Demonstration of high opioid-like activity in isolated peptides from wheat gluten hydrolysates

Because of a possible relationship between schizophrenia and celiac disease, a condition in some individuals who are sensitive to wheat gluten proteins in the diet, there has been interest in observations that peptides derived from wheat gluten proteins exhibit opioid-like activity in in vitro tests. To determine the origin of the peptides exhibiting opioid activity, wheat proteins were fractionated by size (gel filtration), by charge differences (ion exchange chromatography) and by differences in hydrophobicity (reversed-phase HPLC). These fractions were hydrolyzed by pepsin or pepsin and trypsin and the resulting peptides separated by gel filtration chromatography. The separated peptides were tested for opioid-like activity by competitive binding to opioid receptor sites in rat brain tissue in the presence of tritium-labeled dihydromorphine. The peptides showed considerable differences in activity; while some peptides exhibited no activity, 0.5 mg of the most active peptides were equivalent to 1 nM of morphine in the binding assay. The most active peptides were derived from the gliadin fraction of the gluten complex.     

Myxobacterial slime and proteolytic activity

An extracellular protein-polysaccharide-lipide (PPL) complex from exponentially growing cultures of Myxococcus virescens was purified by phosphate precipitation and gel chromatography. The high molecular weight slime polymer appeared homogenous upon isoelectric focusing. The PPL complex exhibited proteolytic activity against gelatin and the activity was only partly reduced by heat treatment. The function of the slime polymer as protein denatured was studied. The complex formed micelles similar to anionic detergents and it inhibited the precipitation and coagulation of proteins by trichloroacetic acid. Lysozyme was totally inactivated when treated with the PPL complex. By gel chromatography binding studies, the PPl complex was found to bind lysozyme in the ratio of 1 to 5.8 (w/w). After separation of added protein from the complex the anticoagulation effect on the protein remained. The biological function of the PPL complex was demonstrated with hemoglobin. When all susceptible peptide bonds in PPL-treated hemoglobin were hydrolyzed by trypsin only 20% in the urea-denatured protein were attacked. The combined role of slime and proteolytic activity is discussed.Abbreviations Used PPL protein-polysaccharide-lipide - TCA trichloroacetic acid - BSA bovine serum albumin - Tris tris-(hydroxymethyl)aminomethane - CMC critical micelle concentration - DNFB 2,4-dinitrofluorobenzene - DNP N-dinitrophenyl - SDS sodium dodecylsulphate - H.U. Hultin units     

Studies on antiviral glycosides. II. Mode of action for virucidal effects on Sendai virus

Treatment of Sendai virus with $\text{p-(sec-}\text{butyl)-phenyl-6-chloro-6-deoxy-β-}\text{d}\text{-glucopyranoside}$, followed by freezing and thawing resulted in a loss of hemolytic and cell fusion activities as well as infectivity without affecting hemagglutinating and neuraminidase activities. The anti-hemolytic activity of this compound was reversed by the addition of phosphatidyl choline to the virus samples. $\text{p-}\text{Azidophenyl-6-chloro-6-deoxy-β-}\text{d}\text{-[}{}^3\text{H]glucopyranoside}$ was successfully used for photoaffinity labeling of a specific virion site, and we confirmed the affected site of the glucoside to be the lipid components in the viral envelopes.     

Incorporation of3H-thymidine into DNA and the activity of alkaline phosphatase in zinc-deficient fetal rat brains

The incorporation of³H-thymidine into DNA in the brains of the 17-day and 20-day old rat fetuses was significantly reduced by maternal zinc restriction during pregnancy. The activity of the enzyme thymidine kinase (EC 2.7.1.21) was similarly reduced in the zine-deprived fetal brains on days 14 and 20 of gestation, but not on day 17. Fetal brain alkaline phosphatase (EC 3.1.3.1) was significantly depressed by maternal zinc deprivation on days 17 and 20 of pregnancy. The data suggest an association between thymidine kinase and the reduced incorporation of³H-thymidine into DNA in the brains of 20-day old fetuses but not in animals on day 17. Alkaline phosphatase was however depressed at this stage. The suggestion is made that because of the complexity of brain development, future biochemical studies in this area should concern specific structures in the brain at particular critical stages during neurogenesis.     

Antimicrobial defense mechanisms in the horseshoe crab, Limulus polyphemus: preliminary observations with heat-derived extracts of Limulus amoebocyte lysate.

Heat-derived (60°C) extracts of Limulus amoebocyte lysate (LAL) were found to contain potent “broad-spectrum” antimicrobial activity. Additional heating of the LAL extracts to 100°C for 30 min completely inactivated the antimicrobial activity and served as a control. Antimicrobial activity was observed over a temperature range of 0° to 37°C (higher temperatures not tested) with greatest activity at 37°C. Antimicrobial activity of LAL extracts was variable when tested against Gram-negative bacteria of the family Enterobacteriaceae. A twofold concentration of the extracts resulted in a significant decrease in antimicrobial effectiveness. Dialysis of single- and double-strength LAL extracts against deionized water produced a marked and significant enhancement of antimicrobial activity against both resistant and sensitive species, confirming the presence of a dialyzable inhibitor(s). Dialyzed LAL extracts were active against 13 of 14 species of Enterobacteriaceae tested. Two strains of Pseudomonas aeruginosa were susceptible as were two of three Gram-positive cocci tested. Highly sensitive bacterial species were rapidly killed with a greater than 90% reduction in viable counts occurring within the first 30 min of reaction time. Dialyzed LAL extracts also possessed considerable antifungal activity. The role of the Limulus polyphemus amoebocyte in defense against microbial invasion and dissemination is discussed.     

Effects of temperature and sink activity on the transport of 14C-labelled indol-3yl-acetic acid in the intact pea plant (Pisum sativum L.)

The velocity and intensity of basipetal transport of ¹⁴C-labelled indol-3yl-acetic acid (IAA) applied to the apical bud of the intact pea plant were influenced by the temperature to which the stem was exposed and were not influenced by changes in the temperature of the root system when this was controlled independently between 5°C and 35°C. The velocity of transport increased steadily with temperature to a maximum in excess of 35°C and then fell sharply with further increase in temperature. The Q₁₀ for velocity, determined from Arrhenius plots, was low (ca. 1.3). Transport intensity increased to a maximum at about 25°C (Q₁₀=2.2) and then declined gradually with further increase in temperature. It is suggested that transport velocity and transport intensity are controlled independently.The characteristics of auxin transport through the stem were not affected by removal of the root system, or by the withdrawl of root aeration. Labelled IAA did not pass a region of the stem cooled to about 1.0°C, or through a narrow zone of stem tissue killed by heat treatment. In the latter case the heat treatment was shown not to interfere with the upward transport of water in the xylem. Labelled IAA continued to move into, and to accumulate in, the tissues immediately above a cooled or heat-killed region of the stem. It was concluded that the long-distance basipetal transport of auxin through the stem of the intact plant is driven by the transporting cells themselves and is independent of the activity of sinks for the transported auxin.The fronts of the observed tracer profiles in the stem were closely fitted by error function diffusion analogue curves. However, diffusion of IAA alone could not account for the observed characteristics of the transport and it is suggested that the curvilinear fronts of the profiles resulted from a diffusive mixing of exogenous IAA (or IAA-carrier complexes) with endogenous IAA already in the transport pathway.Abbreviations IAA indol-3yl-acetic acid - IAAsp indol-3yl-acetyl aspartic acid - CFM methyl 2-chloro-9-hydroxyfluorene-9-carboxylate (morphactin) - TIBA 2,3,5-triiodobenzoic acid - ABA abscisic acid     

Opiate receptor antagonism by l-prolyl-l-leucyl-glycinamide, MIF-I

Chronic administration of l-prolyl-l-leucyl-glycinamide (MIF-I) to mice slightly reduced morphine's antinociceptive activity in the hot-plate test and modified the biphasic motor activity response to morphine. MIF-I antagonized the initial depression of activity and potentiated the increased motor activity phase. Chronic treatment of rats with MIF-I prevented morphine's antinociceptive activity in the tail flick test. MIF-I partly antagonized the inhibition by morphine of the coaxially stimulated guinea-pig ileum preparation. The inhibition of the ileum produced by ethylketocyclazocine was weakly antagonized by MIF-I. In contrast, MIF-I had no effect on the inhibition of the stimulated mouse vas deferens produced by Leu-enkephalin. The findings show that MIF-I weakly and selectively inhibits μ-type opiate receptors which suggests that MIF-I could be an endogenous inhibitor of opiate receptors.     

Rat and human embryo and post-natal sera contain a potent endogenous competitor of estrogen-rat alpha-fetoprotein interactions

A highly active inhibitor of the binding of estrone and estradiol-17β to rat alpha-fetoprotein is demonstrated for the first time in embryo, immature and adult rat sera as well as in fetal and adult human sera. The competitive character and the narrow specificity of this inhibition effect is shown. The major compound responsible for this activity is isolated by successive column Sephadex LH₂₀ and thin layer chromatography : it is characterized as a nonpolar, nonphenolic, dialysable and thermostable substance, unreactive towards anti-estrone and anti-estradiol-17β anti-bodies. The possible biological role of an endogenous non-estrogen ligand of rodent fetoproteins is discussed.     

Isolation and characterization of fluorescent Pseudomonas associated with the roots of rice and banana grown in Sri Lanka

Bacterial populations in different parts of the rhizosphere of rice and banana in Sri lanka were examined. On rice, the number of aerobic bacteria and the population of fluorescent bacteria were higher in the rhizoplane as compared to the exorhizosphere. However, the opposite was observed with banana. Percentage of fluorescent bacteria was significantly higher on banana (10.8%) than on rice from the wet and dry zones of Sri Lanka (4.3% and 2.7%, respectively). In the endorhizosphere fraction of rice, bacterial populations were very low. Fluorescent bacteria were absent.Based on 33 phenotypical tests, 89 fluorescent isolates were grouped into 5 clusters. The three major clusters covered the isolates belonging to the Pseudomonas fluorescens-putida group, whereas the remaining small clusters contained other UV-fluorescent bacteria. SDS-PAGE of total cell proteins enabled classification of the isolates into one of 12 different protein-polymorphic types. Only a partial correlation was found between the latter classification and the phenotypical one. Cyanogenesis was observed with strains of P. fluorescens only. Isolates P. fluorescens RW9S1 and P. cepacia RW5P1 displayed a potent antagonism against several fungi.