Properties of hemocyanins isolated from Amazon river arthropods and molluscs

• 1. Hemocyanins from four organisms inhabiting the Amazon River were isolated and partially characterized.
• 2. Three arthropodan species (Dilocarcinus pagei cristatus, Silviocarcinus pardalinus andMacrobrachium amazonicum) possess hemocyanins whose subunit structure is remarkably simple. Regular and SDS polyacrylamide disc electrophoresis revealed predominantly single bands and no polymorphisms.
• 3. Oxygen-binding experiments showed that the three arthropodan hemocyanins possess large positive Bohr effects and pH dependence in the degree of subunit interaction.
• 4. The hemocyanin of one mollusc,Pila sp., was studied and its subunit size appears to be similar to that of other molluscan hemocyanins, i.e. a polypeptide of mol. wt 400,000. In the hemolymph,Pila hemocyanin probably exists as a mixture of 100 and 124S aggregates.
• 5. The oxygen binding properties of the large molecules ofPila hemocyanin are notable because of their low cooperativity and lack of a strong pH-dependence.

     

Properties of enzymes hydrating epoxides in human epidermis and liver

• 1.1. Cytosolic and microsomal epoxide hydrolyzing enzymes of human skin and liver were compared and found to be different.
• 2.2. Epidermal and hepatic cytosolic epoxide hydrolases were different in terms of substrate selectivity, pI, inhibitor sensitivity and affinity Chromatographic properties.
• 3.3. Microsomal epoxide hydrolases had the same pIs but different substrate selectivities.
• 4.4. Cytosolic epoxide hydrolase from adults had higher specific activity than that from neonates or cultured epidermis, but lower activity than adult hepatic enzymes.
• 5.5. The sizes of cytosolic epoxide hydrolase from epidermis and liver were similar and lower than that from cultured fibroblasts.
• 6.6. Cytosolic epoxide hydrolase from all sources shared similar antigenic determinants.

     

Specific biochemical and immunological properties of some water-soluble glycoproteins produced by rat gastric mucosal scrapings in vitro

• 1.1. Two major glycoprotein components were released in vitro by rat gastric mucosal cells after 4 hr incubation with radioisotopes: a high molecular weight fraction with the characteristics of a fucomucin and a low molecular weight fraction, the latter having a higher specific radioactivity than the former.
• 2.2. Pulse chase experiments indicate that several low and high molecular weight glycoproteins are synthesized simultaneously with no precursor-product relationship between them.
• 3.3. Common antigenic determinants, specific to the stomach were found on the 2 fractions, using immunofluorescence, both fractions appeared to be present in the same cells.

     

Kinetics and stoichiometry of cysteinyldopa formation in the first steps of melanogenesis

• 1.1. Spectra of products obtained during dopa oxidation by mushroom tyrosinase in presence of cysteine or glutathione were recorded for the first minutes of the enzymatic reaction.
• 2.2. Two isosbestic points were defined, indicating the existence of a constant ratio between the disappearance of dopa and the formation of cysteinyl- or glutathione-dopa.
• 3.3. Matrix analysis of these spectra verified that there were two kinetically related absorbing species in solution, these being dopa and either cysteinyldopa or glutathione-dopa.
• 4.4. This stoichiometry (1:1) was confirmed by measuring the lag period in dopachrome accumulation, arising from the presence of thiol.
• 5.5. A kinetic approach has been proposed for the first steps, considered common, in the eumelanin and phaeomelanin biosynthesis pathway, thereby allowing us to establish a quantitative relation between the lag period and thiol concentration.
• 6.6. This relation can be used as a simple kinetic method for thiol evaluation.

     

Polymorphism and tissue specificity of scallop tropomyosin

• 1.1. Molecular polymorphism of tropomyosin from various muscle sources of the scallop, Patinopecten yessoensis, was investigated by electrophoretic and immunochemical methods.
• 2.2. Treatment of the muscle sources with trichloroacetic acid (TCA) prior to tropomyosin preparation was found useful to prevent proteolytic degradation of this protein.
• 3.3. Electrophoretic and immunochemical analysis revealed that at least six kinds of tropomyosin isoforms may exist in scallop muscle tissues.
• 4.4. The tropomyosin isoforms showed tissue-specific distribution in amounts and molecular species among the various muscle sources.

     

Human spleen cathepsin D: Its characterization and localization in human spleen

• 1.1. The cathepsin D was purified 1830-fold under mild conditions by a rapid procedure, based on two-step affinity chromatography.
• 2.2. Its molecular weight, amino acid composition and substrate specificity were shown to display minor differences from materials of other origins.
• 3.3. Inhibition with thiol compounds was found to be a specific phenomenon of the cathepsin D from the human spleen.
• 4.4. Production of antiserum specific for purified cathepsin D was demonstrated by immunodiffusion test, an immunoadsorbent column and immunoblotting of the crude enzyme in SDS gel.
• 5.5. In an immunocytochemical study, the antigenic sites for this enzyme were found to be localized in the reticuloendothelial system of the human spleen.
• 6.6. The role of this enzyme in human spleen cell was discussed.

     

Circadian rhythms of determined blood chemistry values in buzzards and eagle owls

• 1.1. Fifteen values were determined in blood samples from six buzzards (Buteo buteo) and six eagle owls (Bubo bubo) over the 24 hr of the day.
• 2.2. Glucose, urea, uric acid, triglyceride and calcium values showed diurnal rhythms in both species. Their respective patterns of diurnal variation were compared.
• 3.3. Phosphorus, cholesterol and cholinesterase levels underwent circadian rhythms only in the buzzards. Albumin/globulin and amylase exhibited diurnal variations exclusively in the eagle owls.
• 4.4. Glutamatic oxaloacetic transaminase, albumin, globulin, total protein and creatinine concentrations did not show diurnal rhythms in either of the species.
• 5.5. Blood values of the different parameters were studied on the basis of the ranges described in birds.

     

Differential weighting of electrophoretic data in crayfish and fiddler crabs (decapoda: astacidae and ocypopidae)

• 1.1. Muscle proteins from the chelae of six crayfish species and ten species of Uca were compared through disc electrophoresis (split gel technique).
• 2.2. No intraspecies variation of the electrophoretic pattern was found.
• 3.3. In interspecies comparisons all components (bands) were weighted individually and specified as ancestral or derived characters.
• 4.4. In the crayfishes the phylogenetic trees constructed from electrophoretic and classical data were found to be congruent. In Uca some branches of either tree remained undefined. Each tree, however, helped complete the other one.

     

Target localization in different luminous environments

• 1.1. A model for target localization, developed for Squilla mantis, was modified for two other stomatopod species.
• 2.2. While Squilla live in dim light, Odontodactylus scyllarus live in medium bright and Gonodactylus in extremely bright habitats.
• 3.3. The critical zones for prey capture, selected by a decision neuron, were identified. These are bifurcated, stretching out for long distances.
• 4.4. A much more limited critical zone is obtained by introducing a strike command neuron, which receives inputs only from the two central ommatidia in the middle band instead of from all six as does the decision neuron.
• 5.5. The results correspond to the different behavior patterns in prey capture for the different species.

     

Structural features of carbohydrate chains in human salivary mucins

• 1.1. The structure of carbohydrate chains in the low and high molecular weight mucus glycoprotein forms from submandibular-sublingual saliva of individuals with blood group B was investigated.
• 2.2. Alkaline borohydride reductive cleavage of the glycoproteins yielded in each case a population of neutral (55%) and acidic (45%) oligosaccharide alditols ranging in size from 3 to 16 sugar units.
• 3.3. The predominant neutral oligosaccharides in both glycoprotein forms consisted of 16 and 15 sugar units arranged in triantennary fashion, and carried blood group B and I antigenic determinants.
• 4.4. Three of the oligosaccharides in each glycoprotein contained sialic acid and ranged in size from 3 to 12 sugar units. In two oligosaccharides sialic acid was linked to C3 of galactose and in one to C6 of N-acetylgalactosamine. The sulfated oligosaccharide in both glycoproteins was identified as a pentasaccharide with the sulfate ester group at C6 of N-acetylglucosamine.
• 5.5. The results demonstrate that contrary to the earlier view the low and high molecular weight mucus glycoprotein forms of human saliva contain identical carbohydrate chains.

     

Cholesterol metabolism in new world primates: Comparative studies in two tamarin species (Saguinus oedipus and Saguinus fuscicollis) and the squirrel monkey (Saimiri sciureus)

• 1.1. Cholesterol metabolism has been characterized in three species of New World primates, the cotton-top tamarin, the saddle-back tamarin, and the squirrel monkey.
• 2.2. When fed a diet containing cholesterol, the three species exhibited differing responses of plasma cholesterol levels.
• 3.3. Dietary cholesterol absorption was determined and plasma cholesterol die-away kinetics were analyzed in terms of a two-pool model.
• 4.4. The results of the analyses of cholesterol turnover are consistent with the observed species-specific differences in plasma cholesterol values and cholesterol absorption.
• 5.5. Cholesterol metabolism differs between the two tamarin species, as well as between the tamarins and the squirrel monkey.
• 6.6. Implications of species-specific differences between tamarin species are discussed in terms of the use of tamarin species as animal models for comparative studies of cholesterol metabolism and the etiology of cancer and cardiovascular disease.

     

Biochemical variations in three species of prairie dogs (Cynomys)

• 1.1. Blood proteins were studied by polyacrylamide disc gel electrophoresis in three species of prairie dogs, Cynomys gunnisoni, C. leucurus, and C. ludovicianus.
• 2.2. The sera were separated into 13–15 fractions and the three species could be distinguished by both qualitative and quantitative differences in their serum patterns.
• 3.3. Qualitatively, variations in the occurrence and number of slow albumin fractions are diagnostic at the species leel.
• 4.4. Quantitative differences were most apparent in variation in the mobility of the major albumin fraction and the transferrin fraction.

     

The qualitative and quantitative analysis of insect hemolymph sugars by high performance thin-layer chromatography

• 1.1. A procedure is described for the analysis of sugars in insect hemolymph by high performance thin-layer chromatography.
• 2.2. Densitometric scanning of spots after plate developments shows linear responses with most sugars in an analytical range of 125ng-2.0 /gmg; detection limits range from 30/2–60 ng.
• 3.3. Quantitative measurements of trehalose, glucose and fructose can be made on hemolymph sample volumes of less than one microliter, allowing blood sugar analysis of individual insects.
• 4.4. Hemolymph sugar determinations on six species of insects agree well with values reported in the literature, but mean values for a species often showed higher variability because samples were taken from individuals.

     

Comparison of U-small nuclear RNA levels in tissues of the silkmoth,Bombyx mori

• 1.1. Tissue-specific abundance of the capped small RNAs in the silkmoth Bombyx mori was compared using preparative immunoprecipitation with anti-trimethylguanosine antibody.
• 2.2. The yields of total capped small RNAs from larval posterior silk gland, 1. early, 2. late in the fifth-instar, and 3. immortal ovarian-derived cells in culture, were determined to be 187, 50 and 218 ng, respectively, per mg of total cellular RNA.
• 3.3. Separation of immunoprecipitated RNAs by polycrylamide gel electrophoresis, followed by densitometric analysis of the bands, allowed the quantitation of individual capped molecules.
• 4.4. This analysis revealed tissue-specific patterns.
• 5.5.|The data indicate that the total abundance of capped small RNAs in Bombyx is highest in rapidly-dividing cells.

     

Distribution of free l-histidine and related dipeptides in the muscle of fresh-water fishes

• 1.1. From the muscle of 20 species of fresh-water fishes, l-histidine, carnosine, anserine, and balenine were analysed by high-performance liquid chromatography.
• 2.2. All cyprinoidei fishes contained significant amount of l-histidine and trace of dipeptides.
• 3.3. High concentration of anserine was found in salmonoidei fishes, irrespective of salmonidae and osmeridae.
• 4.4. Two species of anguilloidei contained large amount of carnosine, small of l-histidine, and determinable of anserine and balenine.
• 5.5. Only trace amounts of these compounds were found in percoidei fishes.
• 6.6. The levels of these compounds represented no large difference among species belonging to sub-order group as well as family.

     

A comparative study of vertebrate eye lens crystallins using isoelectric focusing and densitometry

• 1.1. The crystallin proteins of numerous species belonging to different classes of vertebrates have been studied.
• 2.2. Species-specific crystallin patterns are revealed which unequivocally characterize the different species.
• 3.3. A marked variability in the number and percentage of alpha-, beta- and gamma-crystallins were found in the various species.
• 4.4. The gamma-crystallin family, with a meagre number of common bands, has proved to be most representative of the species. The beta-crystallins, with their greater number of common bands, have been best preserved throughout vertebrate evolution.
• 5.5. From the similarity coefficient matrix a dendrogram is drawn up, a visual phylogenetic summary of the interrelationships between the vertebrates considered.
• 6.6. In the Discussion, other aspects are considered, such as lens morphology, functionality, animal age, post-synthetic modifications and genetic factors.

     

Intact Transition Epitope Mapping – Targeted High-Energy Rupture of Extracted Epitopes (ITEM-THREE),

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Highlights

• •Multiplex epitope mapping/antigenic determinant identification in the gas phase.
• •Intact transition and controlled dissociation of immune complexes by MS.
• •Simultaneous identification and amino acid sequence determination of epitopes.
• •Simplified in-solution sample handling because of ion manipulation and filtering by MS.

     

Characterization of a membrane fragment of respiratory chain complex I from Neurospora crassa. Insights on the topology of the ubiquinone-binding site

• 1.1. A membrane fragment of complex I from the fungus Neurospora crassa was isolated by immunoprecipitation from alkaline-extracted mitochondrial membranes.
• 2.2. Analysis of the polypeptide composition of this hydrophobic domain of complex I has brought insights on the topology of two subunits of the enzyme, namely the 20.8 and 9.3 kDa components.
• 3.3. Our results indicate that the ubiquinone-binding site of complex I resides in the interface of the peripheral and membrane arms of the enzymes. The significance of these findings are discussed.

     

Cloning and characterization of rabbit pancreatic colipase

• 1.1. Among the digestive enzymes synthesized by pancreas, lipase is the principle lipolytic enzyme which hydrolyses dietary glycerides.
• 2.2. For its action it requires a coenzyme, colipase.
• 3.3. The molecular mechanisms of the interaction of these two are not fully understood.
• 4.4. Further, molecular events that regulate and influence lipid absorption are ill denned.
• 5.5. The rabbit is the conventional animal model for the study of lipid absorption. We have undertaken the molecular cloning, and characterization of rabbit pancreatic colipase, the coenzyme for pancreatic lipase.
• 6.6. Colipase has been cloned from a gt 11 library of an adult rabbit pancreatic cDNA by probing with an oligonucleotide derived from human colipase sequence.
• 7.7. The total reading frame consists of 321 nucleotides coding for 90 amino acids of the functional protein and 17 nucleotides of the leader peptide.
• 8.8. Northern blot analysis revealed a distinct band around 0.5kb. Comparison with other species revealed an over all homology of 75% at the nucleotide level.
• 9.9. At the amino acid level highest conservation is observed at the lipase-binding region (AA 53–73).
• 10.10. Rabbit enzyme also retained the N-terminal pentapeptide of it preform.
• 11.11. The regions of homology and conservation may aid to define the sites of interaction of colipase with lipase.

     

Monoclonal antibodies to antigen binding protein of annelids (Lumbricus terrestris)

• 1.1. Monoclonal antibodies (mAb) to antigen binding protein (ABP) of the earthworm Lumbricus terrestris have been prepared.
• 2.2. The specificity of mAb for a determinant located outside the antigen binding site was determined and verified in inhibition experiments.
• 3.3. The mAb were used for isolation of a 56 kDa ABP by an immunoprecipitation technique.
• 4.4. The binding of mAb to coelomocytes has demonstrated the existence of two cell populations, one with low and the other with high densities of ABP molecules on the cell membranes.