Stomach digestion and its effect upon protein hydrolysis in the intestine of rainbow trout (Salmo gairdneri Richardson)

• 1.1. The in vitro digestion of soya protein by pancreatic proteases of the trout was measured following various conditions of stomach digestion.
• 2.2. Peptic hydrolysis results in an increase of small peptides.
• 3.3. Acid treatment and peptic proteolysis do not affect the degradation of insoluble proteins or the formation of free amino acids during intestinal digestion, but they do lead to a significant shift from soluble polypeptides to di- and oligopeptides.

     

Cotranslational fatty acylation of mucus glycoprotein. Addition of palmitic acid to peptidyl-tRNA occurs prior to peptide chain completion and its release

• 1.1. The fatty acylation of mucus glycoprotein nascent peptides was investigated using [³H]palmitic acid and [³⁵S]methionine-labeled peptidyl-tRNA of rat gastric mucous cells.
• 2.2. The mucus glycoprotein peptidyl-tRNA fraction was found to contain covalently bound palmitic acid in its complexes.
• 3.3. RNase digestion of the mucus glycoprotein peptidyl-tRNA released [³H]palmitic acid labeled peptides which, on SDS-polyacrylamide gel, separated into a multitude of bands ranging in size from 2000 to 60,000 Da.
• 4.4. The analyses of low molecular weight peptides revealed that palmitic acid was present in methionine-labeled peptides containing 30–43 amino acids and those of 18–25 amino acids or larger devoid of methionine, but was not identified in methionine-labeled peptides containing 10–15 amino acids.
• 5.5. The results indicate that the N-terminal fatty acylation of mucus glycoprotein nascent peptides is a cotranslational process which is occuring in an immediate vicinity of the signal peptide fragment.

     

Proteolysis post mortem in North Atlantic krill

• 1.1. The autoproteolytic processes in selected species of North Atlantic krill, Meganyctiphanes norvegica (M. Sars), Thysanoessa inermis (Krøyer) and T. raschii (M. Sars) have been examined at 0°C by following the release of peptides and free amino acids.
• 2.2. The krill contains high levels of peptide hydrolases, and autoproteolysis seems to be due mainly to digestive enzymes localized in the hepatopancreas and the intestinal tract of the animals.
• 3.3. During autoproteolysis the individual amino acids were generally released at rates corresponding to their proportion in the bulk protein of the krill. The major exceptions were alanine which accumulated in amounts larger than was to be expected from the composition of the krill protein, and glutamic acid/glutamine, aspartic acid/asparagine, arginine, and to some extent glycine, proline and serine, which accumulated to a lesser extent than was to be expected.
• 4.4. Storage of krill for 1 week resulted in only minor changes in the total content of amino acids as determined after acid hydrolysis, with the exception of alanine which increased in concentration.
• 5.5. The results suggest that the formation of free alanine is partly due to reactions other than proteolysis.
• 6.6. The release of free amino acids was accompanied by a considerable increase in the amount of small peptides, and glutamic acid/glutamine, aspartic acid/asparagine, glycine and proline tended to accumulate in these peptides.
• 7.7. The autoproteolytic activity of the Thysanoessa species showed seasonal variations, probably in response to food availability. In the case M. norvegica, the results suggest that there are smaller fluctuations in the level of proteolytic enzymes, probably indicating less pronounced variations in the food intake over the year.

     

A comparative study of molluscan and crustacean chitin proteoglycans by carbon-13 NMR spectroscopy. Identification of carbohydrate and amino acid contributions and the determination of amino acid chemical shifts in anhydrous formic acid

• 1.1. ¹³C-NMR spectra of formic acid solutions of chitin proteoglycans from cephalopod pen, lamellibranch siphon sheath and crab cuticle have been determined.
• 2.2. Carbohydrate and amino acid components provide well-defined resonances, completely assignable in the case of hexosamine and partially so for protein amino acids.
• 3.3. The individually unique spectra contain information of compositional and chain environment nature.
• 4.4. Spectral data for each protein amino acid, as a formic acid solution, is presented and compared with values for chemical shifts of amino acids and peptides in water.

     

Specific dynamic action (SDA) in the supralittoral isopod,Ligia pallasii: Identification of components of apparent SDA and effects of dietary amino acid quality and content on SDA

• 1.1. Specific Dynamic Action (SDA) effects of diet were investigated in the supralittoral isopod, Ligia pallasii, using defined chemical diets.
• 2.2. “Apparent SDA”, or the total rise in metabolic rate following a meal, was resolved in animals eating a nutritionally complete chemical diet into three components: 8% mechanical costs of moving food through the gut, 40% “excitement costs” due to investigator disturbance and presence of food, and 52% SDA.
• 3.3. Excitement costs in animals exposed to food but which chose not to eat showed non-significant variation between diets containing different levels of chemical nutrients, but were significantly less on a diet containing only cellulose and agar.
• 4.4. SDA increased with increasing concentration of amino acids in the diet.
• 5.5. Substitution of whole-protein casein for free amino acids in the diet had no significant SDA effect, while substitution of free amino acids in the ratio found in casein more than doubled the SDA effect.
• 6.6. Deletion of alanine from the diet caused no significant effect on SDA, while deletion of phenylalanine caused a highly significant elevation in SDA.

     

The free amino acids of muscle in the cephalochordate Branchiostoma,the cyclostome Myxine and some fishes

• 1.1. Branchiostoma and Myxine have the highest concentrations of amino acids (207 and 234 mM) of the five species investigated.
• 2.2. The predominant amino acids are glycine, proline, alanine, taurine, serine and valine, which form 83–98% of the total, except in Latimeria (60%).
• 3.3. Total amino acids are considered from the point of view of osmotic concentration in relation to other nitrogenous compounds of muscle.

     

Hydrolysis of proteins by peptide hydrolases of Antarctic krill,Euphausia superba

• 1.1. The hydrolysis of casein by peptide hydrolases of Antarctic krill, E. superba, has been
• 2.2. The peptide hydrolases studied included trypsin-like enzymes, carboxypeptidase A-type of enzymes, carboxypeptidase B-type of enzymes, and an aminopeptidase isolated from Antarctic krill.
• 3.3. The trypsin-like enzymes seemed to play a decisive role in the degradation of casein, whereas the carboxypeptidase A, carboxypeptidase B and the aminopeptidase had limited effect when acting on casein alone. When combined with the trypsin-like enzymes, the exopeptidases effected enhanced release of amino acids from the protein.
• 4.4. Based on the pattern of amino acids relased from casein by a crude extract of krill, and by the isolated peptide hydrolases either alone or in combination, it is concluded that the purified peptide hydrolases examined comprise the major enzymes responsible for the autoproteolytic activity of krill at neutral- to weakly alkaline pH.

     

Osmotic changes in an estuarine bivalve,Modiolus fluviatilis

• 1.1.Responses to different salinities monitored by opening and closing of the shell valves were observed in Modiolus fluviatilis.
• 2.2.The osmotic pressure, sodium and chloride ion concentrations were measured in the haemocoelic fluid of Modiolus fluviatilis under similar conditions.
• 3.3.Free amino acids (measured as ninhydrin-positive substances) were determined in the muscle tissue of Modiolus.
• 4.4.It appears that these free amino acids are involved in the ability of the estuarine bivalve Modiolus fluviatilis to osmoregulate in a wide range of salinities.

     

Chemical feeding stimulants for the herbivorous fish,Tilapia zillii

• 1.1. Chemical feeding stimulants for an herbivorous fish, Tilapia zillii have been determined by fractionation and bioassay of substances derived from a model food plant.
• 2.2. Stimulation was produced by amino acids; glutamic acid, aspartic acid, serine, lysine and alanine produced the bulk of stimulatory activity.
• 3.3. These amino acids are among the most abundant in the test plant, and are markedly different from the amino acids found to stimulate feeding in carnivorous fish.
• 4.4. On the basis of these results, a chemically-mediated mechanism of feeding niche separation is postulated.

     

Chemical composition of North Atlantic krill

• 1.1. The main chemical components of Meganyctiphanes norvegica (M. Sars), Thysanoessa inermis (Krøyer) and T. raschii (M. Sars) have been examined.
• 2.2. Protein accounted for 42–47% of the dry weight of M. norvegica and 32–50% of the dry weight of the Thysanoessa species. On a wet weight basis, the protein content was relatively constant and independent of season.
• 3.3. The dominating amino acids in the bulk protein of the krill were glutamic acid/glutamine, aspartic acid/asparagine, glycine, alanine, lysine and leucine.
• 4.4. Lipids were present in amounts of 13–29% of the dry weight in M. norvegica, 15–50% in T. inermis and 12–44% in T. raschii, and the lipid content varied with season.
• 5.5. The main nitrogen extractives in krill, expressed on a dry weight basis, were free amino acids (5–10%), trimethylamine oxide (about 4%), peptides (about 1%) and nucleotides (0.4–1.3%). Trimethylamine and ammonia were present in very low concentrations in living krill.
• 6.6. The amino acids taurine, glycine, proline, arginine, sarcosine and alanine made up 89–93 mol% of the free amino acid pool.
• 7.7. The ash content of krill was in the order of 10–13% of the dry weight, and fluoride represented 1040 and 3200 ppm in the Thysanoessa species and M. norvegioca, respectively.

     

Changes in free amino acid composition in haemolymph of larvae of the wax moth,Galleria mellon ella L., during cold acclimation

• 1.1. Free amino acids were analysed in the haemolymph of Galleria mellonella larvae by HPLC chromatography with o-phthaldialdehyde (OPA)-l-thio-β-d-glucose as derivatization agent.
• 2.2. Fourteen primary amino acids were detected among which glutamine, alanine, γ-aminobutyric acid (GABA) and glycine predominated and constituted 67.7% of the amino acids found.
• 3.3. The concentration of GABA increased significantly with the age of larvae entering the wandering phase and reached a maximum during metamorphosis.
• 4.4. Analysis of cold-acclimated larvae revealed a net increase of free primary amino acids from 96 to 151.8 μmol/ml during consecutive acclimation to 0°C within 20 days and to 205.4μmol/ml during cold shock injury at 0°C (3 hr).
• 5.5. The bulk of this increase was accounted for by alanine, glycine, phenylalanine and lysine.

     

A very stable and potent fibrinolytic enzyme found in earthworm Lumbricus rubellus autolysate

• 1.1. The autolysate of earthworms was found to exhibit powerful fibrin and thrombin substrate hydrolyzing activity.
• 2.2. It also showed a clot-forming activity in the fibrinogen- or plasma-added system.
• 3.3. Zymography revealed that there were three active components with mol. wts of 40,000, 21,000 and 15,000 in the autolysate.
• 4.4. The major form with a mol. wt 35,500 (by SDS-PAGE) was further purified. The N-terminal amino acid sequence of this enzyme (16 residues) was similar to that of the swine pancreatic proelastase.

     

A halogenated amino acid-containing sperm activating peptide and its related peptides isolated from the egg jelly of sea urchins,Tripneustes gratilla,Pseudoboletia maculata,Strongylocentrotus nudus,Echinometra mathaei and Heterocentrotus mammillatus

• 1.1. Twenty-eight peptides were isolated from the egg jelly of sea urchins, Tripneustes gratilla, Pseudoboletia maculata, Strongylocentrotus nudus, Echinometra mathaei (type A and B) and Heterocentrotus mammillatus and their amino acid sequences were determined.
• 2.2. Two of the peptides obtained from T. gratilla egg jelly possessed a bromophenylalanine (Br-Phe) residue in their sequences (Gly-(Br-Phe)-Asn-Leu-Asn-Gly-Gly-Gly-Val-Gly and Gly-(Br-Phe)-Asp-Leu-Asn-Gly-Gly-Gly-Val-Gly).
• 3.3. All of the peptides elevated cyclic GMP concentrations in the spermatozoa of the respective sea urchin and caused a shift in the apparent mol. wt of a major sperm protein of the respective sea urchin.
• 4.4. They stimulated respiration rates of the spermatozoa of Hemicentrotus pulcherrimus as well as their own species.
• 5.5. One-half maximal concentrations of the peptides for respiratory stimulation of H. pulcherrimus spermatozoa were between 10⁻¹¹ M and 10⁻⁹ M except a methionine-containing peptide which was about 10⁻⁷ M.

     

Hyaluronidase degradation of hyaluronic acid from different sources: Influence of the hydrolysis conditions on the production and the relative proportions of tetra- and hexasaccharide produced

• 1.1. Hyaluronic acid (HA) can be digested with a Streptomyces hyaluronidase.
• 2.2. The rate of production and the ratio of tetrasaccharide (T) and hexasaccharide (H), studied by HPLC, varied with the temperature and duration of hydrolysis.
• 3.3. The rates of production and the respective amounts of the two oligosaccharides depended on the rheological properties of the HA from different sources.
• 4.4. A close relationship was found between the initial rate of hydrolysis and the intrinsic viscosity of the HA (η_i).
• 5.5. Our data suggest that enzymatic degradation at a given pH value, temperature, and duration of hydrolysis is dependent on the conformation of HA.
• 6.6. Moreover, under given conditions, the relative proportions of the two oligosaccharides depend on the η_i and may also reflect the degree of hydrolysis of the substrate.

     

Effect of dietary d-amino acids on growth,survival, ammonia excretion and specific dynamic action in the supralittoral isopod,Ligia pallasii

• 1.1. Growth, survival, ammonia excretion and Specific Dynamic Action (SDA) were assessed in the supralittoral isopod Ligia pallasii eating chemical diets with differing proportions of d- and l-amino acids. Growth and survival decreased in direct proportion to increasing dietary intake of d-amino acids.
• 2.2. Survival on diets with greater than 50% content of d-amino acids (based on total amino acids in diet) was lower than that expected based on previous work, suggesting a deleterious effect of the d-isomers.
• 3.3. Ammonia excretion and SDA correlated negatively with increasing dietary content of d-amino acids.
• 4.4. The general conclusion is that d-amino acids play no role in anabolic or energy metabolism in Ligia, and that poor performance at higher dietary levels of d-amino acids may relate to their interference with transport pathways for the normal l-forms.

     

Sulphhydryl protease inhibitors from pineapple plant stem

• 1.1. Pineapple stem extract, consisting of a mixture of the protease bromelain and sulphhydryl protease inhibitors, was fractionated by gel permeation chromatography.
• 2.2. Inhibitor-containing fractions were further resolved by ion exchange chromatography on DEAE-cellulose, giving 12 chromatographically distinct inhibitory fractions.
• 3.3. These 12 inhibitory fractions all show an inhibition specificity towards bromelain.
• 4.4. Reduction, S-carboxymethylation and refractionation of each of these inhibitory fractions gave, for each fraction, two separated peptides of ca 13 and 40 amino acids in length, respectively.
• 5.5. The amino acid compositions and the N-terminal sequences of these peptides show the inhibitors to be a closely homologous set. Both the constituent peptides of each fraction are microheterogeneous. Each DEAE-cellulose chromatogram peak contains a co-eluting set of iso-inhibitors.
• 6.6. Structural microvariations within these isoinhibitors have a minor influence on inhibitor activity towards bromelain.

     

Purification and some properties of isocitrate dehydrogenase of a blowfly Aldrichina grahami

• 1.1. NADH-dependent isocitrate dehydrogenase has been purified 110-fold from the crude extract of the flight muscle mitochondria of Aldrichina grahami.
• 2.2. The purification procedure involved Triton X-100 treatment of isolated mitochondria, column chromatography on DEAE-cellulose, Affi-gel blue, and P-cellulose.
• 3.3. The purified enzyme was homogeneous by criteria of the polyacrylamide gel electrophoresis.
• 4.4. The enzyme of the blowfly contains more acidic amino acids and less hydrophobic amino acids than that of pig heart.
• 5.5. The molecular weight was determined to be 330,000 daltons. The subunit construction differs from ghat of mammalian isocitrate dehydrogenase.

     

Changes in composition and proteolytic enzyme activities of artemia during early development

• 1.1. The proximate composition, total and free amino acids, and proteases of Artemia nauplii were determined during early development.
• 2.2. Moisture increased from 71.0% to 80.8%, crude protein decreased from 13.2% to 8.8%, crude fat and ash varied slightly.
• 3.3. The total amino acids decreased. Free amino acids changed in three patterns.
• 4.4. Trypsin, chymotrypsin, carboxypeptidase A, B and cathepsin B and C increased in activity. The activity of trypsin was lower, while cathepsin B and C were the highest.
• 5.5. The protease activities were maximal at pH 7.5 and 8.0, and at 45°C on casein.
• 6.6. The optimal pH for carboxypeptidase A was 4.0, for carboxypeptidase B was 4.5, for trypsin and chymotrypsin were 7.0–7.5. The protease(s) active at pH 9.0–9.5 were to be determined.

     

Isolation and partial amino acid sequence of A 78 kDa porcine gastrin-binding protein

• 1.1. A 78 kDa protein (p78) has been partially purified from washed membranes isolated from the corpus of porcine gastric mucosa. The purification was monitored by covalent cross-linking of iodinated [Nle¹⁵]-gastrin; 17.
• 2.2. A single N-terminal sequence extending for 33 amino acids was obtained from the p78 preparation. Partial sequences totalling 192 amino acids were also obtained from 14 tryptic and 3 Staphylococcal V8 peptides.
• 3.3. 10 peptides plus the N-terminal sequence were derived from a previously unsequenced protein which was distantly related to the product of the E. coli fadB gene (Baldwin G. S. (1993) Comp. Biochem. Physiol. 104B, 55–61). The remaining 7 peptides were derived from the gb-subunit of the gastric H⁺/K⁺-ATPase.
• 4.4. The gastrin-binding activity remained in association with p78, and could be separated from the P-subunit of the gastric H⁺K⁺-ATPase, during chromatography on tomato lectin-Sepharose.
• 5.5. We conclude that p78 binds gastrin, and is a novel member of the enoyl-CoA hydratase/3-hydroxyacyl-CoA dehydrogenase family of enzymes.

     

Driving forces for the uphill transport of amino acids into epidermal brush border membrane vesicles of the sea anemone,Anemonia sulcata (Cnidaria,anthozoa)

• 1.1. The transport of amino acids into membrane vesicles prepared from epidermal tentacle tissue of the sea anemone, Anemonia sulcata, depends on an electrochemical potential difference caused, e.g. by sodium chloride gradients.
• 2.2. Potassium or choline chloride gradients energized the transport less effectively than sodium chloride gradients. Both Na⁺-ions and Cl⁻-ions were required for the amino acid transport.
• 3.3. The uphill transport of amino acids along the downhill movement of driver ions (sodium chloride gradient conditions) was characterized by an overshoot; under sodium chloride equilibrium conditions, however, an accumulation of amino acids within the vesicles could not be measured.
• 4.4. Potassium diffusion potentials in combination with valinomycin indicated that hyperpolarization (vesicle inside negative) and hypopolarization (vesicle inside positive) enhanced or depressed the accumulation of amino acids within the vesicles.
• 5.5. Being at the phylogenetic base of the Eumetazoa, cnidarians show characteristics for the transmembrane transport of amino acids comparable to those established for vertebrates.