Effect of acute heat stress on heat shock protein 70 messenger RNA and on heat shock protein expression in the liver of broilers

1. The synthesis of heat shock protein 70 (Hsp70) mRNA and the expression of Hsp70 in the liver of broiler chickens submitted to acute heat stress (35 degrees C for 5 h) was investigated.2. Hsp70 expression was detected by SDS-PAGE and Western blot analysis using a polyclonal antiserum against Hsp70 of Blastocladiella emersonii. The specific signal of Hsp70 mRNA was analysed by Northern blot using as probe a Hsp70 cDNA of B. emersonii.3. An increase in the amount of Hsp70 was detected from the first up to the fifth hour of acute heat exposure. This increase in the amount of Hsp70 was accompanied by an increase in Hsp70 mRNA which peaked at 3 h.4. This study shows that the heat induced increase in Hsp70 mRNA and protein in broiler liver, in vivo, are time dependent, similar to that in mammals.

Net requirements of protein and energy for maintenance of wool and hair lambs in a tropical region

The objective of this work was to evaluate the total endogenous N losses, and protein and energy net requirements for maintenance in growing lambs. Thirty-four castrated lambs, 17 F1 Ideal X Ile de France wool and 17 Santa Inas hair lambs, averaging 20 +/- 0.14 kg BW, were used in the experiment. Five animals from each genotype were slaughtered at the beginning of the experiment and taken as controls. Diets (D) were composed of concentrate mix (C) and Cynodon sp. c.v. Tifton 85 hay (R), combined in three different ratios: D1 = 60C:40R; D2 = 40C:60R and D3 = 20C:80R. Animals of each group of three lambs, that showed BW of 20 kg at the beginning of the dietary regimen, were slaughtered when one of them reached 35 kg, what always happened to be the one fed with D1. Total endogenous N losses estimated for wool lambs were 250 mg N/kg BW0.75. For hair lambs, total endogenous N losses reached 324 mg N/kg BW0.75 . Hair lambs showed higher (P < 0.01) (29.9%) net requirements of protein for maintenance than wool lambs. In contrast, net energy (NE) requirement for maintenance was similar (P > 0.05) for both genotypes (74.27 kcal/kg BW0.75 per day), the average of the antilog of the two intercept values obtained from the estimated regression equations of heat production for zero metabolizable energy (ME) consumption. Further studies should be done to check if this trend is also true for metabolizable energy and protein in animals exhibiting BW gains in tropical region. (C) 2003 Elsevier B.V. B.V. All rights reserved.

Growth and metabolism of pacu (Piaractus mesopotamicus Holmberg 1887) juveniles fed diets containing different protein, lipid and carbohydrate levels

To verify the potential of lipids and carbohydrates to spare dietary protein and to understand the intermediary metabolism of interaction of these nutrients in pacu juveniles, an experiment was carried out to evaluate pacu physiological and performance parameters. The experimental design was completely randomized with 12 treatments in a 2 x 2 x 3 factorial arrangement, consisting of diets containing two digestible protein levels (200 and 230 g kg(-1) PD), two lipid levels (40 and 80 g kg(-1)) and three carbohydrate levels (410, 460 and 500 g kg(-1)). Fish-fed 230 g kg(-1) digestable protein (DP) showed increased glycaemia, decreased hepatic glycogen, as well as a smaller intake index and better feed conversion ratio. The higher dietary lipid level (80 g kg(-1)) reduced protein intake and serum protein concentration, increased liver and body fat content, but did not affect growth. At a lipid level of 80 g kg(-1), the increase in dietary carbohydrate levels promoted greater weight gain (WG), crude protein intake (CPI) and better feed conversion ratio (FCR). For fish fed diets containing 40 g kg(-1) lipid, the best energy-productive values (EPV) were obtained at 460 g kg(-1) carbohydrate. Increased levels of the main nutrients in the diets reduced the levels of serum triglycerides, while the increase in energy concentration increased the hepatosomatic (HSI) and glycaemia index values. Pacu used lipids as effectively as carbohydrates in the maximization of protein usage, as long as dietary protein was at a level of 230 g kg(-1) DP. The physiological parameters indicated that the best balance between the DP, dietary lipid and carbohydrate levels within the ranged this trial was obtained at 230, 40 and 460 g kg(-1), respectively, without lower growth.

REEVALUATION OF CHLORIDES REGULATION OF HEMOGLOBIN OXYGEN-UPTAKE - THE NEGLECTED CONTRIBUTION OF PROTEIN HYDRATION IN ALLOSTERISM

We have measured hemoglobin oxygen uptake vs. The partial pressure of oxygen, with independently controlled activities of chloride and water. This control is effected by combining different concentrations of NaCl and sucrose in the bathing solution to achieve: (i) water activities were varied and CI- activity was fixed, (ii) both water and CI- activities were varied with a traditional NaCI titration, or (iii) CI- activities were varied and water activity was fixed by adding compensating sucrose. Within this analysis, the CI--regulated loading of four oxygens can be described by the reaction Hb.CI- + 4 O-2 + 65 H2O reversible arrow Hb.4O(2).65H(2)O + CI-. The dissociation of a neatly integral chloride, rather than the nonintegral 1.6 chlorides inferred earlier from simple salt titration, demonstrates the need to recognize the potentially large contribution from changes in water activity when titrating weakly binding solutes. The single-chloride result might simplify structural considerations of the action of CI- in hemoglobin regulation.

Apparent digestibility of protein, energy, and amino acids in some selected feed ingredients for pacu Piaractus mesopotamicus

The apparent digestibility coefficients (ADCs) of amino acids (AA), protein, and energy in six feed ingredients (Brazilian fish meal, soybean meal, corn gluten meal, alcoholic yeast, corn, and wheat bran) we evaluated for pacu juveniles. In general, all ingredients showed high digestibility values for all AA, and differences among ADCs of individual amino acid were detected (P < 0.01). Corn gluten, soybean, and fish meals had the highest ADCs of AA. The ADCs of protein in fish meal, yeast, and corn gluten meal should not be used as AA digestibility indicators, because those showed differences up to 6.7% between the ADCs of protein and AA. All ingredients had lower ADCs of energy than corn gluten meal (P < 0.01). Lysine was the first limiting amino acid in fish meal, corn gluten meal, wheat bran, and corn, and the second limiting amino acid in soybean meal, as methionine was the first limiting amino acid in soybean meal and yeast. However, the soybean meal was the best quality protein source, as it had the highest digestible essential amino acid index. This demonstrated that digestible amino acid values can be used to formulate practical diets for pacu, preventing potential deficiencies or excess that might cause environmental and economic losses.

Protein synthesis studies in skeletal muscle of aging rats. II. In vitro studies with 0.5M potassium chloride washed polyribosomes

To investigate further the age-related reduction in muscle protein synthesis activity found previously using a crude polyribosome/pH 5 system (Pluskal et al., 1984), a 0.5M KCl washing procedure was utilized to remove the nonribosomal factors from polyribosomes isolated from male Sprague-Dawley rats in the following age groups: young (1 to 2 months), mature (12 months), and aged (22 to 24 months). Using a common source of enriched elongation factor fraction from young animals, it was not possible to demonstrate any significant difference (p > .05) in protein synthesis between the 0.5M KCl-washed polyribosomes isolated from the various age groups. Using a cell-free system containing young salt washed polyribosomes stimulated by the addition of 0.5M KCl-wash fractions, however, it was shown that the mature and aged salt-wash fractions were less (p < .05) active than material from young animals. Thus, the observed decline in protein synthesis efficiency during aging may be attributed to a reduced capacity to promote initiation/elongation by the nonribosomal salt wash fractions of muscle polyribosomes.

Effects of protein malnutrition on glucose tolerance in rats with alloxan-induced diabetes

Protein-calorie malnutrition produces glucose intolerance and reduced insulin release in response to glucose. Rats adapted to low- or high-protein diets show an increased resistance to the diabetogenic action of a single dose of streptozotocin or alloxan. To determine the effects of dietary protein level on pancreatic function, we measured serum glucose levels under basal conditions and during the oral glucose tolerance test (GTT) performed before and after a single dose of alloxan administered to rats fed a 25% or a 6% protein diet for a period of 8 weeks. The incidence of mild hyperglycemia (serum glucose > 250 mg/dl) was greater among the rats fed the 25% protein diet (81%) than among those fed the 6% protein diet (42%). During the GTT performed before alloxan administration the serum glucose levels of the rats fed the 6% protein diet were not found to be significantly different from those of rats fed the 25% protein diet. During the GTT performed after alloxan injection all rats showed intolerance to the substrate (serum glucose > 160 mg/dl 120 min after glucose administration) regardless of whether basal serum glucose was normal or high. In summary, alloxan was less effective in producing basal hyperglycemia in the rats fed the 6% protein diet than in those fed the 25% protein diet but caused glucose intolerance during the oral GTT in both groups. Thus, it seems that feeding a 6% protein diet to rats offers only partial protection against the toxic effects of alloxan.

Conserved sequences in the β subunit of archaeal and eukaryal translation initiation factor 2 (elF2), absent from elF5, mediate interaction with elF2γ

The eukaryotic translation initiation factor 2 (eIF2) binds the methionyl-initiator tRNA in a GTP-dependent mode. This complex associates with the 40 S ribosomal particle, which then, with the aid of other factors, binds to the 5' end of the mRNA and migrates to the first AUG codon, where eIF5 promotes GTP hydrolysis, followed by the formation of the 80 S ribosome. Here we provide a comparative sequence analysis of the β subunit of eIF2 and its archaeal counterpart (aIF2β). aIF2β differs from eIF2β in not possessing an N-terminal extension implicated in binding RNA, eIF5 and eIF2B. The remaining sequences are highly conserved, and are shared with eIF5. Previously isolated mutations in the yeast eIF2β, which allow initiation of translation at UUG codons due to the uncovering of an intrinsic GTPase activity in eIF2, involve residues that are conserved in aIF2β, but not in eIF5. We show that the sequence of eIF2B homologous to aIF2β is sufficient for binding eIF2γ, the only subunit with which it interacts, and comprises, at the most, 78 residues, eIF5 does not interact with eIF2γ, despite its similarity with eIF2β, probably because of a gap in homology in this region. These observations have implications for the evolution of the mechanism of translation initiation.

Changes in protein fractions, trypsin inhibitor and proteolytic activity in the cotyledons of germinating chickpea

The chickpea seed germination was carried out in 6 days. During the period it was observed a little variation on total nitrogen contents, however the non protein nitrogen was double. A decrease of 19.1 and 20.6% in relation to total nitrogen was observed to the total globulin and albumin fractions, respectively. The gel filtration chromatography on Sepharose CL-6B and SDS-PAGE demonstrated alterations on the distribution patterns of the albumin and total globulin fractions between the initial and the sixth day of germination suggesting the occurrence of protein degradation in the germination process.The assay for acid protease only appeared in the albumin fraction with casein and chickpea total globulin as substrates, whereas the former was more degradated than the latter, however the transformations detected in the protein fractions apppear indicated that others enzymes could be acting during the process. The trypsin inhibitor activity had a little drop after six day of germination indicating a possible increase on the digestibility of the proteins.

Evidence for involvement of Saccharomyces cerevisiae protein kinase C in glucose induction of HXT genes and derepression of SUC2

The PKC1 gene in the yeast Saccharomyces cerevisiae encodes protein kinase C that is known to control a mitogen-activated protein (MAP) kinase cascade consisting of Bck1, Mkk1 and Mkk2, and Mpk1. This cascade affects the cell wall integrity but the phenotype of Pkc1 mutants suggests additional targets which have not yet been identified. We show that a pkc1Δ mutant, as opposed to mutants in the MAP kinase cascade, displays two major defects in the control of carbon metabolism. It shows a delay in the initiation of fermentation upon addition of glucose and a defect in derepression of SUC2 gene after exhaustion of glucose from the medium. After addition of glucose the production of both ethanol and glycerol started very slowly. The V max of glucose transport dropped considerably and Northern blot analysis showed that induction of the HXT1, HXT2 and HXT4 genes was strongly reduced. Growth of the pkc1Δ mutant was absent on glycerol and poor on galactose and raffinose. Oxygen uptake was barely present. Derepression of invertase activity and SUC2 transcription upon transfer of cells from glucose to raffinose was deficient in the pkc1Δ mutant as opposed to the wild-type. Our results suggest an involvement of Pkc1p in the control of carbon metabolism which is not shared by the downstream MAP kinase cascade. © 2002 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.