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Mg Protein (mg + protein)
Kinds of Mg Protein Selected AbstractsGallbladder Na+/H+ exchange activity is up-regulated prior to cholesterol crystal formationEUROPEAN JOURNAL OF CLINICAL INVESTIGATION, Issue 8 2005S. C. Narins Abstract Background, Gallbladder Na+ and H2O absorption are increased prior to gallstone formation and may promote cholesterol nucleation. Na+/H+ exchange (NHE) isoforms NHE2 and NHE3 are involved in gallbladder Na+ transport in prairie dogs. We examined whether increased gallbladder Na+ absorption observed during early gallstone formation is the result of NHE up-regulation. Materials and methods, Native gallbladder and primary cultures of gallbladder epithelial cells (GBECs) harvested from prairie dogs fed nonlithogenic (CON) or 1·2% cholesterol diet for varying lengths of time to induce cholesterol-saturated bile (PreCRYS), cholesterol crystals (CRYS), or gallstones (GS) were used. NHE activity was assessed by measuring dimethylamiloride-inhibitable 22Na+ uptake under H+ gradient in primary GBECs. HOE-694 was used to determine NHE2 and NHE3 contributions. NHE protein and mRNA expression were examined by Western and Northern blots, respectively. Results, Gallbladder total NHE activity was 25·1 ± 1·3 nmol mg protein,1 min,1 in the control and increased during gallstone formation peaking at the PreCRYS stage (98·4 ± 3·9 nmol mg protein,1 min,1). There was a shift in NHE activity from NHE2 to NHE3 as the animals progressed from no stones through the PreCRYS and CRYS stages to gallstones. The increase in NHE activity was partly caused by an increased Vmax without any change in KNam. Both NHE2 and NHE3 protein increased moderately during the PreCRYS stage without increases in mRNA expression. Conclusions, Increased gallbladder Na+ absorption observed prior to crystal formation is in part caused by an increase NHE activity which is not fully accounted for by an increase in NHE proteins and mRNA levels but may be explained by enhanced localization in the membranes and/or altered regulation of NHE. [source] ,- d -Mannopyranosyl-(1,2)-,- d -glucopyranosyl-(1,2)-glycerate in the thermophilic bacterium Petrotoga miotherma , structure, cellular content and functionFEBS JOURNAL, Issue 12 2007Carla D. Jorge The intracellular accumulation of low molecular mass organic compounds in response to stressful conditions was investigated in the thermophilic bacterium Petrotoga miotherma, a member of the order Thermotogales. This led to the discovery of a new solute, whose structure was established as ,- d -mannopyranosyl-(1,2)-,- d -glucopyranosyl-(1,2)-glycerate (MGG) by MMR spectroscopy and MS. Under optimum growth conditions (3% NaCl; 55 °C), MGG was the major solute [up to 0.6 µmol·(mg protein),1]; ,-glutamate and proline were also present but in minor amounts [below 0.08 µmol·(mg protein),1]. The level of MGG increased notably with the salinity of the growth medium up to the optimum NaCl concentration. At higher NaCl concentrations, however, the level of MGG decreased, whereas the levels of proline and ,-glutamate increased about five-fold and 10-fold, respectively. MGG plays a role during low-level osmotic adaptation of Petrotoga miotherma, whereas ,-glutamate and, to a lesser extent, proline are used for osmoprotection under salt stress. MGG is not part of the cell strategy for coping with heat or oxidative stress. Nevertheless, MGG was an efficient protector of pig heart malate dehydrogenase against heat inactivation and freeze-drying, although mannosylglycerate was better. This is the first report on the occurrence of MGG in living systems. [source] Characterization of DNA transport in the thermophilic bacterium Thermus thermophilus HB27FEBS JOURNAL, Issue 18 2006Cornelia Schwarzenlander Horizontal gene transfer has been a major force for genome plasticity over evolutionary history, and is largely responsible for fitness-enhancing traits, including antibiotic resistance and virulence factors. In particular, for adaptation of prokaryotes to extreme environments, lateral gene transfer seems to have played a crucial role. Recently, by performing a genome-wide mutagenesis approach with Thermus thermophilus HB27, we identified the first genes in a thermophilic bacterium for the uptake of free DNA, a process called natural transformation. Here, we present the first data on the biochemistry and bioenergetics of the DNA transport process in this thermophile. We report that linear and circular plasmid DNA are equally well taken up with a high maximal velocity of 1.5 µg DNA·(mg protein),1·min,1, demonstrating an extremely efficient binding and uptake rate of 40 kb·s,1·cell,1. Uncouplers and ATPase inhibitors immediately inhibited DNA uptake, providing clear evidence that DNA translocation in HB27 is an energy-dependent process. DNA uptake studies with genomic DNA of Bacteria, Archaea and Eukarya revealed that Thermus thermophilus HB27 takes up DNA from members of all three domains of life. We propose that the extraordinary broad substrate specificity of the highly efficient Thermus thermophilus HB27 DNA uptake system may contribute significantly to thermoadaptation of Thermus thermophilus HB27 and to interdomain DNA transfer in hot environments. [source] Purification, characterization, cDNA cloning and nucleotide sequencing of a cellulase from the yellow-spotted longicorn beetle, Psacothea hilarisFEBS JOURNAL, Issue 16 2003Masahiro Sugimura A cellulase (endo-,-1,4-glucanase, EC 3.2.1.4) was purified from the gut of larvae of the yellow-spotted longicorn beetle Psacothea hilaris by acetone precipitation and elution from gels after native PAGE and SDS/PAGE with activity staining. The purified protein formed a single band, and the molecular mass was estimated to be 47 kDa. The purified cellulase degraded carboxymethylcellulose (CMC), insoluble cello-oligosaccharide (average degree of polymerization 34) and soluble cello-oligosaccharides longer than cellotriose, but not crystalline cellulose or cellobiose. The specific activity of the cellulase against CMC was 150 µmol·min,1·(mg protein),1. TLC analysis showed that the cellulase produces cellotriose and cellobiose from insoluble cello-oligosaccharides. However, a glucose assay linked with glucose oxidase detected a small amount of glucose, with a productivity of 0.072 µmol·min,1·(mg protein),1. The optimal pH of P. hilaris cellulase was 5.5, close to the pH in the midgut of P. hilaris larvae. The N-terminal amino-acid sequence of the purified P. hilaris cellulase was determined and a degenerate primer designed, which enabled a 975-bp cDNA clone containing a typical polyadenylation signal to be obtained by PCR and sequencing. The deduced amino-acid sequence of P. hilaris cellulase showed high homology to members of glycosyl hydrolase family 5 subfamily 2, and, in addition, a signature sequence for family 5 was found. Thus, this is the first report of a family 5 cellulase from arthropods. [source] The expression of glutathione reductase in the male reproductive system of rats supports the enzymatic basis of glutathione function in spermatogenesisFEBS JOURNAL, Issue 5 2002Tomoko Kaneko Glutathione reductase (GR) recycles oxidized glutathione (GSSG) by converting it to the reduced form (GSH) using an NADPH as the electron source. The function of GR in the male genital tract of the rat was examined by measuring its enzymatic activity and examining the gene expression and localization of the protein. Levels of GR activity, the protein, and the corresponding mRNA were the highest in epididymis among testes, vas deferens, seminal vesicle, and prostate gland. The localization of GR, as evidenced by immunohistochemical techniques, reveals that it exists at high levels in the epithelia of the genital tract. In testis, GR is mainly localized in Sertoli cells. The enzymatic activity and protein expression of GR in primary cultured testicular cells confirmed its predominant expression in Sertoli cells. Intracellular GSH levels, expressed as mol per mg protein, was higher in spermatogenic cells than in Sertoli cells. As a result of these findings, the effects of buthionine sulfoximine (BSO), an inhibitor for GSH synthesis, and 1,3-bis(2-chlorethyl)-1-nitrosourea (BCNU), an inhibitor for GR, on cultured testicular cells were examined. Sertoli cells were prone to die as the result of BCNU, but not BSO treatment, although intracellular levels of GSH declined more severely with BSO treatment. Spermatogenic cells were less sensitive to these agents than Sertoli cells, which indicates that the contribution of these enzymes is less significant in spermatogenic cells. The results herein suggest that the GR system in Sertoli cells is involved in the supplementation of GSH to spermatogenic cells in which high levels of cysteine are required for protamine synthesis. In turn, the genital tract, the epithelia of which are rich in GR, functions in an antioxidative manner to protect sulfhydryl groups and unsaturated fatty acids in spermatozoa from oxidation during the maturation process and storage. [source] Isolation of the dxr gene of Zymomonas mobilis and characterization of the 1-deoxy- D -xylulose 5-phosphate reductoisomeraseFEMS MICROBIOLOGY LETTERS, Issue 1 2000Sigrid Grolle Abstract The gene encoding the second enzyme of the 2C -methyl- D -erythritol 4-phosphate (MEP) pathway for isopentenyl diphosphate biosynthesis, 1-deoxy- D -xylulose 5-phosphate (DXP) reductoisomerase, was cloned and sequenced from Zymomonas mobilis. The deduced amino acid sequence showed the highest identity (48.2%) to the DXP reductoisomerase of Escherichia coli. Biochemical characterization of the purified DXP reductoisomerase showed a strict dependence of the enzyme on NADPH and divalent cations (Mn2+, Co2+ or Mg2+). The enzyme is a dimer with a molecular mass of 39 kDa per subunit and has a specific activity of 19.5 U mg protein,1. Catalysis of the intramolecular rearrangement and reduction of DXP to MEP is competitively inhibited by the antibiotic fosmidomycin with a Ki of 0.6 ,M. [source] Generation of the improved recombinant xylose-utilizing Saccharomyces cerevisiae TMB 3400 by random mutagenesis and physiological comparison with Pichia stipitis CBS 6054FEMS YEAST RESEARCH, Issue 3 2003C.Fredrik Wahlbom Abstract The recombinant xylose-utilizing Saccharomyces cerevisiae TMB 3399 was constructed by chromosomal integration of the genes encoding d -xylose reductase (XR), xylitol dehydrogenase (XDH), and xylulokinase (XK). S. cerevisiae TMB 3399 was subjected to chemical mutagenesis with ethyl methanesulfonate and, after enrichment, 33 mutants were selected for improved growth on d -xylose and carbon dioxide formation in Durham tubes. The best-performing mutant was called S. cerevisiae TMB 3400. The novel, recombinant S. cerevisiae strains were compared with Pichia stipitis CBS 6054 through cultivation under aerobic, oxygen-limited, and anaerobic conditions in a defined mineral medium using only d -xylose as carbon and energy source. The mutation led to a more than five-fold increase in maximum specific growth rate, from 0.0255 h,1 for S. cerevisiae TMB 3399 to 0.14 h,1 for S. cerevisiae TMB 3400, whereas P. stipitis grew at a maximum specific growth rate of 0.44 h,1. All yeast strains formed ethanol only under oxygen-limited and anaerobic conditions. The ethanol yields and maximum specific ethanol productivities during oxygen limitation were 0.21, 0.25, and 0.30 g ethanol g xylose,1 and 0.001, 0.10, and 0.16 g ethanol g biomass,1 h,1 for S. cerevisiae TMB 3399, TMB 3400, and P. stipitis CBS 6054, respectively. The xylitol yield under oxygen-limited and anaerobic conditions was two-fold higher for S. cerevisiae TMB 3399 than for TMB 3400, but the glycerol yield was higher for TMB 3400. The specific activity, in U mg protein,1, was higher for XDH than for XR in both S. cerevisiae TMB 3399 and TMB 3400, while P. stipitis CBS 6054 showed the opposite relation. S. cerevisiae TMB 3400 displayed higher specific XR, XDH and XK activities than TMB 3399. Hence, we have demonstrated that a combination of metabolic engineering and random mutagenesis was successful to generate a superior, xylose-utilizing S. cerevisiae, and uncovered distinctive physiological properties of the mutant. [source] Characterization of Arginine Transport in Helicobacter pyloriHELICOBACTER, Issue 4 2003George L. Mendz ABSTRACT Background. The amino acid L-arginine is an essential requirement for growth of Helicobacter pylori. Several physiological roles of this amino acid have been identified in the bacterium, but very little is known about the transport of L-arginine and of other amino acids into H. pylori. Methods. Radioactive tracer techniques using L-(U- 14C) arginine and the centrifugation through oil method were employed to measure the kinetic parameters, temperature dependence, substrate specificity, and effects of analogues and inhibitors on L-arginine transport. Results. The transport of arginine at millimolar concentrations was saturable with a Km of 2.4 ± 0.3 mM and Vmax of 1.3 ± 0.2 pmole min,1 (µl cell water),1 or 31 ± 3 nmole per minute (mg protein),1 at 20°C, depended on temperature between 4 and 40°C, and was susceptible to inhibitors. These characteristics suggested the presence of one or more arginine carriers. The substrate specificity of the transport system was studied by measuring the effects of L-arginine analogues and amino acids on the rates of transport of L-arginine. The absence of inhibition in competition experiments with L-lysine and L-ornithine indicated that the transport system was not of the Lysine-Arginine-Ornithine or Arginine-Ornithine types. The presence of different monovalent cations did not affect the transport rates. Several properties of L-arginine transport were elucidated by investigating the effects of potential inhibitors. Conclusions. The results provided evidence that the transport of L-arginine into H. pylori cells was carrier-mediated transport with the driving force supplied by the chemical gradient of the amino acid. [source] Stable maintenance of 5, -reductase activity in long-term subcultures of fibroblasts derived from the foreskinINTERNATIONAL JOURNAL OF UROLOGY, Issue 6 2002Kazumi Nakae Abstract Background: There is up to a 50-fold variation in control subjects in current assays of 5,-reductase activity which makes interpretation difficult. It was therefore attempted in this study to establish an assay method which produced stable 5,-reductase activity in long-term subcultured foreskin fibroblasts. Methods: Foreskin fibroblasts were obtained from three boys with phimosis (control subjects), three patients with Reifenstein syndrome and one patient with 5,-reductase deficiency (due to mutation L113P in exon 2 of the SRD5A2 gene). To maximize the number of cells in the DNA synthesis phase, cells were subcultured consistently to approximately 70% confluency. Thawed cells, frozen after the third subculture, were incubated for 24 h with [1,,2,- 3H] testosterone. 5,-Reductase activity was expressed as the sum of formed [3H] 5,-reduced metabolites (separated by thin-layer chromatography). Results: The full range of 5,-reductase activity in controls and patients with Reifenstein syndrome was 3.44,15.59 pmol/h per mg protein: a 4.53-fold variation. The activity in the patient with 5,- reductase deficiency was 0.52 pmol/h per mg protein. Conclusion: By the cell culture methods used in this study, which aimed to increase the number of cells in the DNA synthesis phase, foreskin fibroblasts maintained a considerably stable level of 5,-reductase activity during long-term subculture. Therefore, this assay method can be used for differential diagnosis of 5,-reductase deficiency from other relevant entities. [source] Sucrose phosphorylase of the rumen bacterium Pseudobutyrivibrio ruminis strain AJOURNAL OF APPLIED MICROBIOLOGY, Issue 3 2009A. Kasperowicz Abstract Aims:, To verify the taxonomic affiliation of bacterium Butyrivibrio fibrisolvens strain A from our collection and to characterize its enzyme(s) responsible for digestion of sucrose. Methods and Results:, Comparison of the 16S rRNA gene of the bacterium with GenBank showed over 99% sequence identity to the species Pseudobutyrivibrio ruminis. Molecular filtration, native electrophoresis on polyacrylamide gel, zymography and thin layer chromatography were used to identify and characterize the relevant enzyme. An intracellular sucrose phosphorylase with an approximate molecular mass of 52 kDa exhibiting maximum activity at pH 6·0 and temperature 45°C was identified. The enzyme was of inducible character and catalysed the reversible conversion of sucrose to fructose and glucose-1-P. The reaction required inorganic phosphate. The Km for glucose-1-P formation and fructose release were 3·88 × 10,3 and 5·56 × 10,3 mol l,1 sucrose, respectively , while the Vmax of the reactions were ,0·579 and 0·9 ,mol mg protein,1 min,1. The enzyme also released free glucose from glucose phosphate. Conclusion:,Pseudobutyrivibrio ruminis strain A utilized sucrose by phosphorolytic cleavage. Significance and Impact of the Study:, Bacterium P. ruminis strain A probably participates in the transfer of energy from dietetary sucrose to the host animal. [source] Covalent immobilization of ,-galactosidase on carrageenan coated with chitosanJOURNAL OF APPLIED POLYMER SCIENCE, Issue 1 2009Magdy M.M. Elnashar Abstract ,-Galactosidase was covalently immobilized to carrageenan coated with chitosan for the hydrolysis of lactose. The chitosan-carrageenan polyelectrolyte interaction was found to be dependent on the chitosan pH. At pH 4, the chitosan reached its maximum binding of 28.5% (w/w) where the chitosan surface density was 4.8 mg chitosan/cm2 g of carrageenan gel disks, using Muzzarelli method. Glutaraldehyde was used as a mediator to incorporate new functionality, aldehydic carbonyl group, to the bio-polymers for covalent attachment of ,-galactosidase. The enzyme was covalently immobilized to the biopolymer at a concentration of 2.73 mg protein per g of wet gel. FTIR proved the incorporation of the aldehydic carbonyl group to the carrageenan coated with chitosan at 1720 cm,1. The optimum time for enzyme immobilization was found to be 16 h, after which a plateau was reached. The enzyme loading increased from 2.65 U/g (control gel) to 10.92 U/g gel using the covalent technique. The gel's modification has shown to improve the carrageenan gel thermal stability as well as the immobilized enzyme. For example, the carrageenan gel treated with chitosan showed an outstanding thermal stability at 95°C compared with 35°C for the untreated carrageenan gel. Similarly, the immobilization process shifted the enzyme's optimum temperature from 50°C for the free enzyme towards a wider temperature range 45,55 °C indicating that the enzyme structure is strengthened by immobilization. In brief, the newly developed immobilization method is simple; the carrier is cheap, yet effective and can be used for the immobilization of other enzymes. © 2009 Wiley Periodicals, Inc. J Appl Polym Sci, 2009 [source] Uptake of lamivudine by rat renal brush border membrane vesiclesJOURNAL OF PHARMACY AND PHARMACOLOGY: AN INTERNATI ONAL JOURNAL OF PHARMACEUTICAL SCIENCE, Issue 1 2002Takatoshi Takubo Uptake of lamivudine, a nucleoside analogue antiviral agent, by brush border membrane vesicles (BBMV) prepared from rat renal cortex was investigated. Initial uptake of lamivudine by BBMV was stimulated in the presence of an outward pH gradient. Determination of the kinetic parameters of the initial uptake yielded apparent Km and Vmax values of 2.28 mM and 1.56 nmol (mg protein),1 (20 s),1, respectively. The pH-driven uptake of lamivudine was inhibited by organic cations such as trimethoprim and cimetidine. The inhibitory effect of trimethoprim on lamivudine uptake was competitive, with an apparent Ki of 27.6 ,M. The uptake of lamivudine was also inhibited by nitrobenzylthioinosine, a representative inhibitor of nucleoside transport, and by other nucleoside analogues, such as azidothymidine and dideoxycytidine, that are excreted by renal tubular secretion. These findings suggest that efflux of lamivudine at the brush border membrane of renal tubular epithelium is mediated by an H+/lamivudine antiport system, which may correspond to the H+/organic cation antiport system, and that this system is also involved in the renal secretion of other nucleoside analogues. [source] The role of cellular polysaccharides in the formation and stability of aerobic granulesLETTERS IN APPLIED MICROBIOLOGY, Issue 3 2001J.-H. Tay Aims:,This paper attempts to investigate the role of cellular polysaccharides in the formation and stability of aerobic granules. Methods and Results:,Three column sequential aerobic sludge blanket reactors (R1, R2 and R3) were operated at a superficial air upflow velocity of 0·3 cm s,1, 1·2 cm s,1 and 2·4 cm s,1, respectively. Aerobic granules appeared at cycle 42 in R2 and R3 with a mean size of 0·37 mm in R2 and 0·35 mm in R3, however, aerobic granulation was not observed in R1. After the formation of aerobic granules, the sludge volume index (SVI) decreased to 55 ml g,1 in R2 and 46 ml g,1 in R3. Aerobic granulation was concurrent with a sharp increase of cellular polysaccharides normalized to cellular proteins, which increased from 5·7 to 13·0 mg per mg proteins in R2, and 7·5,13·9 mg per mg protein in R3. The content of polysaccharides in aerobic granules was 2,3 times higher than that in the bioflocci cultivated in R1. The disappearance of aerobic granules in R2 was tightly coupled to a drop in cellular polysaccharides. After the reappearance of bioflocci in R2, the content of cellular polysaccharides were found to be restored to the level observed in R1. Conclusions:,It appears that the production of cellular polysaccharides could be stimulated by hydrodynamic shear force and contributes to the formation and stability of aerobic granules. Significance and Impact of the Study:,It is expected that this study would provide useful information for better understanding the mechanisms of aerobic granulation. [source] Successful oral tolerance induction in severe peanut allergyALLERGY, Issue 8 2009A. T. Clark Background:, Peanut allergy is common, potentially severe and rarely resolves causing impaired quality of life. No disease-modifying treatment exists and there is therefore a need to develop a therapeutic intervention. Aims of the study:, The aim of this study was to investigate whether peanut oral immunotherapy (OIT) can induce clinical tolerance to peanut protein. Methods:, Four peanut-allergic children underwent OIT. Preintervention oral challenges were performed to confirm clinical allergy and define the amount of protein required to cause a reaction (dose thresholds). OIT was then administered as daily doses of peanut flour increasing from 5 to 800 mg of protein with 2-weekly dose increases. After 6 further weeks of treatment, the oral challenge was repeated to define change in dose threshold and subjects continued daily treatment. Results:, Preintervention challenges confirmed peanut allergy and revealed dose thresholds of 5,50 mg (1/40,1/4 of a whole peanut); one subject had anaphylaxis during challenge and required adrenaline injection. All subjects tolerated immunotherapy updosing to 800 mg protein and i.m. adrenaline was not required. Each subject tolerated at least 10 whole peanuts (approximately 2.38 g protein) in postintervention challenges, an increase in dose threshold of at least 48-, 49-, 55- and 478-fold for the four subjects. Conclusions:, We demonstrated a substantial increase in dose threshold after OIT in all subjects, including the subject with proven anaphylaxis. OIT was well tolerated and conferred protection against at least 10 peanuts, more than is likely to be encountered during accidental ingestion. [source] Differential induction of superoxide dismutase in downy mildew-resistant and -susceptible genotypes of pearl milletPLANT PATHOLOGY, Issue 4 2002M. P. Babitha Differential induction of superoxide dismutase (SOD) in downy mildew-resistant and -susceptible genotypes of pearl millet (Pennisetum glaucum) was observed on inoculation with Sclerospora graminicola. SOD activity was studied in resistant (IP18292) and susceptible (23B) pearl millet seedlings inoculated with S. graminicola. SOD activity increased by 2·3-fold in resistant seedlings upon inoculation. SOD activity was greatest in roots, with a specific activity of 3182 U per mg protein, after inoculation. SOD activity increased in all the resistant genotypes upon inoculation with S. graminicola. Native PAGE analysis showed four isozymes of SOD, three of which (SOD-1, -2 and -4) were Cu/Zn-SOD, whereas isozyme SOD-3 was Mn-SOD. This study also revealed increased intensity of all four isozymes of SOD in the resistant genotype upon inoculation. The involvement of SOD in pearl millet (host),downy mildew pathogen interaction is discussed. [source] Energy and protein demands for optimal egg production including maintenance requirements of female tilapia Oreochromis niloticusAQUACULTURE RESEARCH, Issue 5 2010Ingrid Lupatsch Abstract The daily requirements of a spawning tilapia female are quantified from the sum of the requirements for maintenance plus production of eggs. The protein and energy requirements for maintenance and the cost of depositing energy and protein towards growth or gonadal products were determined by supplying feed at increasing levels from zero to the maximum intake. Comparative body composition analyses of the females in addition to the amount and the content of eggs enabled us to quantify the total energy and protein channelled into weight gain or alternatively into egg production. The amount of eggs produced increased with increasing feeding levels and ranged between 0.7 and 1.1 g eggs per kg,1 fish day,1. Regardless of feed allowance, the composition of eggs was similar and contained 235 mg protein and 10.5 kJ g,1 wet weight. In contrast, the whole body of tilapia contained 167 mg protein and 6.7 kJ g,1 on average. The energy requirement for maintenance was calculated to be 59.46 kJ × BW (kg)0.80 and 0.98 g × BW (kg)0.70 for digestible protein. The partial efficiency of producing gonads was 0.67 and 0.59 for digestible energy and digestible protein respectively. [source] Digestive enzyme activity at different developmental stages of blackspot seabream, Pagellus bogaraveo (Brunnich 1768)AQUACULTURE RESEARCH, Issue 4 2008Laura Ribeiro Abstract Blackspot seabream, Pagellus bogaraveo (Brunnich), has been identified as a potential species to diversify European aquaculture production. Although rearing aspects have been widely investigated, little information exists on the nutritional requirements for this species. The aim of this study was to build up information on the activity of digestive enzymes at certain developmental stages of blackspot seabream in order to understand the nutritional needs of larvae and post larvae. Fish larvae were reared from hatching to 55 days after hatching (dah), and the feeding plan consisted in rotifers (5,35 dah), Artemia naupli (30,35 dah) metanaupli (35,45) and Gemma microdiet (45,55 dah). At 7, 11, 21, 45 and 55 days after hatching (dah), pooled samples of fish larvae were collected for analysis of trypsin, amylase, lipase, alkaline phosphatase and leucine,alanine peptidase activity. Up to 21 dah, the whole larvae body was used for enzymatic analysis, whereas in older larvae only the dissected abdominal cavity was used. Blackspot seabream body dry weight growth was exponential, increasing from 60 ,g at 5 dah to 30±9.7 mg at 55 dah. Amylase specific activity decreased significantly during development, exhibiting at 11 dah (0.6 U mg,1 protein) an average value 2.7 times lower than at 7 dah, and remaining stable between 45 and 55 dah (0.7 U mg protein,1). Trypsin specific activity remained constant until 21 dah (between 38 and 44 mU mg protein,1), which could be related to the larvae feeding regime. At later stages of development, lipase-specific activity exhibited a significant increase (P<0.05), being three times higher at 55 dah (8 U mg protein,1) than at 45 dah. The total activity of the studied digestive enzymes increased significantly during larval development (until 21 dah), whereas afterwards only lipase and leucine,alanine peptidase increased significantly between 45 and 55 dah. The pattern of digestive enzymes activity was related to organogenesis and the type of food used at different developmental stages. [source] Evaluation of SupermixTM as an in vitro model of human liver microsomal drug metabolismBIOPHARMACEUTICS AND DRUG DISPOSITION, Issue 5 2002Karthik Venkatakrishnan Abstract SUPERMIXTM is a commercially available formulation of insect cell-expressed human drug-metabolizing cytochrome P450 (CYP) isoforms, mixed in proportions that are optimized to parallel their relative activities in human liver microsomes. We have evaluated the apparent functional affinity and capacity of individual CYP isoforms in SUPERMIXTM in comparison with microsomes from a panel of 12 human livers, using enzyme kinetic studies of isoform-selective index reactions. In addition, we have measured the concentration of NADPH cytochrome P450 oxidoreductase (OR) in SUPERMIXTM and compared it with the concentrations of this accessory electron transfer protein in human liver microsomes. No important differences were evident in the catalytic activities of CYPs 1A2, 2C8, 2C9, 2C19, 2D6 and 3A4 between SUPERMIXTM and human liver microsomes. However, SUPERMIXTM lacks CYP2B6 activity and did not hydroxylate the antidepressant bupropion, a clinically relevant substrate of this enzyme. In addition, the concentration of OR in SUPERMIXTM (1198 pmol mg protein,1) is 17-fold higher than the mean value in human liver microsomes (70 pmol mg protein,1). In conclusion, SUPERMIXTM lacks CYP2B6 activity and contains supraphysiological concentrations of the accessory electron transfer protein OR. These factors should be considered when this formulation is used as an in vitro model in human liver microsomal drug metabolism studies. Copyright © 2002 John Wiley & Sons, Ltd. [source] A critical evaluation of the brain efflux index method as applied to the nitric oxide synthase inhibitor, aminoguanidineBIOPHARMACEUTICS AND DRUG DISPOSITION, Issue 9 2001Joseph J. Raybon Abstract The Brain Efflux Index (BEI) method is an in vivo procedure designed to quantitate saturable efflux mechanisms resident at the blood,brain barrier (BBB). The present work utilized the BEI method to assess the BBB efflux mechanisms of [14C]aminoguanidine, a nitric oxide synthase inhibitor. The BEI for [14C]aminoguanidine was >100% (relative to [3H]inulin diffusion) over a range of 41,184 pmol after 40 min. The unusually high retention (>100%) of [14C]aminoguanidine suggested brain parenchymal sequestration, either by neuronal uptake or tissue protein binding. The uptake of [14C]aminoguanidine in dendritic neuronal endings (synaptosomes) showed a saturable concentration dependency, consistent with a carrier-mediated process. Nonlinear least-squares regression yielded the following Michaelis,Menten and diffusional (kns) parameters for synaptosomal [14C]aminoguanidine uptake: Vmax=118.50± 28.77 pmol x mg protein,1/3 min; Km=58.34±8.33 ,M; kns=0.15±0.029 pmol x mg protein,1/3 min/,M; mean±SEM; n=3 concentration profiles). Protein binding studies using brain tissue showed negligible binding. In summary, this work identified three principle findings: (1) An apparent lack of quantifiable aminoguanidine BBB efflux; (2) a previously undescribed synaptosomal accumulation process for aminoguanidine; and (3) an interesting limitation of the BEI technique where unusual brain parenchymal sequestration yields values >100%. Copyright © 2001 John Wiley & Sons, Ltd. [source] Effects of cellulase and xylanase enzymes on the deconstruction of solids from pretreatment of poplar by leading technologiesBIOTECHNOLOGY PROGRESS, Issue 2 2009Rajeev Kumar Abstract Comparative data is presented on glucose and xylose release for enzymatic hydrolysis of solids produced by pretreatment of poplar wood by ammonia fiber expansion (AFEX), ammonia recycled percolation (ARP), controlled pH, dilute acid, flowthrough (FT), lime, and sulfur dioxide (SO2) technologies. Sugar solubilization was measured for times of up to 72 h using cellulase supplemented with ,-glucosidase at an activity ratio of 1:2, respectively, at combined protein mass loadings of 5.8,116 mg/g of glucan in poplar wood prior to pretreatment. In addition, the enzyme cocktail was augmented with up to 11.0 g of xylanase protein per gram of cellulase protein at combined cellulase and ,-glucosidase mass loadings of 14.5 and 29.0 mg protein (about 7.5 and 15 FPU, respectively)/g of original potential glucose to evaluate cellulase,xylanase interactions. All pretreated poplar solids required high protein loadings to realize good sugar yields via enzymatic hydrolysis, and performance tended to be better for low pH pretreatments by dilute sulfuric acid and sulfur dioxide, possibly due to higher xylose removal. Glucose release increased nearly linearly with residual xylose removal by enzymes for all pretreatments, xylanase leverage on glucan removal decreased at high cellulase loadings. Washing the solids improved digestion for all pretreatments and was particularly beneficial for controlled pH pretreatment. Furthermore, incubation of pretreated solids with BSA, Tween 20, or PEG6000 prior to adding enzymes enhanced yields, but the effectiveness of these additives varied with the type of pretreatment. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009 [source] Production and Molecular Characterization of Clinical Phase I Anti-Melanoma Mouse IgG3 Monoclonal Antibody R24BIOTECHNOLOGY PROGRESS, Issue 5 2001Sven E. Kemminer R24 is a mouse IgG3 monoclonal antibody (mab) that reacts with the ganglioside GD3 expressed by cells of neuroectodermal origin. The anti-tumor activity of R24 has been demonstrated in initial phase I and pilot trials in patients suffering from metastatic melanoma. The purpose of this study was to investigate the biotechnological production and particularly the glycosylation of this clinically important antibody. Growth, metabolism, and IgG production of R24 secreting hybridoma cells were analyzed on 1 L bioreactor bench scale using repeated-batch mode. The amount of 57 mg of pure mab was obtained from 1.6 L crude supernatant by protein A chromatography. Western blot binding assays with sugar-specific lectins revealed glycosylation of the heavy chains, whereas no carbohydrates were detectable on the light chains. Because glycosylation is essential for antibody effector functions in vivo (such as complement fixation or binding to macrophage Fc receptors), mab R24 was subjected to both enzymatic deglycosylation using PNGase F and chemical deglycosylation by hydrazinolysis. Released glycans were structurally characterized by high pH anion exchange chromatography with pulsed amperometric detection (HPAEC-PAD), matrix assisted laser desorption ionization time-of-flight (MALDI-TOF), and electrospray ionization quadrupole time-of-flight (ESI-QTOF) mass spectrometry. Six major biantennary chains of the complex glycosylation phenotype were found with variations in galactosylation and core fucosylation. The predominant N-linked structure, indicating the high degree of agalactosyl glycoforms, was the agalacto biantennary chain with a relative percentage of 57% (51% core-fucosylated, 6% nonfucosylated). The second most abundant oligosaccharide was the monogalacto biantennary chain amounting to 30% (26% core- and 4% nonfucosylated). The antibody contained 0.46 ,g sialic acid per mg protein, which splits into 0.243 ,g Neu5Gc and 0.217 ,g Neu5Ac, corresponding to a Neu5Ac:Neu5Gc ratio of 1:1.06. Furthermore, the antigen specificity of R24 was determined by immunodetection of GD3 on thin-layer chromatograms, and real time GD3-antibody binding interactions were measured with an optical biosensor (BIAcore). From the structural data obtained in this study it is concluded that glycosylation of the antibody may be important in the clinical outcome of targeted anti-cancer immunotherapy. [source] Precipitation diagram and optimization of crystallization conditions at low ionic strength for deglycosylated dye-decolorizing peroxidase from a basidiomyceteACTA CRYSTALLOGRAPHICA SECTION F (ELECTRONIC), Issue 8 2005Shinya Saijo The growth of suitably sized protein crystals is essential for protein structure determination by X-ray crystallography. In general, crystals are grown using a trial-and-error method. However, these methods have been modified with the advent of microlitre dispensing-robot technology and of protocols that rapidly screen for crystal nucleation conditions. The use of one such automatic dispenser for mixing protein drops (1.3,2.0,µl in volume) of known concentration and pH with precipitating solutions (ejecting 2.0,µl droplets) containing salt is described here. The results of the experiments are relevant to a crystallization approach based on a two-step procedure: screening for the crystal nucleation step employing robotics followed by optimization of the crystallization conditions using incomplete factorial experimental design. Large crystals have successfully been obtained using quantities as small as 3.52,mg protein. [source] Catecholamine synthesis and metabolism in the central nervous system of mice lacking ,2 -adrenoceptor subtypesBRITISH JOURNAL OF PHARMACOLOGY, Issue 3 2009MA Vieira-Coelho Background and purpose:, This study investigates the role of ,2 -adrenoceptor subtypes, ,2A, ,2B and ,2C, on catecholamine synthesis and catabolism in the central nervous system of mice. Experimental approach:, Activities of the main catecholamine synthetic and catabolic enzymes were determined in whole brains obtained from ,2A -, ,2B - and ,2C -adrenoceptor knockout (KO) and C56Bl\7 wild-type (WT) mice. Key results:, Although no significant differences were found in tyrosine hydroxylase activity and expression, brain tissue levels of 3,4-dihydroxyphenylalanine were threefold higher in ,2A - and ,2C -adrenoceptor KO mice. Brain tissue levels of dopamine and noradrenaline were significantly higher in ,2A and ,2CKOs compared with WT [WT: 2.8 ± 0.5, 1.1 ± 0.1; ,2AKO: 6.9 ± 0.7, 1.9 ± 0.1; ,2BKO: 2.3 ± 0.2, 1.0 ± 0.1; ,2CKO: 4.6 ± 0.8, 1.5 ± 0.2 nmol·(g tissue),1, for dopamine and noradrenaline respectively]. Aromatic L-amino acid decarboxylase activity was significantly higher in ,2A and ,2CKO [WT: 40 ± 1; ,2A: 77 ± 2; ,2B: 40 ± 1; ,2C: 50 ± 1, maximum velocity (Vmax) in nmol·(mg protein),1·h,1], but no significant differences were found in dopamine ,-hydroxylase. Of the catabolic enzymes, catechol- O -methyltransferase enzyme activity was significantly higher in all three ,2KO mice [WT: 2.0 ± 0.0; ,2A: 2.4 ± 0.1; ,2B: 2.2 ± 0.0; ,2C: 2.2 ± 0.0 nmol·(mg protein),1·h,1], but no significant differences were found in monoamine oxidase activity between all ,2KOs and WT mice. Conclusions and implications:, In mouse brain, deletion of ,2A - or ,2C -adrenoceptors increased cerebral aromatic L-amino acid decarboxylase activity and catecholamine tissue levels. Deletion of any ,2 -adrenoceptor subtypes resulted in increased activity of catechol- O -methyltransferase. Higher 3,4-dihydroxyphenylalanine tissue levels in ,2A and ,2CKO mice could be explained by increased 3,4-dihydroxyphenylalanine transport. [source] Role of hydrogen sulphide in haemorrhagic shock in the rat: protective effect of inhibitors of hydrogen sulphide biosynthesisBRITISH JOURNAL OF PHARMACOLOGY, Issue 7 2004Ying-Yuan Pamela Mok Haemorrhagic shock (60 min) in the anaesthetized rat resulted in a prolonged fall in the mean arterial blood pressure (MAP) and heart rate (HR). Pre-treatment (30 min before shock) or post-treatment (60 min after shock) with inhibitors of cystathionine , lyase (CSE; converts cysteine into hydrogen sulphide (H2S)), dl-propargylglycine or , -cyanoalanine (50 mg kg,1, i.v.), or glibenclamide (40 mg kg,1, i.p.), produced a rapid, partial restoration in MAP and HR. Neither saline nor DMSO affected MAP or HR. Plasma H2S concentration was elevated 60 min after blood withdrawal (37.5±1.3 ,m, n=18 c.f. 28.9±1.4 ,m, n=15, P<0.05). The conversion of cysteine to H2S by liver (but not kidney) homogenates prepared from animals killed 60 min after withdrawal of blood was significantly increased (52.1±1.6 c.f. 39.8±4.1 nmol mg protein,1, n=8, P<0.05), as was liver CSE mRNA (2.7 ×). Both PAG (IC50, 55.0±3.2 ,m) and BCA (IC50, 6.5±1.2 ,m) inhibited liver H2S synthesizing activity in vitro. Pre-treatment of animals with PAG or BCA (50 mg kg,1, i.p.) but not glibenclamide (40 mg kg,1, i.p., KATP channel inhibitor) abolished the rise in plasma H2S in animals exposed to 60 min haemorrhagic shock and prevented the augmented biosynthesis of H2S from cysteine in liver. These results demonstrate that H2S plays a role in haemorrhagic shock in the rat. CSE inhibitors may provide a novel approach to the treatment of haemorrhagic shock. British Journal of Pharmacology (2004) 143, 881,889. doi:10.1038/sj.bjp.0706014 [source] Pharmacological effects of carcinine on histaminergic neurons in the brainBRITISH JOURNAL OF PHARMACOLOGY, Issue 5 2004Zhong Chen Carcinine (, -alanyl histamine) is an imidazole dipeptide. The present study was designed to characterize the pharmacological effects of carcinine on histaminergic activity in the brain and on certain neurobehavior. Carcinine was highly selective for the histamine H3 receptor over H1 or H2 receptor (Ki (,M)=0.2939±0.2188 vs 3621.2±583.9 or 365.3±232.8 ,M, respectively). Carcinine at a dose of 20 mg kg,1 slightly increased histidine decarboxylase (HDC) activity in the cortex (from 0.186±0.069 to 0.227±0.009 pmol mg protein,1 min,1). In addition, carcinine (10, 20, and 50 mg kg,1) significantly decreased histamine levels in mice brain. Like thioperamide, a histamine H3 receptor antagonist, carcinine (20, 50 ,M) significantly increased 5-HT release from mice cortex slices, but had no apparent effect on dopamine release. Carcinine (20 mg kg,1) significantly inhibited pentylenetetrazole-induced kindling. This inhibition was completedly reversed by (R)- , -methylhistamine, a representative H3 receptor agonist, and , -fluromethylhistidine, a selective HDC inhibitor. Carcinine (20 mg kg,1) ameliorated the learning deficit induced by scopolamine. This amelioration was reversed by (R)- , -methylhistamine as evaluated by the passive avoidance test in mice. Like thioperamide, carcinine dose-dependently increased mice locomotor activity in the open-field test. The results of this study provide first and direct evidence that carcinine, as a novel histamine H3 receptor antagonist, plays an important role in histaminergic neurons activation and might be useful in the treatment of certain diseases, such as epilepsy, and locomotor or cognitive deficit. British Journal of Pharmacology (2004) 143, 573,580. doi:10.1038/sj.bjp.0705978 [source] Characterisation of the binding of [3H]-SB-674042, a novel nonpeptide antagonist, to the human orexin-1 receptorBRITISH JOURNAL OF PHARMACOLOGY, Issue 2 2004Christopher J Langmead This study characterises the binding of a novel nonpeptide antagonist radioligand, [3H]SB-674042 (1-(5-(2-fluoro-phenyl)-2-methyl-thiazol-4-yl)-1-((S)-2-(5-phenyl-(1,3,4)oxadiazol-2-ylmethyl)-pyrrolidin-1-yl)-methanone), to the human orexin-1 (OX1) receptor stably expressed in Chinese hamster ovary (CHO) cells in both a whole cell assay and in a cell membrane-based scintillation proximity assay (SPA) format. Specific binding of [3H]SB-674042 was saturable in both whole cell and membrane formats. Analyses suggested a single high-affinity site, with Kd values of 3.76±0.45 and 5.03±0.31 nM, and corresponding Bmax values of 30.8±1.8 and 34.4±2.0 pmol mg protein,1, in whole cell and membrane formats, respectively. Kinetic studies yielded similar Kd values. Competition studies in whole cells revealed that the native orexin peptides display a low affinity for the OX1 receptor, with orexin-A displaying a ,five-fold higher affinity than orexin-B (Ki values of 318±158 and 1516±597 nM, respectively). SB-334867, SB-408124 (1-(6,8-difluoro-2-methyl-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) and SB-410220 (1-(5,8-difluoro-quinolin-4-yl)-3-(4-dimethylamino-phenyl)-urea) all displayed high affinity for the OX1 receptor in both whole cell (Ki values 99±18, 57±8.3 and 19±4.5 nM, respectively) and membrane (Ki values 38±3.6, 27±4.1 and 4.5±0.2 nM, respectively) formats. Calcium mobilisation studies showed that SB-334867, SB-408124 and SB-410220 are all functional antagonists of the OX1 receptor, with potencies in line with their affinities, as measured in the radioligand binding assays, and with approximately 50-fold selectivity over the orexin-2 receptor. These studies indicate that [3H]SB-674042 is a specific, high-affinity radioligand for the OX1 receptor. The availability of this radioligand will be a valuable tool with which to investigate the physiological functions of OX1 receptors. British Journal of Pharmacology (2004) 141, 340,346. doi:10.1038/sj.bjp.0705610 [source] Arachidonic acid-mediated cooxidation of all- trans -retinoic acid in microsomal fractions from human liverBRITISH JOURNAL OF PHARMACOLOGY, Issue 4 2000Louise Nadin The quantitative importance of prostaglandin H synthase (PGHS)-mediated cooxidation of all- trans -retinoic acid (ATRA) was evaluated in human liver microsomes (n=17) in relation to CYP-dependent ATRA 4-hydroxylation. Observed rates of ATRA cooxidation (4.6,20 pmol mg protein,1 min,1) and 4-hydroxylation (8.7,45 pmol mg protein,1 min,1) were quantitatively similar and exhibited similar individual variation (4 and 5 fold, respectively). From kinetic studies cooxidation was an efficient process in human hepatic microsomes (VmaxKm,1=0.25) compared with NADPH- and NADH-mediated 4-hydroxylation by CYP (VmaxKm,1=0.14 and 0.02, respectively). The capacity of lipid hydroperoxide metabolites of arachidonic acid to mediate ATRA oxidation was established directly, but downstream products (D, E, F and I-series prostaglandins) were inactive. cDNA-expressed CYPs supported ATRA oxidation by lipid hydroperoxides. Whereas CYPs 2C8, 2C9 and 3A4, but not CYPs 1A2 or 2E1, were effective catalysts of the NADPH-mediated reaction, cooxidation supported by 15(S)-hydroperoxyeicosatetraenoic acid was mediated by all five CYPs. The cooxidation reaction in human hepatic microsomes was inhibited by the CYP inhibitor miconazole. These findings indicate that ATRA oxidation is quantitatively significant in human liver. Lipid hydroperoxides generated by intracellular enzymes such as prostaglandin synthase and lipoxygenases are sources of activated oxygen for CYP-mediated deactivation of ATRA to polar products. British Journal of Pharmacology (2000) 131, 851,857; doi:10.1038/sj.bjp.0703579 [source] Matrix metalloproteinase 8 (neutrophil collagenase) in the pathogenesis of abdominal aortic aneurysm,BRITISH JOURNAL OF SURGERY (NOW INCLUDES EUROPEAN JOURNAL OF SURGERY), Issue 7 2005W. R. W. Wilson Background: Loss of elastin is the initiating event in abdominal aortic aneurysm (AAA) formation, whereas loss of collagen is required for continued expansion. The elastolytic matrix metalloproteinases (MMPs) 2 and 9 are well described, but the source of excessive collagenolysis remains undefined. The aim of this study was to determine the expression of MMP-8, a potent type I collagenase, in normal aorta and AAA. Methods: Infrarenal aortic biopsies were taken from 40 AAA and ten age-matched normal aortas. The concentrations of MMP-8 protein and its inhibitors, tissue inhibitor of metalloproteinase (TIMP) 1 and TIMP-2, were quantified by enzyme-linked immunosorbent assay. Immunohistochemistry was used to localize MMP-8 expression. Results: MMP-8 concentrations were significantly raised in AAA compared with normal aorta (active MMP-8: 4·5 versus 0·5 ng per mg protein, P < 0·001; total MMP-8: 16·6 versus 2·8 ng per mg protein, P < 0·001). Levels of TIMP-1 and TIMP-2 were significantly lower in AAA than in normal aortic samples (TIMP-1: 142·2 versus 302·8 ng per mg protein; P = 0·010; TIMP-2: 9·2 versus 33·1 ng per mg protein, P < 0·001). Immunohistochemistry localized MMP-8 to mesenchymal cells within the adventitia of the aortic wall. Conclusion: The high concentration of MMP-8 in aortic aneurysms represents a potent pathway for collagen degradation, and hence aneurysm formation and expansion. Copyright © 2005 British Journal of Surgery Society Ltd. Published by John Wiley & Sons, Ltd. [source] Is correction for protein concentration appropriate for protein adduct dosimetry?CANCER SCIENCE, Issue 2 2007Hypothesis, clues from an aflatoxin B1-exposed population Protein adducts are useful biomarkers for assessing exposure, metabolism and risk of carcinogens. Aflatoxin B1,albumin adducts (AAA) and protein carbonyl content (PCC) have long been used for assessing aflatoxin exposure and oxidative stress to proteins, and the quantitative data are almost exclusively expressed per mg protein. Given the large variation in protein concentrations in plasma among populations, this may not be the most appropriate method. The objective was to test the hypothesis that AAA and PCC should be expressed per mL plasma in population studies. AAA and PCC were analyzed among 402 subjects from three regions of China with a gradient in hepatocellular carcinoma (HCC) mortality ranging from 21 to 97 per 100 000. When biomarker values were expressed per mL plasma, the AAA level was significantly associated with plasma PCC (r = 0.262, P < 0.001), and adjusted levels of AAA and PCC paralleled HCC mortalities in the three regions, suggesting a role for aflatoxin-related oxidative stress in hepatocarcinogenesis in this population. In addition, there were statistically significant associations between both protein biomarkers, expressed per mL plasma, and the levels of alanine aminotransferase and aspartate aminotransferase in hepatitis B virus-infected subjects, suggesting roles for aflatoxin exposure, oxidative stress and hepatitis B virus infection in the development of HCC. The present data suggest that interindividual variation in plasma protein concentration may influence the dosimetry and relevant interpretation of protein biomarkers. (Cancer Sci 2007; 98: 140,146) [source] Optimum Dietary Protein Levels and Protein to Energy Ratios in Olive Flounder Paralichthys olivaceusJOURNAL OF THE WORLD AQUACULTURE SOCIETY, Issue 2 2005Kang-Woong Kim The olive flounder Paralichthys olivaceus is one of the most commercially important fish species in Korea. In order to formulate better diets for cultured olive flounder we evaluated the optimum dietary protein requirements for larval, fry and juvenile olive flounder, and the optimum dietary protein to energy ratio for juvenile olive flounder. Results of four separate experiments suggested that the optimum dietary protein requirements were 60% in larvae (0.3 g), 46.4,51.2% in 4.1-g juvenile, and 40,44% in 13.3 g growing olive flounder. The optimum dietary protein to energy ratio based on weight gain, feed efficiency, specific growth rate, and protein retention efficiency was 27,28 mg protein/kJ 2 energy (35 and 45% CP for diets containing 12.5 and 16.7 kJ energylg diet, respectively). [source] | |||||||||||||||||||