel feature of CCR5 may modulate its signaling potential and could explain its preferential use by HIV for person-to-person transmission of disease.     

An AFLP-based database of Shigella flexneri and Shigella sonnei isolates and its use for the identification of untypable Shigella strains

Amplified fragment length polymorphism (AFLP) can be used to assess the genetic diversity of closely related microbial genomes. In this study, the first of its kind for identification of Shigella, the high discriminatory power of AFLP has been used to determine the genetic relatedness of 230 isolates of Shigella flexneri and Shigella sonnei strains. An AFLP database was generated to demonstrate its utility in the discrimination of closely related strains. Based on AFLP, S. flexneri strains could be grouped into separate clusters according to their serotypes. Within each serotype, strains demonstrated 80-100% similarity indicating that identical strains and closely related strains could be distinguished by this technique. S. flexneri 6 formed a distinct cluster with 55% similarity to the rest of the S. flexneri strains showing significant divergence from the rest of the S. flexneri strains. Significantly, S. sonnei isolates formed a distinct group and showed approximately the same level of genetic linkage to S. flexneri as Escherichia coli strains. Untypable isolates that showed conflicting agglutination reactions with conventional typing sera were identifiable by AFLP. Thus AFLP can be used for genetic fingerprinting of Shigella strains and aid in the identification of variant untypable isolates.     

Characterization of sex-based differences in the mechanical properties of human finger glabrous tissue using a fiberoptic sensor

TAC-Cell is a custom-built somatosensory stimulator that delivers pneumatic cutaneous tactile inputs to virtually any skin target on the body and by virtue of its non-ferrous materials is compatible with functional magnetic resonance imaging (fMRI) and magnetoencephalography (MEG) brain scanners. In this study, we describe the method to measure apparent skin displacement induced by TAC-Cell stimulation of the glabrous surface of the distal phalanx of the index finger. Specifically, we studied the effect of four servo controller input voltages (0.4 V–1.0 V) on resultant skin displacement among eighteen, neurotypical adult male and female participants. A fiberoptic displacement sensor, commonly used in industrial applications, was coupled to the TAC-Cell to measure the glabrous skin?s kinematic response to different stimulus amplitudes. Skin displacement was significantly dependent on stimulus amplitudes and sex (p<0.0001). Power spectrum and kinematic analysis of skin displacement showed that the pneumatic TAC-Cell stimulus consists of a spectrally rich, high velocity signal. In related work, we have shown that this dynamic pneumocutaneous stimulus is highly effective in evoking a cortical brain response for neurodiagnostic applications and somatosensory pathway analysis in health and disease.