Aspects of tautomerism. 8. Solvolysis of some pseudo and normal acid chlorides

Pseudo acid chlorides derived from levulinic acid ando-benzoyl-benzoic acid, solvolyse in aqueous acetone, aqueous dioxane and aqueous dimethylformamide by aS Nl process. Their reaction pattern is distinct from that of typical normal acid chlorides, viz.,p-benzoylbenzoyl chloride and fluorene-9-one-1-carboxylic acid chloride, which solvolyse by aS N2 pathway. No evidence for tautomerism could be obtained either between the normal and pseudo forms of the acid chlorides or the derived ion pairs.

Analysis of the amino acid sequences of plant Bowman-Birk inhibitors

Plant seeds contain a large number of protease inhibitors of animal, fungal, and bacterial origin. One of the well-studied families of these inhibitors is the Bowman-Birk family(BBI). The BBIs from dicotyledonous seeds are 8K, double-headed proteins. In contrast, the 8K inhibitors from monocotyledonous seeds are single headed. Monocots also have a 16K, double-headed inhibitor. We have determined the primary structure of a Bowman-Birk inhibitor from a dicot, horsegram, by sequential edman analysis of the intact protein and peptides derived from enzymatic and chemical cleavage. The 76-residue-long inhibitor is very similar to that ofMacrotyloma axillare. An analysis of this inhibitor along with 26 other Bowman-Birk inhibitor domains (MW 8K) available in the SWISSPROT databank revealed that the proteins from monocots and dicots belong to related but distinct families. Inhibitors from monocots show larger variation in sequence. Sequence comparison shows that a crucial disulphide which connects the amino and carboxy termini of the active site loop is lost in monocots. The loss of a reactive site in monocots seems to be correlated to this. However, it appears that this disulphide is not absolutely essential for retention of inhibitory function. Our analysis suggests that gene duplication leading to a 16K inhibitor in monocots has occurred, probably after the divergence of monocots and dicots, and also after the loss of second reactive site in monocots.

The hydrogen bond: a molecular beam microwave spectroscopist's view with a universal appeal

In this manuscript, we propose a criterion for a weakly bound complex formed in a supersonic beam to be characterized as a `hydrogen bonded complex'. For a `hydrogen bonded complex', the zero point energy along any large amplitude vibrational coordinate that destroys the orientational preference for the hydrogen bond should be significantly below the barrier along that coordinate so that there is at least one bound level. These are vibrational modes that do not lead to the breakdown of the complex as a whole. If the zero point level is higher than the barrier, the `hydrogen bond' would not be able to stabilize the orientation which favors it and it is no longer sensible to characterize a complex as hydrogen bonded. Four complexes, Ar-2-H2O, Ar-2-H2S, C2H4-H2O and C2H4-H2S, were chosen for investigations. Zero point energies and barriers for large amplitude motions were calculated at a reasonable level of calculation, MP2(full)/aug-cc-pVTZ, for all these complexes. Atoms in molecules (AIM) theoretical analyses of these complexes were carried out as well. All these complexes would be considered hydrogen bonded according to the AIM theoretical criteria suggested by Koch and Popelier for C-H center dot center dot center dot O hydrogen bonds (U. Koch and P. L. A. Popelier, J. Phys. Chem., 1995, 99, 9747), which has been widely and, at times, incorrectly used for all types of contacts involving H. It is shown that, according to the criterion proposed here, the Ar-2-H2O/H2S complexes are not hydrogen bonded even at zero kelvin and C2H4-H2O/H2S complexes are. This analysis can naturally be extended to all temperatures. It can explain the recent experimental observations on crystal structures of H2S at various conditions and the crossed beam scattering studies on rare gases with H2O and H2S.

Absence of parallelism between polyamine and nucleic acid contents during induced growth of cucumber cotyledons

The cytokinins (benzyladenine or benzyladenosine) decreased spermidine and spermine contents despite increasing putrescine content, when administered to isolated cotyledons of Cucumis sativus L. var. Guntur in organ culture. KCl decreased putrescine contents, although marginally increasing polyamine contents. The cytokinins and/or KCl augmented nucleic acid biosynthesis and accumulation, resulting in enhanced growth and differentiation of the isolated cotyledons. These observations show that polyamine accumulation and growth are not always coupled.

Solution NMR studies of acetohydroxy acid synthase I: Identification of the sites of inter-subunit interactions using multidimensional NMR methods

The novel multidomain organization in the multimeric Escherichia coli AHAS I (ilvBN) enzyme has been dissected to generate polypeptide fragments. These fragments when cloned, expressed and purified reassemble in the presence of cofactors to yield a catalytically competent enzyme. Structural characterization of AHAS has been impeded due to the fact that the holoenzyme is prone to dissociation leading to heterogeneity in samples. Our approach has enabled the structural characterization using high-resolution nuclear magnetic resonance methods. Near complete sequence specific NMR assignments for backbone H-N, N-15, C-13 alpha and C-13(beta) atoms of the FAD binding domain of ilvB have been obtained on samples isotopically enriched in H-2, C-13 and N-15. The secondary structure determined on the basis of observed C-13(alpha) secondary chemical shifts and sequential NOEs indicates that the secondary structure of the FAD binding domain of E. coli AHAS large Subunit (ilvB) is similar to the structure of this domain in the catalytic subunit of yeast AHAS. Protein-protein interactions involving the regulatory subunit (ilvN) and the domains of the catalytic subunit (ilvB) were studied using circular dichroic and isotope edited solution nuclear magnetic resonance spectroscopic methods. Observed changes in circular dichroic spectra indicate that the regulatory subunit (ilvN) interacts with ilvB alpha and ilvB beta domains of the catalytic subunit and not with the ilvB gamma domain. NMR chemical shift mapping methods show that ilvN binds close to the FAD binding site in ilvB beta and proximal to the intrasubunit ilvB alpha/ilvB beta domain interface. The implication of this interaction on the role of the regulatory subunit oil the activity of the holoenzyme is discussed. NMR studies of the regulatory domains show that these domains are structured in solution. Preliminary evidence for the interaction of ilvN with the metabolic end product of the pathway, viz., valine is also presented.

Gabapentin: A Stereochemically Constrained gamma Amino Acid Residue in Hybrid Peptide Design

Nature has used the all-alpha-polypeptide backbone of proteins to create a remarkable diversity of folded structures. Sequential patterns of 20 distinct amino adds, which differ only in their side chains, determine the shape and form of proteins. Our understanding of these specific secondary structures is over half a century old and is based primarily on the fundamental elements: the Pauling alpha-helix and beta-sheet. Researchers can also generate structural diversity through the synthesis of polypeptide chains containing homologated (omega) amino acid residues, which contain a variable number of backbone atoms. However, incorporating amino adds with more atoms within the backbone introduces additional torsional freedom into the structure, which can complicate the structural analysis. Fortunately, gabapentin (Gpn), a readily available bulk drug, is an achiral beta,beta-disubstituted gamma amino add residue that contains a cyclohexyl ring at the C-beta carbon atom, which dramatically limits the range of torsion angles that can be obtained about the flanking C-C bonds. Limiting conformational flexibility also has the desirable effect of increasing peptide crystallinity, which permits unambiguous structural characterization by X-ray diffraction methods. This Account describes studies carried out in our laboratory that establish Gpn as a valuable residue in the design of specifically folded hybrid peptide structures. The insertion of additional atoms into polypeptide backbones facilitates the formation of intramolecular hydrogen bonds whose directionality is opposite to that observed in canonical alpha-peptide helices. If hybrid structures mimic proteins and biologically active peptides, the proteolytic stability conferred by unusual backbones can be a major advantage in the area of medicinal chemistry. We have demonstrated a variety of internally hydrogen-bonded structures in the solid state for Gpn-containing peptides, including the characterization of the C-7 and C-9 hydrogen bonds, which can lead to ribbons in homo-oligomeric sequences. In hybrid alpha gamma sequences, district C-12 hydrogen-bonded turn structures support formation of peptide helices and hairpins in longer sequences. Some peptides that include the Gpn residue have hydrogen-bond directionality that matches alpha-peptide helices, while others have the opposite directionality. We expect that expansion of the polypeptide backbone will lead to new classes of foldamer structures, which are thus far unknown to the world of alpha-polypeptides. The diversity of internally hydrogen-bonded structures observed in hybrid sequences containing Gpn shows promise for the rational design of novel peptide structures incorporating hybrid backbones.

Encapsulation and release of rifampicin using poly(vinyl pyrrolidone)-poly (methacrylic acid) polyelectrolyte capsules

Poly(vinyl pyrrolidone) and poly(methacrylic acid) multilayer capsules based on hydrogen bonding have been prepared by the layer-by-layer approach and used to encapsulate and release rifampicin, an antituberculosis drug. Removal of silica core using a buffer of ammonium fluoride and hydrofluoric acid at about pH 3 was found to produce better capsules than hydrofluoric acid alone. An eight-layered capsule had a wall thickness of 20 rim. Maximum encapsulation was found to be about 86 mu g at 40 degrees C with 1 +/- 0.2 x 10(6) capsules. Release studies showed a burst kind of release and maximum release was obtained above pH 7 where the capsules disintegrate rapidly thereby releasing the drug in a short period. Interactions studies with Mycobacterium smegmatis showed that the capsules were cytocompatible and the released drug functioned with the same efficacy as the free drug.

Studies on boundary lubrication properties of oxidised coconut and soy bean oils

Vegetable oils are a potential source of base oils for biodegradable lubricants, with limited oxidative stability. This study focuses on the effect of long-term ageing and the influence of oxidation products on the boundary lubrication performance of coconut and soy bean oils, by subjecting them to accelerated ageing in a dark oven at elevated temperature. The samples were collected at regular intervals and analysed for the changes in viscosity, percentage of free fatty acid and peroxide number compared to fresh oil samples. The boundary lubrication properties of these samples were evaluated using a four-ball tester. Increased wear observed with aged oil samples was linked to the destruction of triglyceride structure and formation of peroxides. The difference in the wear properties of soy bean oil to coconut oil was accounted by its high content of unsaturated fatty acids and its susceptibility to undergo oxidation. It was concluded that the coconut oil can perform as a better lubricant and has got a better storage life compared to soy bean oil.

NUVIEW:software for display and interactive manipulation of nucleic acid models

The NUVIEW software package allows skeletal models of any double helical nucleic acid molecule to be displayed out a graphics monitor and to apply various rotations, translations and scaling transformations interactively, through the keyboard. The skeletal model is generated by connecting any pair of representative points, one from each of the bases in the basepair. In addition to the above mentioned manipulations, the base residues can be identified by using a locator and the distance between any pair of residues can be obtained. A sequence based color coded display allows easy identification of sequence repeats, such as runs of Adenines. The real time interactive manipulation of such skeletal models for large DNA/RNA double helices, can be used to trace the path of the nucleic acid chain in three dimensions and hence get a better idea of its topology, location of linear or curved regions, distances between far off regions in the sequence etc. A physical picture of these features will assist in understanding the relationship between base sequence, structure and biological function in nucleic acids.

NUPARM and NUCGEN: software for analysis and generation of sequence dependent nucleic acid structures

Software packages NUPARM and NUCGEN, are described, which can be used to understand sequence directed structural variations in nucleic acids, by analysis and generation of non-uniform structures. A set of local inter basepair parameters (viz. tilt, roll, twist, shift, slide and rise) have been defined, which use geometry and coordinates of two successive basepairs only and can be used to generate polymeric structures with varying geometries for each of the 16 possible dinucleotide steps. Intra basepair parameters, propeller, buckle, opening and the C6...C8 distance can also be varied, if required, while the sugar phosphate backbone atoms are fixed in some standard conformation ill each of the nucleotides. NUPARM can be used to analyse both DNA and RNA structures, with single as well as double stranded helices. The NUCGEN software generates double helical models with the backbone fixed in B-form DNA, but with appropriate modifications in the input data, it can also generate A-form DNA ar rd RNA duplex structures.

Identification of alpha- and beta-Hydroxy Acid Containing Cyclodepsipeptides in Natural Peptide Mixtures Using Negative Ion Mass Spectrometry

Natural peptide libraries often contain cyclodepsipeptides containing alpha or beta hydroxy residues. Extracts of fungal hyphae of Isaria yield a microheterogenous cyclodepsipeptide mixture in which two classes of molecules can be identified by mass spectral fragmentation of negative ions. In the case of isaridins, which contain an alpha-hydroxy residue and a beta-amino acid residue, a characteristic product ion corresponding to a neutral loss of 72 Da is obtained. hi addition, neutral loss of water followed by a 72 Da loss is also observed. Two distinct modes of fragmentation rationalize the observed product ion distribution. The neutral loss of 72 Da has also been obtained for a roseotoxin component, which is also an alpha-hydroxy residue containing cyclodepsipeptide. In the case of isariins, which contain a beta-hydroxy acid residue, ring opening and subsequent loss of the terminal residue as an unsaturated ketene fragment, rationalizes the observed product ion formation. Fragmentation of negative ions provide characteristic neutral losses, which are diagnostic of the presence of alpha-hydroxy or beta-hydroxy residues.

Microwave absorption studies of iron encapsulating carbon-polymer nanocomposite films

Iron encapsulated carbon nanoparticle polyvinyl chloride composite films have been prepared by solvent mixing and drying method. The films were characterized by scanning electron microscope (SEM) and high resolution transmission electron microscope (HRTEM). A 5 nm thin graphitic carbon coating is observed on cubic Fe nanoparticles. The microwave absorption studies by wave guide technique in the Ka band range showed highest electromagnetic interference shielding efficiency of 18dB on a 300 micron thick film. The shielding efficiency depends on weight % of the filler in the composite. The data obtained for different films indicate that these lightweight materials are good candidates for potential electromagnetic interference shielding applications.

Surfactant-Mediated Synthesis of Functional Metal Oxide Nanostructures Via Microwave Irradiation-Assisted Chemical Synthesis

In the present work we report a rapid microwave irradiation-assisted chemical synthesis technique for the growth of nanoparticles, nanorods, and nanotubes of a variety of metal oxides in the presence of an appropriate surfactant (cationic, anionic, non ionic and polymeric), without the use of any templates. The method is simple, inexpensive, and helps one to prepare nanostructures in quick time, measured in seconds and minutes. This method has been applied successfully to synthesize nanostructures of a variety of binary and ternary metal oxides such as ZnO, CdO, Fe2O3, CuO, Ga2O3, Gd2O3, ZnFe2O4, etc. There is an observed variation in the morphology of the nanostructures with changes in different process parameters, such as microwave power, irradiation time, identity of solvent, type of surfactant, and its concentration.

HDAC inhibitor valproic acid enhances tumor cell kill in adenovirus-HSVtk mediated suicide gene therapy in HNSCC xenograft mouse model

Safety, efficacy and enhanced transgene expression are the primary concerns while using any vector for gene therapy. One of the widely used vectors in clinical. trials is adenovirus which provides a safe way to deliver the therapeutic gene. However, adenovirus has poor transduction efficiency in vivo since most tumor cells express low coxsackie and adenovirus receptors. Similarly transgene expression remains low, possibly because of the chromatization of adenoviral genome upon infection in eukaryotic cells, an effect mediated by histone deacetylases (HDACs). Using a recombinant adenovirus (Ad-HSVtk) carrying the herpes simplex thymidine kinase (HSVtk) and GFP genes we demonstrate that HDAC inhibitor valproic acid can bring about an increase in CAR expression on host cells and thereby enhanced Ad-HSVtk infectivity. It also resulted in an increase in transgene (HSVtk and GFP) expression. This, in turn, resulted in increased cell kill of HNSCC cells, following ganciclovir treatment in vitro as well as in vivo in a xenograft nude mouse model.

Microbial aspects of acid generation and bioremediation with relevance to indian mining

The role of Acidithiobacillus group of bacteria in acid generation and heavy metal dissolution was studied with relevance to some Indian mines. Microorganisms implicated in acid generation such as Acidithiobacillus Acidithicibacillus thiooxidans and Leptospirillum ferrooxidans were isolated from abandoned mines, waste rocks and tailing dumps. Arsenite oxidizing Thiomonas and Bacillus group of bacteria were isolated and their ability to oxidize As (111) to As (V) established. Mine isolated Sulfate reducing bacteria were used to remove dissolved copper, zinc, iron and arsenic from solutions.