Mitochondrial NAD(P)-malic enzyme from herring skeletal muscle

Mitochondrial NAD(P)-dependent malic enzyme [EC 1.1.1.39, L-malate: NAD⁺ oxidoreductase (decarboxylating)] was purified from herring skeletal muscle to a specific activity of 8.2 mol/min/mg. The purification procedure involved chromatography on DEAE-cellulose, Red Agarose and a Sephacryl S-300 with a final recovery of 38% of enzyme activity. This enzyme catalyzes the oxidative decarboxylation of malate in the presence of either NAD or NADP in the presence of Mn²⁺. Some kinetic characteristics of this enzyme were determined. The pH optimum of activity is 7.0. ATP was shown to be a competitive inhibitor with malate. The inhibition by ATP displayed hyperbolic competitive kinetics with a K_i (ATP) of 0.28 mM in the presence of NAD and 0.75 mM in the presence of NADP. Fumarate reversed ATP inhibition.In vivo, regulation of NAD(P)-dependent malic enzyme might respond to changing levels of mitochondrial ATP and fumarate with the enzyme undergoing kinetic activation by an increase in the concentration of mitochondrial fumarate which could reverse enzyme inhibition by ATP.     

Purification and properties of β-galactosidase from Tilapia intestine: Digestive enzyme of Tilapia-X

ABSTRACT:   β-galactosidase of the intestine of Tilapia nilotica was purified by ammonium sulfate precipitation, followed by PAPTG-Sepharose 4B affinity chromatography, ethylenediamineetetraacetic acid ion-exchange chromatography, polyexchanger PBE 94 chromatofocusing, and Sephadex G-100 gel filtration. β-galactosidase was found to be a single band when examined by poly-acrylamide gel electrophoresis. The purifications of β-galactosidase were 27-fold from the crude extract. β-galactosidase showed optimum activity at pH 5.0 at 40°C, and was specifically found to be able to hydrolyze p -nitrophenyl β-galactopyranoside. It degrades galactan and agarose, and produces galactose. β-galactosidase was strongly inhibited by Hg²⁺ and PCMB. β-galactosidase is considered to be secreted by the upper and middle parts of the intestine and most of the activity was detected in the intestinal juice.     

An investigation of the biological basis of recruitment,growth and adult survival rate variability of Pacific herring (Clupea pallasi) from British Columbia: a synthesis

I explored the biological basis of variation in recruitment (age 3 abundance), growth and age‐specific adult survival rate for the major populations [West Coast Vancouver Island (WCVI), Strait of Georgia, Central Coast, North Coast and Haida Gwaii] of Pacific herring (Clupea pallasi) that inhabit British Columbian waters. The analyses were based on a synthesis of time series of empirical observations of herring population characteristics (egg deposition, age‐specific abundance and size) and prey, competitor and predator biomass/abundance. Recruitment was not correlated among populations. Recruitment variability was explained for WCVI herring only, as a consequence of prey (the euphausiid Thysanoessa spinifera) biomass during August in each of the first 3 years of life, and the biomass of piscivorous Pacific hake (Merluccius productus) during the first year of life. Recruit mass and adult mass‐at‐age were correlated among populations and over ages within populations. Recruit mass was affected by T. spinifera biomass in August of the first and third years of life. Adult mass‐at‐age variability was determined mainly by size at the beginning of the growth season, but also by T. spinifera biomass in August. Age‐specific adult survival rates were not correlated among the five populations. Survival rates decreased with age; there were additional population‐specific effects of somatic mass and T. spinifera biomass in August. The analyses were repeated using physical oceanographic explanatory variables. Only recruit mass variation was explained significantly by physical oceanographic variables, and the biological‐based explanation of recruit mass variability accounted for more of the variation.     

Application of a sensitive,specific and controlled real‐time PCR assay to surveillance indicates a low prevalence of viral haemorrhagic septicaemia virus (VHSV) in wild herring,Clupea harengus L., in Scottish waters

Surveillance data on the distribution of viral haemorrhagic septicaemia virus (VHSV) in the North Sea (UK), targeting Atlantic herring in areas with previous virus detection, were obtained from research cruises conducted during 2005. The sensitive molecular approach of real‐time RT‐PCR (qRT‐PCR) was applied alongside a newly developed endogenous positive control assay specific for herring (elongation factor 1α) to ensure integrity of template. Three hundred and five pools from 1937 individual herring were tested, and no evidence of VHSV in association with wild Atlantic herring was detected. Samples were obtained from Scottish waters where marine aquaculture is conducted. The results confirm that previous tissue culture studies have most likely not significantly underestimated the prevalence of carrier herring in this area. The significance of migratory species such as herring as a reservoir species for VHSV, with the potential to translocate virus genotypes between geographical areas, is discussed.     

Feeding gelatinized carbohydrate in the diets of magur,Clarias batrachus (Linnaeus, 1758): Effects on growth performance,enzyme activities and expression of muscle regulatory factors

This study was conducted to illustrate the effect of dietary gelatinized starch (GS) on the growth performance, enzyme activities and expression of MyoD and Myf5 in magur, Clarias batrachus fingerlings. Four iso‐nitrogenous (37%) and iso‐lipidic (6%) diets containing 15% (D‐1), 25% (D‐2), 35% (D‐3) or 45% (D‐4) GS were fed to 240 fingerlings (2.5 ± 0.5 g) in triplicates groups for 8 weeks. The maximum weight gain %, specific growth rate, protein efficiency ratio, lower FCR and higher mRNA expression of MyoD or Myf5 were found in the D‐3 group fed with 35% GS. Higher hepatosomatic index, viscerosomatic index, body lipid and lower moisture content were found in the D‐4 group. Aspartate transaminase and alanine transaminase activities were found to be higher in the D‐1 group. Amylase, glucose‐6‐phosphate dehydrogenase and blood glucose were higher in the D‐4 group. Superoxide dismutase, catalase and hexokinase activities remain unaffected by the dietary GS levels. Hence, the overall results indicate that 35% GS can improve growth performance and upregulate myogenic regulatory factors, but 45% GS level will favor lipogenesis and compromise growth. Furthermore, Myf5 gene showed more immediate response than the MyoD to the dietary carbohydrate in magur.     

Effect of dietary iron levels on growth,body composition and intestinal enzyme activities of juvenile Jian carp (Cyprinus carpio var. Jian)

A 60‐day feeding trial was carried out to investigate the effect of iron on growth, body composition and digestive enzyme activities. Diets with seven levels of iron (53.9, 90.0, 115.6, 146.1, 176.0, 215.8 and 266.0 mg iron kg^?1 diet) were fed to Jian carp (initial weight 11.4 ± 0.0 g). Per cent weight gain (PWG), feed efficiency (FE) and protein efficiency ratio were the lowest in fish fed the basal diet (P < 0.05). Body protein content was increased with the increasing iron levels (P < 0.05), but moisture, lipid and ash of fish were not significantly affected by dietary iron levels (P > 0.05). Activities of trypsin, lipase, α‐amylase, Na⁺, K⁺‐ATPase, alkaline phosphatase and gamma‐glutamyl transpeptidase were improved with increasing dietary iron levels. Serum iron were significantly enhanced with dietary iron levels up to 146.1 mg iron kg^?1 diet, and plateaued. In conclusion, iron improved digestive enzyme activities of juvenile Jian carp and the dietary iron requirement for serum iron of juvenile Jian carp (11.4–64.0 g) was 147.4 mg iron kg^?1 diet with ferrous fumarate as the iron source.     

Calcium-sensitive extracellular poly(α-L-guluronate)lyase from a marine bacterium Pseudomonas sp. strain F6: Purification and some properties

ABSTRACT: Poly(α- L -guluronate)lyase, as one of alginate lyases, was purified from the culture medium of a marine bacterium, Pseudomonas sp. strain F6, to an electrophoretically homogeneous state. The enzyme was shown to have a molecular mass of 36 kDa by sodium dodecylsulfate–polyacrylamide gel electrophoresis (SDS-PAGE), and was most active at around pH 7.5 and was stable between pH 6.5 and pH 8.5. In the thermal stability experiments, the enzyme's activity diminished through an intermediate state with increasing incubation temperatures and was finally lost when heated at 100°C for 15 min. The addition of hen egg-white lysozyme to the enzyme decreased thermal stability dramatically. The apparent retention of enzyme activity (approximately 50%) was observed after the addition of 6 M guanidine hydrochloride and 8 M urea. Enzyme activity was lost completely with 10 mMSDS, while the ordered structure, which is considered likely to be β-structure, was markedly created. The similar conformational feature has also been created in marine bacterial and mollusc enzymes and the β-structure is commonly observed in polyuronate lyases. The divalent cation (Ca²⁺) promoted the activity of the calcium chelator-treated enzyme significantly, suggesting that Ca²⁺ is involved in the formation of the active intermediate between the acidic uronate(s) and amino acid side-chain(s) of the enzyme.     

Myo-inositol prevents oxidative damage, inhibits oxygen radical generation and increases antioxidant enzyme activities of juvenile Jian carp (Cyprinus carpio var. Jian)

This study was conducted to evaluate the effects of dietary myo -inositol (MI) on the antioxidant status of juvenile Jian carp ( Cyprinus carpio var. Jian). A total of 1050 Jian carp (22.28±0.07 g) were randomly distributed into seven groups of three replicates each, feeding diets containing graded levels of MI (163.5, 232.7, 384.2, 535.8, 687.3, 838.8 and 990.3 mg kg⁻¹ diet) for 60 days. Results indicated that the malondialdehyde content was the lowest for fish fed diets containing ≥384.2 mg MI kg⁻¹, and the highest for fish fed the MI-unsupplemented basal diet ( P <0.05). The protein carbonyl content was decreased with increasing dietary MI levels up to 535.8 mg kg⁻¹ diet, and no differences were found with a further increase in the MI concentration. The anti-superoxide anion capacity (ASA) and anti-hydroxyl radical capacity (AHR) were increased with increasing MI levels up to 535.8 mg kg⁻¹ diet, and plateaued thereafter. The superoxide dismutase and glutathione- S -transferase activities showed the same tendency with the ASA capacity. Catalase, glutathione peroxidase and glutathione reducase activities were improved with increasing MI levels up to 838.8, 384.2 and 687.3 mg kg⁻¹ diet, respectively, and remained nearly constant thereafter. These results suggested that MI could inhibit oxygen radical generation, increase enzymatic antioxidant capacity and prevent oxidative damage of carp. Dietary MI requirements for ASA and AHR activities of juvenile Jian carp were 567.94 and 517.22 mg MI kg⁻¹ diet respectively.