Pazhani Sundaram

Glycosulfatase activity of helicobacter pylori toward gastric mucin

A glycosulfatase activity toward sulfated gastric mucus glycoprotein was identified in the extracellular material elaborated by H. pylori, a bacteria implicated in the etiology of gastric disease. Upon acetone precipitation, an active enzyme fraction at 64% acetone was obtained which on SDS-PAGE gave a major 30kDa protein band. The H. pylori glycosulfatase exhibited maximum activity (314.8 pmol/mg protein/h) at pH 5.7 in the presence of Triton X-100 and CaCl2, and was capable of removal of the sulfate ester groups situated at C-6 of N-acetylglucosamine, galactose and glucose. However, the enzyme was ineffective toward galactosylceramide and lactosylceramide sulfates which contain the sulfate ester group on C-3 of galactose. The results suggest that H. pylori is capable of overcoming the interference by sulfated mucus glycoprotein with its colonization of gastric mucosa.

Novel Detox Gel Depot sequesters β-Amyloid Peptides in a mouse model of Alzheimer's Disease.

Alzheimer’s disease (AD), a debilitating neurodegenerative disease is caused by aggregation and accumulation of a 39–43 amino acid peptide (amyloid β or Aβ) in brain parenchyma and cerebrovasculature. The rational approach would be to use drugs that interfere with Aβ–Aβ interaction and disrupt polymerization. Peptide ligands capable of binding to the KLVFF (amino acids 16–20) region in the Aβ molecule have been investigated as possible drug candidates. Retro-inverso (RI) peptide of this pentapeptide, ffvlk, has been shown to bind artificial fibrils made from Aβ with moderate affinity. We hypothesized that a ‘detox gel’, which is synthesized by covalently linking a tetrameric version of RI peptide ffvlk to poly(ethylene glycol) polymer chains will act like a ‘sink’ to capture Aβ peptides from the surrounding environment. We previously demonstrated that this hypothesis works in an in vitro system. The present study extended this hypothesis to an in vivo mouse model of AD and determined the therapeutic effect of our detox gel. We injected detox gel subcutaneously to AD model mice and analyzed brain levels of Aβ-42 and improvement in memory parameters. The results showed a reduction of brain amyloid burden in detox gel treated mice. Memory parameters in the treated mice improved. No undesirable immune response was observed. The data strongly suggest that our detox gel can be used as an effective therapy to deplete brain Aβ levels. Considering recent abandonment of failed antibody based therapies, our detox gel appears to have the advantage of being a non-immune based therapy.

Detoxification Depot for β -Amyloid Peptides

Alzheimers Disease (AD) is caused by the deposition of insoluble and toxic amyloid peptides (Aβ) in the brain leading to memory loss and other associated neurodegenerative symptoms. To date there is limited treatment options and strategies for treating AD. Studies have shown that clearance of the amyloid plaques from the brain and thus from the blood could be effective in stopping and or delaying the progression of the disease. Small peptides derived from the Aβ- 42 sequence, in particular KLVFF, have shown to be effective binders of Aβ peptides and thus could be useful in delaying progression of the disease. We have taken advantage of this property by generating the retro-inverso (RI) version of this peptide, ffvlk, in different formats. We are presenting a new detox gel system using poly ethylene glycol (PEG), polymerized and cross linked with the RI peptides. We hypothesize that detox gel incorporating RI peptides will act like a ‘sink’ to capture the Aβ peptides from the surrounding environment. We tested these detox gels for their ability to capture biotinylated Aβ-42 peptides in vitro. The results showed that the detox gels bound Aβ-42 peptides effectively and irreversibly. Gels incorporating the tetramer RI peptide exhibited maximum binding capacity. The detox gel could be a potential candidate for treatment strategies to deplete the brain of toxic amyloid peptides.

Protein Stains and Applications

Staining of proteins separated on gels provides the basis for determination of the critical properties of these biopolymers, such as their molecular weight and/or charge. Detection of proteins on gels and blots require stains. These stains vary in sensitivity, ease of use, color, stability, versatility, and specificity. This review discusses different stains and applications with details on how to use the advantages and disadvantages of each stain. It also compiles some important points to be considered in imaging and evaluation. Commonly used colorimetric and fluorescent dyes for general protein staining, and posttranslational modification-specific detection methods are also discussed.

Calcium transport and calcium activated ATPase activity in microsomal vesicles of rat gastric mucosa

1. Microsomal and plasma membrane vesicles, isolated from rat gastric mucosa, were found to exhibit Ca(2+)-dependent ATPase activities of 14.1 +/- 1.4 and 7.8 +/- 1.1 mumol/mg/hr, respectively. The optimum conditions for the microsomal Ca(2+)-ATPase was pH 6-7, and required Mg2+, while divalent cation such as Cu2+, Zn2+, Fe2+, Ba2+ and Cd2+ had no significant effect. 2. As in the case of Ca2+, Mg(2+)-ATPase, the Ca2+ uptake activity of the microsomal membrane required Mg2+. Both processes were stimulated by submicro molar concentrations of Ca2+ and the apparent Km for Ca2+, Mg2+ ATPase and Ca2+ uptake activities were 0.06 microM and 0.02 microM, respectively. 3. Divalent cations Ba2+ and Fe2+, inhibited both microsomal activities, while Zn2+ and Cd2+ showed no effect on them. However, the monovalent cation K+ did not stimulate Ca2+, Mg(2+)-ATPase and Ca2+ uptake activities. 4. The Ca2+ pumping ATPase of rat gastric mucosal microsome cross-reacted with a monoclonal antibody (mAb-5F10) against the human erythrocyte Ca2+ pump. The apparent molecular weight of mucosal Ca2+ pump was 98 kDa. 5. Close relationship between the kinetic parameters of Ca2+, Mg(2+)-ATPase and Ca2+ uptake activities, and the cross reaction of 98 kDa protein of mucosal microsome with erythrocyte Ca2+ pump antibody, strongly suggest the expression of Ca2+ pump in rat gastric mucosa.

Patents on Potential Drugs to Treat Alzheimer’s Disease: Special Emphasis on Small Peptides

The primary objective of this article is to review patents and the related scientific work on naturally occurring compounds, heterocyclic compounds and small peptides that are being explored for their utility for the treatment of Alzheimers disease (AD). In this review, a special emphasis is given to the patents, including our patent issued in 2013, on peptides that bind to A. Utility of the peptides that prevent aggregation or those that help in clearance of Aβ is discussed as the latter can be considered as an important arm in combination therapy for AD.