Growth hormone binding sites are present in tissues of the porcupine Hystrix hodgesoni,the snakehead Channa maculata and the winter flounder Pleuronectes americanus but not in those of the lamprey Petromyzon marinus

• 1.1. Various tissues of the porcupine Hystrix hodgesoni including liver, intestine, stomach, spleen, kidney, brain and lung were examined for the presence of growth hormone binding sites.
• 2.2. Membranes were prepared from the aforementioned tissues and tested for binding to ¹²⁵I-bovine growth hormone (¹²⁵I-bGH).
• 3.3. Porcupine kidney membranes yielded 1.3 and 2.7% specific binding when tested at 1000 and 2500 μg protein, respectively. Porcupine liver membranes demonstrated approximately 1% specific binding at 3000 μg protein. The other tissues gave low specific binding. The results indicate that porcupine kidney contained binding sites for growth hormone.
• 4.4. Various tissues of two teleosts, the snakehead Channa maculata and the winter founder Pleuronectes americanus, were similarly processed and tested for binding to ¹²⁵I-bGH. It was found that among the different tissues studied, the liver membranes of Channa maculata and the gonad membranes of Pleuronectes americanus gave the highest specific binding of ¹²⁵I-bGH.
• 5.5. Liver and intestine membranes of the lamprey Petromyzon marinus did not bind ¹²⁵I-bGH.

     

Response of rainbow trout to source and level of supplemental dietary methionine

• 1.1. Methionine and total sulfur amino acid (TSAA) requirements of rainbow trout (Salmo gairdneri) were investigated by feeding graded isosulfurous levels of l- and dl-methionine, l-cystine, and the free acid and calcium forms of methionine hydroxy analog (MHA).
• 2.2. Added cystine did not promote growth, survival or prevent cataracts.
• 3.3. l-methionine produced fastest growth, followed by dl-methionine, CaMHA and free acid MHA.
• 4.4. Trout fed CaMHA gained 85.7 and 92.3% as much as those fed l-methionine and dl-methionine.
• 5.5. Within each experiment, the level of L-methionine isomer that prevented cataracts was constant (1.86 g/100g protein in experiment (1), 1.45 in experiment (2) and was lower than for maximum growth (2.89 and 2.15 g) regardless of methionine source.

     

A simplified two-time-point method for measuring whole-body protein synthesis in chicken embryos cultured in vitro: Response to fragmented bovine growth hormone

• 1.1. A simplified two-time-point method for measuring whole-body protein synthesis of chicken embryos cultured in vitro was developed.
• 2.2. The chicken embryos at 7 days of egg incubation age were cultured with a synthetic medium containing l-[4-³H]phenylalanine in a rotatory whole-embryo culture apparatus for a period of up to 60 min.
• 3.3. An adequate combination of measurement time points was examined by comparing fractional synthesis rates calculated by the simplified two-time-point method with those estimated by a full curve-fitting method which would give best estimates. The effect of fragmented bovine growth hormone added to the culture medium on fractional synthesis rates was also tested.
• 4.4. The results indicated that the closest fractional synthesis rates by the simplified two-time-point method to the one by the full curve-fitting method were obtained by taking the time points of t₁ at 10 min and t₂ at 30 or 60 min with intraperitoneal injection of the tracer prior to the culture period.
• 5.5. With the simplified two-time-point method, the fragmentation of bovine growth hormone was shown to increase the biopotency in inducing fractional synthesis rates approximately 100 times as high as that of the intact growth hormone.
• 6.6. It was concluded, therefore, that the present assay method would be convenient and sensitive for searching physiologically active compounds in promoting growth and protein synthesis in the chicken.

     

Essential tyrosyl residues of human placental alkaline phosphatase

• 1.1. Human placental alkaline phosphatase was inactivated with tetranitromethane in a biphasic process.
• 2.2. Spectral and amino acid analysis demonstrated that the inactivation was due to the conversion of tyrosine residues to 3-nitrotyrosine.
• 3.3. The inactivation process showed saturation kinetics.
• 4.4. Protection of the enzyme against tetranitromethane inactivation was afforded by inorganic phosphate.
• 5.5. The binding affinity between the modified enzyme and inorganic phosphate was decreased.
• 6.6. Our results suggest the involvement of tyrosyl residues in the locus of phosphoryl site of the phosphorylated enzyme forms.

     

Analysis of steroid receptor domains with the aid of antihormones

• 1.1. The receptors for steroid hormones consist of well defined domains with overlapping functions.
• 2.2. Contrary to the classical view, it is now becoming increasingly evident that agonist binding regions of the ligand binding domain are not identical to those that bind steroid antagonists.
• 3.3. The DNA binding domain can be activated equally well in presence of both agonists and antagonists, again contradicting the classical view where only the physiologically active hormone was believed to induce such a change.
• 4.4. In some cases, a synthetic antagonist is a more specific ligand for the receptor than the natural hormone.
• 5.5. Synthetic antagonists are therefore important not only to alleviate disease in the human subject, they have also become an important tool to elucidate the mechanism of transactivation by steroid hormones.

     

Vitellogenin synthesis in female larvae of the gypsy moth,Lymantria dispar (L.): suppression by juvenile hormone

• 1.1. Fat body from feeding-phase, last instar gypsy moth females incorporates l-[³⁵S]methionine in vitro into two vitellogenins with the same molecular masses (165 and 180 kDa) as the apo-vitellogenins found in teh hemolymph and the apo-vitellins in teh eggs.
• 2.2. Both apo-vitellogenins are observed in the medium of fat body cultures, but only the 180 kDa apo-vitellogenin is observed in extracts of cultured tissue.
• 3.3. Synthesis and accumulation of the apo-vitellogenins are suppressed in a dose-dependent manner by topical treatment with the juvenile hormone analog, methoprene, prior to day 4.
• 4.4. This suppression suggests that a declining juvenile hormone titre is involved in the initiation of vitellogenin synthesis.

     

Prostaglandin E2/parathyroid hormone-induced suppression of alkaline phosphatase activity is mediated by protein kinase C

• 1.1. Bone resorptive factors, prostaglandin E2 and parathyroid hormone are shown to suppress alkaline phosphatase activity in a rat osteoblastic cell line.
• 2.2. Phorbol myristate acetate, but not dibutyryl cAMP or calcium ionophore can suppress alkaline phosphatase activity.
• 3.3. The protein kinase C inhibitors (H89, staurosporine) are able to block the suppression of alkaline phosphatase activity induced by prostaglandin E2 and parathyroid hormone.
• 4.4. These data suggest that protein kinase C is involved in the inhibition of alkaline phosphatase activity induced by prostaglandin E2 and parathyroid hormone.

     

Inactivation of ornithine decarboxylase by intermediates of tyrosinase-catalyzed reaction

• 1.1. Intermediates in the process of melanin synthesis formed through oxidation of catechols by tyrosinase produced the inactivation of ornithine decarboxylase (ODC), a key enzyme in the polyamine biosynthesis pathway.
• 2.2. The inactivation was dependent on the substrate used (dihydroxybenzylamine ⪢ l-3,4-dihydroxy-phenylalanine ⪢ l-tyrosine) and on the concentration of intermediate produced rather than on the rate of formation.
• 3.3. Sulfhydryl compounds (dithiothreitol and glutathione) or quinone-reducing agents (ascorbic acid) prevented the inactivation of ODC; l-ornithine, but not other aminoacids, also protected partially ODC. The results suggest that different cysteine residues in ODC molecule are implicated in the inactivatory event.
• 4.4. When ¹⁴C-labeled catechols were used, numerous polypeptides resulted labeled, showing that the reactive quinones formed as intermediates in the process of melanin biosynthesis bind covalently to many cellular proteins.

     

Oral administration of insulin in winter-acclimatized carp (Cyprinus carpio) induces hepatic ultrastructural changes

• 1.1. The intestinal absorption of insulin in carps was assessed examining the transepithelial passage of ingested gold-labeled hormone by electron microscopy. Insulin transfer occurred mainly through the intercellular spaces between the enterocytes.
• 2.2. When reaching the lamina propria, the gold-labeled hormone gathered predominantly around the granules of the granular cells, and therefore can enter the circulatory system via the blood capillaries which are found in close contact with these cells.
• 3.3. Winter-acclimatized carp were also capable of internalizing the hormone when fed with insulin.
• 4.4. Furthermore, the absorbed hormone revealed full activity in regard to the observed changes in the ultrastructure of the liver cells of the treated cold-adapted fish.
• 5.5. The fish ingesting the hormone underwent the same type of hepatic ultrastructure reprogramming observed when winter-acclimatized carps are injected intraperitoneally with insulin, i.e. conversion to a phenotype corresponding to hepatocytes from summer-adapted carp.
• 6.6. The oral absorption of insulin by winter-acclimatized fish and its effect in reversing the cold-adaptive state might be useful for the fish culturing industry.

     

Stimulatory effect of amp on the copper ion dependent oxidation of adrenaline to adrenochrome

• 1.1. AMP markedly activated copper ion dependent oxidation of adrenaline to adrenochrome. It is proposed that a Cu(II)-AMP chelate forms a reactive complex with adrenaline.
• 2.2. AMP did not affect copper stimulated oxidation of noradrenaline, dopamine and l-dopa.

     

Solvent perturbation spectra of yellow mealworm α-amylase and of wheat protein inhibitors

• 1.1. The number of tyrosine and tryptophan residue-equivalents on the surfaces of the α-amylase and of two of its protein inhibitors has been determined.
• 2.2. The solvent-exposed tyrosine and tryptophan residue-equivalents in the dimeric inhibitor are respectively four and two times as much as those ones of the monomeric inhibitor.
• 3.3. On the basis of the homology among their polypeptide chains, it is suggested that the outer tryptophan residues in either inhibitors are located at the positions 4 and 51 of the primary structure.
• 4.4. On the interaction of the monomeric inhibitor with the amylase, two tryptophan residue-equivalents became no more accessible to the solvent.

     

Effects of recombinant salmon growth hormone on hypophysectomized male Fundulus heteroclitus

• 1.1. Recombinant salmon growth hormone at doses of 0.8 and 2.1 μg/g significantly enhanced linear growth in hypophysectomized male killifish, Fundulus heteroclitus, over that of controls and a significant regression was found between growth and the logarithm of dose.
• 2.2. Bovine growth hormone elicited significant growth enhancement at all three dosages tested (1,4 and 10 μg/g) and a significant log/dose relationship was also observed.
• 3.3. Observations on the relative weight of the gonads indicate that whole salmon pituitary extract (25 μg/g) possesses strong gonadotropic activity and that both bGH and rsGH had smaller but significant effects on the gonads.
• 4.4. It is suggested that growth hormone may play a subsidiary synergistic role to other pituitary hormones in gonadal development.

     

Selective modification of tryptophan-150 in ovine placental lactogen

• 1.1. Ovine placental lactogen was modified by reaction with o-nitrophenylsulfenyl chloride. Fluorescence measurements indicated that one of the two tryptophan residues of the molecule had reacted. Besides, there was some reagent not covalently bound.
• 2.2. The reagent was covalently bound to Trp-150. No evidence of modification of Trp-90 was found.
• 3.3. Binding capacity to lactogenic as well as somatogenic receptors was diminished but not abolished upon modification, indicating that absolute molecular integrity of Trp-150 is not required for binding.
• 4.4. This behavior is similar to that of the tryptophan residues of ovine prolactin.

     

Sex-specific androgen binding in spotted hyaena (Crocuta crocuta) plasma

• 1.1. A charcoal adsorption assay demonstrated a large variance in androgen binding ability in female spotted hyaenas.
• 2.2. A positive correlation between plasma androgen binding ability and ovarian steroid concentrations was demonstrated in adult females.
• 3.3. The strong plasma binding affinity for testosterone and dihydrotestosterone (DHT) (nM) together with the lack of cortisol and weaker oestradiol-17β binding suggests that a specific androgen binding substance, possibly a protein, is present in adult females of this species.
• 4.4. The lack of high affinity binding in male spotted hyaenas is unusual and deserves further investigation.
• 5.5. Some androgen binding in all, including males and immature animals suggests that albumin may bind some plasma androgens in this species.

     

δ-Aminolevulinic acid dehydratase inactivation by uroporphyrin I in light and darkness

• 1.1. The action of uroporphyrin I on erythrocytic ALA-D activity under dark and light conditions was examined.
• 2.2. Photo and non-photoinactivation of ALA-D induced by uroporphyrin I were observed.
• 3.3. Both effects were dependent on uroporphyrin concentration, temperature and time of exposure of the protein to the porphyrin.
• 4.4. Light-dependent effect of uroporphyrin I is related with the phototoxicity of porphyrins and could be produced by primary amino acid photooxidation followed by secondary cross-linking of the protein.
• 5.5. Light-dependent effect of uroporphyrin I could be ascribed to a direct enzyme inhibition due to binding of the porphyrin to the protein inducing structural changes at or near its active site.

     

Main serum protein of rainbow trout (Salmo gairdnerii): its biological properties and significance

• 1.1. Main serum α₁-protein (α₁P) of rainbow trout was purified and its biochemical and physico-pathological properties were studied.
• 2.2. α₁P was suggested to be a primitive protein having both properties of albumin and AFP in serum proteins of mammals according to the following results.
• 3.3. Molecular weight (75,000), two kinds of molecules (pI 4.55 and 5.05) and amino acid composition.
• 4.4. Dye- or ConA binding activity.
• 5.5. Estrogen binding activity and inhibitory effect on lymphoblastoid-forming activity.
• 6.6. Possible osmotic regulator.
• 7.7. Significant elevation of blood α₁P level in the course of hepatoma induction.

     

Action of free radical generating systems upon the biological and immunological properties of caeruloplasmin

• 1.1. Exposure of human caeruloplasmin, an acute phase protein with antioxidant properties, to a mixture of xanthine/hypoxanthine and xanthine oxidase as a source of reactive oxygen intermediates decreased its ferroxidase and ascorbate oxidase activities and its ability to inhibit lipid peroxidation.
• 2.2. Immunological reactivity was also altered.
• 3.3. Exposure to hydrogen peroxide mimicked these effects.
• 4.4. Exposure to low-intensity u.v. irradiation depressed caeruloplasmin's ability to inhibit iron-catalysed hyaluronic acid degradation.
• 5.5. The results may explain the mechanism of the observed inactivation of caeruloplasmin within human rheumatoid synovial fluid.

     

Interaction of N-methyl-anthraniloyl-labelled porcine apolipoprotein A-I with porcine lecithin: Cholesterol acyltransferase: An energy transfer study

• 1.1. To investigate whether a direct protein-protein interaction between apoA-I and lecithin: cholesterol acyltransferase (LCAT) is necessary for the activation of the enzyme, apoA-I was labelled with N-methylisatoic anhydride at lysine residues. The intermolecular resonance energy transfer from tryptophan residues of LCAT (donor) to N-methyl-anthraniloyl (NMA)-labelled apoA-I (NMA-apoA-I) (acceptor) was used as a sensitive fluorescence method for studying molecular interactions.
• 2.2. In the absence of lipids no fluorescence energy transfer was measurable.
• 3.3. Fluorescence energy transfer occurred from LCAT to NMA-apoA-I in the presence of liposomes with phospholipid/cholesterol ratios ranging from 5:1 to 18:1 and regardless whether only 1 or up to 5 NMA-apoA-I molecules resided at the liposome surface.
• 4.4. This indicates a preferred binding of the enzyme directly to or in spatial proximity to the activator protein NMA-apoA-I even if enough space at the liposome surface is available to allow LCAT binding at a distance, where no energy transfer is measurable.

     

Localization of the glucocorticoid receptor in rat liver cells: Evidence for plasma membrane bound receptor

• 1.1. The cytoplasmic glucocorticoid receptor of rat liver cells is in part recovered in the plasma membrane fraction.
• 2.2. After in vivo administration of [³H]dexamethasone, 0.35% of the radioactivity recovered is bound on plasma membranes.
• 3.3. Dexamethasone also binds in vitro specifically to plasma membranes. Expressed as fmol/mg protein, binding of dexamethasone to plasma membranes is comparable to binding to the soluble cytoplasmic fraction (cytosol).
• 4.4. Using polyclonal antibody to the glucocorticoid receptor and the indirect immunofluorescence technic, an intense decoration of the plasma membranes is observed, denoting a high concentration of glucocorticoid receptor on plasma membranes.
• 5.5. The localization of the receptor on plasma membranes could be of potential importance for its interaction with agents (mitogens, growth factors) initially acting on the cell membrane, regulating subsequent cell proliferation and growth at the level of the cell nucleus.

     

The binding curves in relation to protein concentration. the interaction of human serum albumin with surfactants

• 1.1. The binding curves of three surfactants (natrium dodecyl sulphate, Triton X-100 and Slovafol 909) to human serum albumin at protein concentrations of 0.01–10% were measured.
• 2.2. Contrary to other authors' findings, the results showed the courses of the binding curves to be independent of protein concentration.
• 3.3. The present values of the concentration-dependent binding curves require special accuracy in the experimental techniques.