The Isolation and Characterization of Glycosylated Phosphoproteins from Herring Fish Bones

Past studies of bone extracellular matrix phosphoproteins such as osteopontin and bone sialoprotein have yielded important biological information regarding their role in calcification and the regulation of cellular activity. Most of these studies have been limited to proteins extracted from mammalian and avian vertebrates and nonvertebrates. The present work describes the isolation and purification of two major highly glycosylated and phosphorylated extracellular matrix proteins of 70 and 22 kDa from herring fish bones. The 70-kDa phosphoprotein has some characteristics of osteopontin with respect to amino acid composition and susceptibility to thrombin cleavage. Unlike osteopontin, however, it was found to contain high levels of sialic acid similar to bone sialoprotein. The 22-kDa protein has very different properties such as very high content of phosphoserine (∼270 Ser(P) residues/1000 amino acid residues), Ala, and Asx residues. The N-terminal amino acid sequence analysis of both the 70-kDa (NPIMA(M)ETTS(M)DSKVNPLL) and the 22-kDa (NQDMAMEASSDPEAA) fish phosphoproteins indicate that these unique amino acid sequences are unlike any published in protein databases. An enzyme-linked immunosorbent assay revealed that the 70-kDa phosphoprotein was present principally in bone and in calcified scales, whereas the 22-kDa phosphoprotein was detected only in bone. Immunohistological analysis revealed diffusely positive immunostaining for both the 70- and 22-kDa phosphoproteins throughout the matrix of the bone. Overall, this work adds additional support to the concept that the mechanism of biological calcification has common evolutionary and fundamental bases throughout vertebrate species.     

Long-term enhancement of pulmonary gas exchange after high-altitude residence during maturation.

In a previous study, our laboratory showed that young dogs born at sea level (SL) and raised from 2.5 mo of age to beyond somatic maturity at a high altitude (HA) of 3,100 m show enhanced resting lung function (Johnson RL Jr, Cassidy SS, Grover RF, Schutte JE, and Epstein RH. J Appl Physiol 59: 1773-1782, 1985). To examine whether HA-induced adaptation improves pulmonary gas exchange during exercise and whether adaptation is reversible when animals return to SL before somatic maturity, we raised 2.5-mo-old foxhounds at HA (3,800 m) for 5 mo (to age 7.5 mo) before returning them to SL. Lung function was measured under anesthesia 1 mo and 2 yr after return to SL and during exercise approximately 1 yr after return. In animals exposed to HA relative to simultaneous litter-matched SL controls, resting circulating blood and erythrocyte volumes, lung volumes, septal volume estimated by a rebreathing technique, and lung tissue volume estimated by high-resolution computed tomography scan were persistently higher. Lung diffusing capacity, membrane diffusing capacity, and pulmonary capillary blood volume estimated at a given cardiac output were significantly higher in animals exposed to HA, whereas maximal oxygen uptake and hematocrit were similar between groups. We conclude that relatively short exposure to HA during somatic maturation improves long-term lung function into adulthood.     

Effects of aminoglutethimide on ovarian histology in the rat

Aminoglutethimide, a corticosteroid inhibitor, administered at a daily dose of 150 mg per kg body weight, showed a dramatic interference with ovarian histology in the rat. The drug caused thinning of the germinal epithelium and disruption of the layers of the follicular cells. It also resulted in the appearance of cytoplasmic vacuoles and distortion of follicle cell nuclei. In the corpus luteum, it caused shrinkage of the lutein cells, resulting in large spaces between the cords of cells. It also arrested immature follicles at various stages of their development. Our results indicate that aminoglutethimide, by modifying ovarian structure in the rat, interferes with the ovulation process.     

Differences in the chemical and catalytic characteristics of two crystallographically ''identical'' enzyme catalytic sites. Characterization of actinidin and papain by a combination of pH-dependent substrate catalysis kinetics and reactivity probe studies targeted on the catalytic-site thiol group and its immediate microenvironment.

The characteristics of actinidin (EC 3.4.22.14) and papain (EC 3.4.22.2), two cysteine proteinases whose catalytic-site regions appear to superimpose to a degree that approaches atomic co-ordinate accuracy of both crystal structures, were evaluated by determining (a) the pH-dependence in acid media of the acylation process of the catalytic act (k+2/Ks) using N alpha-benzoyl-L-arginine p-nitroanilide (L-Bz-Arg-Nan) as substrate and (b) the sensitivity of the reactivity of the catalytic-site thiol group and its pH-dependence to structural change in small, thiol-specific, two-protonic-state reactivity probes (2,2'-dipyridyl disulphide and methyl 2-pyridyl disulphide) where enzyme-probe contacts should be restricted to areas close to the catalytic site. Distortion of the catalytic sites of the two enzymes at pH less than 4 was evaluated over time-scales appropriate for both stopped-flow reactivity probe kinetics (less than or equal to 1-2 s) and steady-state substrate catalysis kinetics (3-5 min) by using the 2,2'-dipyridyl disulphide monocation as a titrant for non-distorted catalytic sites. This permitted a lower pH limit to be defined for valid kinetic analysis of both types. The behaviour of the enzymes at pH less than 4 requires a kinetic model in which the apparently biomolecular reaction of enzyme with probe reagent is separated from the process leading to loss of conformational integrity by a potentially reversible step. The acylation of actinidin with L-Bz-Arg-Nan in acidic media occurs in two protonic states, one produced by raising the pH across pKa less than 4 which probably characterizes the formation of -S-/-ImH+ ion pair (pKa approx. 3) and the other, of higher reactivity, produced by raising the pH across pKa 5.5, which may characterize rearrangement of catalytic-site geometry. The pH-dependence of the acylation of papain by L-Bz-Arg-Nan is quite different and is not influenced by protonic dissociation with pKa values in the range 5-6. The earlier conclusion that the acylation of papain depends on two protonic dissociations each with pKa approx. 4 was confirmed. This argument is now more firmly based because titration with 2,2'-dipyridyl disulphide permits the loss of conformational integrity to be taken into account in the analysis of the kinetic data at very low pH. Methyl 2-pyridyl disulphide was synthesized by reaction of pyridine-2-thione with methyl methanethiolsulphonate and its pKa at I = 0.1 was determined by spectral analysis at 307 nm to be 2.8.(ABSTRACT TRUNCATED AT 400 WORDS)     

Growth and development of maxipak wheat as affected by soil salinity and moisture levels

The effect of soil salinity and soil moisture on the growth and yield of maxipak wheat (Triticum aestivum L.) was studied in a lath-house experiment in whih, chloride-sulphate salt mixtures were used to artificially salinize a sandy loam soil from Al-Jadyriah Baghdad. Five soil salinity levels of ECe's equal to 1.7 (Control) 4.2, 5.8, 8.1, 9.4 and 11.0dSm^–1 were prepared and used at 3 levels of available soil moisture depletion, namely, 25, 50, and 75% as determined by weight. Both growth (vegetative) and yield components were studied throughout the growing season.Results showed that increasing the soil salinity from 1.7 to 11.0 dSm^–1, and decreasing the available soil water from 75 to 25% resulted in independent and significant decreases in Mazipak wheat growth and yield components at different stages of plant development. Root growth showed more sensitivity to both available soil water and soil salinity level than other components. It has been concluded that at soil salinity levels of more than 8.0 dSm^–1, available soil water became a limiting factor on wheat growth and the maintenance of 75% of available soil water during the growth period is recommended to obtain satisfactory grain yield.     

Underestimation of D-glucose phosphorylation as measured by 3H2O production from D-[2-3H]glucose

In rat pancreatic islets, tumoral islet cells (RINm5F line), parotid gland, and in human erythrocytes, but not in rat hepatocytes, the production of 3H2O from D-[2-3H]glucose is 20-30% lower than from D-[5-3H]glucose. This coincides with the production of tritiated lactic acid from D-[2-3H]glucose and may be attributable to an intramolecular hydrogen transfer in the phosphoglucoisomerase reaction. It is concluded that the production of 3H2O from D-[2-3H]glucose is not a reliable tool to assess the total rate of hexose phosphorylation.     

Climate–ecosystem modelling made easy: The Land Sites Platform

Dynamic Global Vegetation Models (DGVMs) provide a state-of-the-art process-based approach to study the complex interplay between vegetation and its physical environment. For example, they help to predict how terrestrial plants interact with climate, soils, disturbance and competition for resources. We argue that there is untapped potential for the use of DGVMs in ecological and ecophysiological research. One fundamental barrier to realize this potential is that many researchers with relevant expertize (ecology, plant physiology, soil science, etc.) lack access to the technical resources or awareness of the research potential of DGVMs. Here we present the Land Sites Platform (LSP): new software that facilitates single-site simulations with the Functionally Assembled Terrestrial Ecosystem Simulator, an advanced DGVM coupled with the Community Land Model. The LSP includes a Graphical User Interface and an Application Programming Interface, which improve the user experience and lower the technical thresholds for installing these model architectures and setting up model experiments. The software is distributed via version-controlled containers; researchers and students can run simulations directly on their personal computers or servers, with relatively low hardware requirements, and on different operating systems. Version 1.0 of the LSP supports site-level simulations. We provide input data for 20 established geo-ecological observation sites in Norway and workflows to add generic sites from public global datasets. The LSP makes standard model experiments with default data easily achievable (e.g., for educational or introductory purposes) while retaining flexibility for more advanced scientific uses. We further provide tools to visualize the model input and output, including simple examples to relate predictions to local observations. The LSP improves access to land surface and DGVM modelling as a building block of community cyberinfrastructure that may inspire new avenues for mechanistic ecosystem research across disciplines.     

A brief review on the development of stronglid nematode eggs and larvae under constant and changing temperature conditions-I. Egg development

The present work gives a brief review on the major aspects of the effect of constant and changing temperatures on the development of stronglid nematode eggs. The effect of culture media on the range of temperatures at which the eggs may hatch as well as the rate of their hatching is also included. In addition, a revision of resistance of the eggs to low and high temperatures is also tackled.     

The F38-Like Group,a New Group of Caprine Mycoplasmas?

This paper concerns the taxonomic status of the F38-like group (MacOwan), a prime determinant of contagious caprine pleuropneumonia (CCPP). Extensive biochemical and serological investigations on strain F38 are reported. Some complex serological relationships with other mycoplasma species are revealed. The results, taken in conjunction with earlier published work on geno-typic characters, lead to the conclusion that final classification of these organisms should await further comparative studies of a number of field strains with a related group of strains classified as M. capri-colum. The characterization of F38 confirms its partial relationship to the “M. mycoides group” of ovine/caprine/bovine mycoplasmas, and has also revealed a very close phenotypic relationship to the bovine mycoplasma serogroup 7, a finding of potential diagnostic and epidemiological importance.     

General occurrence of binding to acetylcholinesterase-substrate complex in noncompetitive inhibition and in inhibition by substrate

To assess the relative importance of binding to enzyme-substrate complex (E.S) and to acetylenzyme (EA), noncompetitive inhibition has been studied in hydrolysis by acetylcholinesterase (AcChE) of cationic and uncharged substrates - acetylcholine (AcCh), 3,3-dimethylbutyl acetate, n-butyl acetate, 2-(methylammonio)ethyl acetate, 2- (N,N-diethyl-N-n-butylammonio)ethyl acetate (DEBAAc) and 2-(methylsulfonyl)ethyl acetate. For the N-trimethyl quaternary ions related to AcCh, tetramethylammonium ion, choline and choline ethyl ether, noncompetitive inhibition (Ki(nonc) is more favorable with the slower substrates than with AcCh, i.e., when E.S greater than EA, and is attributed to formation of enzyme-substrate-inhibitor complexes, E.S.I'. Noncompetitive inhibition by tetraethyl-, tert-butyl- and isopropylammonium ions, and acetamidocholine and its lower dimethyl analogue, is also attributed to E.S.I' complexes. Peripheral binding of these inhibitors decreases acylation more than deacylation. Some tertiary dimethylamonio ions have more favorable Ki(nonc) values with AcCh, decreasing deacylation more than acylation. The substrate DEBAAc is a more effective noncompetitive than competitive inhibitor in hydrolysis of AcCh, indicating that it binds more strongly in a peripheral site than in the active site of the free enzyme. In its hydrolysis by AcChE, it acts as its own noncompetitive inhibitor, by this non-productive binding. Formation of E.S.I' complexes is a general characteristic of hydrolysis by AcChE and decrease in rates at high concentrations of AcCh and related substrates is attributed to peripheral regulatory site binding, formation of E.S.S' complexes, rather than to binding to the acetylenzyme.