Antiallergic/antiasthmatic effect of novel antiallergic hexapeptide-95/220 in various experimental models

The effects of newly synthesized antiallergic hexapeptide 95/220 was investigated on various allergic and asthmatic test models. This newly developed peptide was found to be more potent than clinically used drug disodium cromoglycate (DSCG). Hexapeptide 95/220 inhibited immediate hypersensitivity reactions such as passive cutaneous anaphylaxis (PCA) and mast cell degranulation in rats, antigen-induced bronchoconstriction in actively sensitized guinea pigs in dose dependent manner like DSCG. Antigen-induced contraction of guinea pig ileum was also markedly inhibited by this newly developed hexapeptide in the same fashion as ketotifen and DSCG did but at comparatively lower dose. Egg albumin-induced histamine release was also blocked by this hexapeptide from chopped lung tissues of sensitized guinea pigs. These results suggest that hexapeptide' 95/220 has potent inhibitory effect on immediate hypersensitivity reactions thereby inhibiting mediator release from mast cell. Moreover, this newly synthesized peptide is orally active and effective at lower doses as compared to standard drugs.     

Hydrogen bonding and equilibrium protium-deuterium fractionation factors in the immunoglobulin G binding domain of protein G

Protium-deuterium fractionation factors (phi) were determined for more than 85% of the backbone amide protons in the IgG binding domains of protein G, GB1 and GB2, from NMR spectra recorded over a range of H2O/D2O solvent ratios. Previous studies suggest a correlation between phi and hydrogen bond strength; amide and hydroxyl groups in strong hydrogen bonds accumulate protium (phi < 1), while weak hydrogen bonds accumulate deuterium (phi > 1). Our results show that the alpha-helical residues have slightly lower phi values (1.03 +/- 0.05) than beta-sheet residues (1.12 +/- 0.07), on average. The lowest phi value obtained (0.65) does not involve a backbone amide but rather is for the interaction between two side chains, Y45 and D47. Fractionation factors for solvent-exposed residues are between the alpha-helix and beta-sheet values, on average, and are close to those for random coil peptides. Further, the difference in phiav between alpha-helix and solvent-exposed residues is small, suggesting that differences in hydrogen bond strength for intrachain hydrogen bonds and amide...water hydrogen bonds are also small. Overall, the enrichment for deuterium suggests that most backbone...backbone hydrogen bonds are weak.     

Novel ligands for the opioid receptors: synthesis and structure-activity relationships among 5'-aryl and 5'-heteroaryl 17-cyclopropylmethyl-4,5 alpha-epoxypyrido[2',3':6,7]morphinans

A series of pyridomorphinans possessing an aryl (10a-s) or heteroaryl (11a-h) substituent at the 5'-position of the pyridine ring of 17-cyclopropylmethyl-4,5 alpha-epoxypyrido[2',3':6,7]morphinan was synthesized and evaluated for binding and functional activity at the opioid delta, mu, and kappa receptors. All of these pyridomorphinans bound with higher affinity at the delta site than at mu or kappa sites. The binding data on isomeric compounds revealed that there exists greater bulk tolerance for substituents placed at the o-position of the phenyl ring than at m- or p-positions. Among the ligands examined, the 2-chlorophenyl (10l), 2-nitrophenyl (10n), 2-pyridyl (11a), and 4-quinolinyl (11g) compounds bound to the delta receptor with subnanomolar affinity. Compound 10c with the p-tolyl substituent displayed the highest mu/delta selectivity (ratio=42) whereas compound 10l with the 2-chlorophenyl substituent displayed the highest kappa/delta selectivity (ratio=23). At 10 microM concentration, the in vitro functional activity determined using [(35)S]GTP-gamma-S binding assays showed that all of the compounds were antagonists devoid of any significant agonist activity at the delta, mu, and kappa receptors. Antagonist potency determinations of three selected ligands revealed that the p-tolyl compound 10c is a potent delta selective antagonist. In the [(35)S]GTP-gamma-S assays this compound had a functional antagonist K(i) value of 0.2, 4.52, and 7.62 nM at the delta, mu, and kappa receptors, respectively. In the smooth muscle assays 10c displayed delta antagonist potency with a K(e) value of 0.88 nM. As an antagonist, it was 70-fold more potent at the delta receptors in the MVD than at the mu receptors in the GPI. The in vitro delta antagonist profile of this pyridomorphinan 10c resembles that of the widely used delta selective antagonist ligand naltrindole.     

Folding of Cu, Zn superoxide dismutase and familial amyotrophic lateral sclerosis

Cu, Zn superoxide dismutase (SOD1) has been implicated in the familial form of the neurodegenerative disease amyotrophic lateral sclerosis (ALS). It has been suggested that mutant mediated SOD1 misfolding/aggregation is an integral part of the pathology of ALS. We study the folding thermodynamics and kinetics of SOD1 using a hybrid molecular dynamics approach. We reproduce the experimentally observed SOD1 folding thermodynamics and find that the residues which contribute the most to SOD1 thermal stability are also crucial for apparent two-state folding kinetics. Surprisingly, we find that these residues are located on the surface of the protein and not in the hydrophobic core. Mutations in some of the identified residues are found in patients with the disease. We argue that the identified residues may play an important role in aggregation. To further characterize the folding of SOD1, we study the role of cysteine residues in folding and find that non-native disulfide bond formation may significantly alter SOD1 folding dynamics and aggregation propensity.     

Optical properties of Erwinia herbicola bacteria at 0.190-2.50 microm

We measure the complex index of refraction of Erwina herbicola (also known as Enterobacter agglomerans or Pantoea agglomerans) bacteria (ATTC 33243) over the spectral region from 0.190 to 2.50 microm (4000-52,632 cm(-1)). Transmission measurements are made on solid films of E. herbicola and on suspensions of the bacteria in water. These measurements, combined with spectral reflectance and Kramers-Kr?nig analysis, allow the determination of the real and imaginary parts over the entire wavelength interval. Accurate and consistent results are obtained for this complex and difficult to measure material. This is part of a continuing series of measurements of the optical constants of representative biological materials that are applicable to the development of methods for detection of airborne biological contaminants, where the material under study is used as a surrogate for a pathogenic agent.     

Proline-rich peptide from the coral pathogen Vibrio shiloi that inhibits photosynthesis of Zooxanthellae

The coral-bleaching bacterium Vibrio shiloi biosynthesizes and secretes an extracellular peptide, referred to as toxin P, which inhibits photosynthesis of coral symbiotic algae (zooxanthellae). Toxin P was produced during the stationary phase when the bacterium was grown on peptone or Casamino Acids media at 29 degrees C. Glycerol inhibited the production of toxin P. Toxin P was purified to homogeneity, yielding the following 12-residue peptide: PYPVYAPPPVVP (molecular weight, 1,295.54). The structure of toxin P was confirmed by chemical synthesis. In the presence of 12.5 mM NH(4)Cl, pure natural or synthetic toxin P (10 microM) caused a 64% decrease in the photosynthetic quantum yield of zooxanthellae within 5 min. The inhibition was proportional to the toxin P concentration. Toxin P bound avidly to zooxanthellae, such that subsequent addition of NH(4)Cl resulted in rapid inhibition of photosynthesis. When zooxanthellae were incubated in the presence of NH(4)Cl and toxin P, there was a rapid decrease in the pH (pH 7.8 to 7.2) of the bulk liquid, suggesting that toxin P facilitates transport of NH(3) into the cell. It is known that uptake of NH(3) into cells can destroy the pH gradient and block photosynthesis. This mode of action of toxin P can help explain the mechanism of coral bleaching by V. shiloi.     

Involvement of a bifunctional,paired-like DNA-binding domain and a transpositional enhancer in Sleeping Beauty transposition

Sleeping Beauty (SB) is the most active Tc1/mariner-like transposon in vertebrate species. Each of the terminal inverted repeats (IRs) of SB contains two transposase-binding sites (DRs). This feature, termed the IR/DR structure, is conserved in a group of Tc1-like transposons. The DNA-binding region of SB transposase, similar to the paired domain of Pax proteins, consists of two helix-turn-helix subdomains (PAI + RED = PAIRED). The N-terminal PAI subdomain was found to play a dominant role in contacting the DRs. Transposase was able to bind to mutant sites retaining the 3' part of the DRs; thus, primary DNA binding is not sufficient to determine the specificity of the transposition reaction. The PAI subdomain was also found to bind to a transpositional enhancer-like sequence within the left IR of SB, and to mediate protein-protein interactions between transposase subunits. A tetrameric form of the transposase was detected in solution, consistent with an interaction between the IR/DR structure and a transposase tetramer. We propose a model in which the transpositional enhancer and the PAI subdomain stabilize complexes formed by a transposase tetramer bound at the IR/DR. These interactions may result in enhanced stability of synaptic complexes, which might explain the efficient transposition of Sleeping Beauty in vertebrate cells.     

Three phase partitioning for extraction of oil from soybean

Three phase partitioning, a method generally used for protein separation, has been evaluated for extraction of oil from soybean. 82% oil was extracted within 1 h using this process which required simultaneous addition of t-butanol (1:1, v/v) and 30% ammonium sulphate to the soybean slurry.