Biochemical characterization of a semi-purified aspartic protease from sea catfish Bagre panamensis with milk-clotting activity.

Pepsin from stomach of Bagre panamensis was semi-purified and biochemically characterized. The acid proteolytic activity and purification fold were 3875 U/mg protein and 91.85, respectively, after purification process. The optimum pH and temperature for semi-purified protease were 2-3 and 65 °C, respectively. The enzyme activity was stable after heating proteases at 50 °C for 120 min, but only 30% residual activity was detected after heating at 65 °C for 30 min. SDS-PAGE analysis showed two proteins bands after dialysis (26.1 and 38.6 kDa). Only the band of 38.6 kDa had proteolytic activity, which was inhibited using pepstatin A. Organic solvents, surfactants and reducing agents affect the proteolytic activity at different extent; however, metal ions or EDTA have no impact on protease activity. The semi-purified protease exhibited milk coagulant activity, with a maximum activity at 45 °C. The obtained results highlight the potential biotechnological use of B. panamensis pepsin.

Developmental changes in oxygen consumption and hypoxia tolerance in the heat and hypoxia-adapted tabasco line of the Nile tilapia Oreochromis niloticus, with a survey of the metabolic literature for the genus Oreochromis.

The genus Oreochromis is among the most popular of the tilapiine cichlid tribe for aquaculture. However, their temperature and hypoxia tolerance, if tested at all, is usually tested at temperatures of 20-25°C, rather than at the considerably higher temperatures of 30-35°C typical of tropical aquaculture. We hypothesized that both larvae and adults of the heat and hypoxia-adapted Tabasco-line of the Nile tilapia Oreochromis niloticus would be relatively hypoxia-tolerant. Oxygen consumption rate ( M ˙ O 2 ), Q10 and aquatic surface respiration (ASR) was measured using closed respirometry at 2 (c. 0.2 g), 30 (c. 2-5 g), 105 c. (10-15 g) and 240 (c. 250 g) days of development, at 25°C, 30°C and 35°C. M ˙ O 2 at 30°C was inversely related to body mass: c. 90 µM O2 g-1 /h in larvae down to c. 1 µM O2 g-1 /h in young adults. Q10 for M ˙ O 2 was typical for fish over the range 25-35°C of 1.5-2.0. ASR was exhibited by 50% of the fish at pO2 of 15-50 mmHg in a temperature-dependent fashion. However, the largest adults showed notable ASR only when pO2 fell to below 10 mmHg. Remarkably, pcrit for M ˙ O 2 was 12-17 mmHg at 25-30°C and still only 20-25 mmHg across development at 35°C. These values are among the lowest measured for teleost fishes. Noteworthy is that all fish maintain equilibrium, ventilated their gills and showed routine locomotor action for 10-20 min after M ˙ O 2 ceased at near anoxia and when then returned to oxygenated waters, all fish survived, further indicating a remarkable hypoxic tolerance. Remarkably, data assembled for M ˙ O 2 from >30 studies showed a > x2000 difference, which we attribute to calculation or conversion errors. Nonetheless, pcrit was very low for all Oreochromis sp. and lowest in the heat and hypoxia-adapted Tabasco line.

Fish trypsins: potential applications in biomedicine and prospects for production.

In fishes, trypsins are adapted to different environmental conditions, and the biochemical and kinetic properties of a broad variety of native isoforms have been studied. Proteolytic enzymes remain in high demand in the detergent, food, and feed industries; however, our analysis of the literature showed that, in the last decade, some fish trypsins have been studied for the synthesis of industrial peptides and for specific biomedical uses as antipathogenic agents against viruses and bacteria, which have been recently patented. In addition, innovative strategies of trypsin administration have been studied to ensure that trypsins retain their properties until they exert their action. Biomedical uses require the production of high-quality enzymes. In this context, the production of recombinant trypsins is an alternative. For this purpose, E. coli-based systems have been tested for the production of fish trypsins; however, P. pastoris-based systems also seem to show great potential in the production of fish trypsins with higher production quality. On the other hand, there is a lack of information regarding the specific structures, biochemical and kinetic properties, and characteristics of trypsins produced using heterologous systems. This review describes the potential uses of fish trypsins in biomedicine and the enzymatic and structural properties of native and recombinant fish trypsins obtained to date, outlining some prospects for their study.

Intestinal absorption of amino acids in the Pacific bluefin tuna (Thunnus orientalis): in vitro lysine-arginine interaction using the everted intestine system.

The interaction between lysine (Lys) and arginine (Arg) in the proximal intestinal region of Pacific bluefin tuna (Thunnus orientalis) was evaluated using the everted intestine method. This in vitro intestinal system has been shown to be an effective tool for studying the nutrient absorption without the need to handle the tuna fish in marine cages as needed for digestibility and amino acid (AA) absorption. We used a factorial design with two sets of variables: low and high Lys concentration (10 and 75 mM) and four different Arg concentrations (3, 10, 20, and 30 mM). Both amino acids were dissolved in marine Ringer solution with a basal amino acidic composition consisting of a tryptone solution (9 mg mL(-1)). No interaction was observed between the absorption of Lys and Arg during the first 10 min of the experiment when low concentration of Lys and Arg was used in the hydrolyzate solution. However, there seemed to be a positive effect on Lys absorption when both amino acids were at high concentrations (30 and 75 mM, respectively). This type of studies will led us to test different formulations and/or additives to better understand the efficiency of AA supplementation as an alternative to in situ studies that are difficult to follow to design with the Pacific Bluefin Tuna.