Reduced use of antibiotic growth promoters in diets fed to weanling pigs: dietary tools, part 1

Antibiotics improve health and productive performance of pigs. There is increasing evidence that other dietary ingredients may provide similar, but probably not identical, benefits. A properly prepared immune egg product, spray-dried plasma, or a high level of dietary zinc can provide protection against enteric infection in young pigs. Spray-dried plasma also increases growth rate dramatically, and zinc increases it substantially. The evidence in refereed publications also indicates that organic acids increase growth rate substantially, but does not clearly identify the most effective acid or combination of acids. Whey proteins, lactose, mannan oligosaccharide, or a high level of dietary copper provides a modest but useful increase in growth rate. It is not clear whether conventional egg products or other milk proteins may be beneficial. There are many other feed ingredients commercially available that may improve health and productive performance of pigs.     

Paracoccus pantotrophus pseudoazurin is an electron donor to cytochrome c peroxidase

The gene for pseudoazurin was isolated from Paracoccus pantotrophus LMD 52.44 and expressed in a heterologous system with a yield of 54.3 mg of pure protein per liter of culture. The gene and protein were shown to be identical to those from P. pantotrophus LMD 82.5. The extinction coefficient of the protein was re-evaluated and was found to be 3.00 mM(-1) cm(-1) at 590 nm. It was confirmed that the oxidized protein is in a weak monomer/dimer equilibrium that is ionic-strength-dependent. The pseudoazurin was shown to be a highly active electron donor to cytochrome c peroxidase, and activity showed an ionic strength dependence consistent with an electrostatic interaction. The pseudoazurin has a very large dipole moment, the vector of which is positioned at the putative electron-transfer site, His81, and is conserved in this position across a wide range of blue copper proteins. Binding of the peroxidase to pseudoazurin causes perturbation of a set of NMR resonances associated with residues on the His81 face, including a ring of lysine residues. These lysines are associated with acidic residues just back from the rim, the resonances of which are also affected by binding to the peroxidase. We propose that these acidic residues moderate the electrostatic influence of the lysines and so ensure that specific charge interactions do not form across the interface with the peroxidase.     

Effect of diazepam treatment on metabolic indices in trained and untrained rats

Exercise training, like diazepam, is commonly employed as a means of reducing anxiety. Both diazepam and exercise training have been shown to modify carbohydrate and lipid metabolism as well as influence calcium metabolism in skeletal muscle. As receptor binding and thereby efficacy of diazepam has been demonstrated to be modulated by the lipid environment of the receptor, and changes in calcium levels can affect a number of intracellular signalling pathways, we sought to determine if the interaction of both chronic diazepam and exercise training would modify selected metabolic indices in an animal model. For this purpose, muscle and liver glycogen, blood glucose and plasma free fatty acids (FFA) were measured in sedentary, exercise trained and exercise trained, acutely exhausted animals. Alterations in lipid and carbohydrate metabolism were observed in all experimental groups. Diazepam treatment alone exerts metabolic consequences, such as elevated muscle glycogen and plasma FFA and depressed blood glucose levels, which are similar to those observed with exercise training. When animals are acutely exercised to exhaustion, however, differences appear, including a reduced rise in plasma FFA, which suggests that long-term diazepam treatment does influence exercise metabolism, possibly as a result of effects on the sympatho-adrenal system.     

Electron transfer complexes of cytochrome c peroxidase from Paracoccus denitrificans containing more than one cytochrome

According to the model proposed in previous papers [Pettigrew, G. W., Prazeres, S., Costa, C., Palma, N., Krippahl, L., and Moura, J. J. (1999) The structure of an electron-transfer complex containing a cytochrome c and a peroxidase, J. Biol. Chem. 274, 11383-11389; Pettigrew, G. W., Goodhew, C. F., Cooper, A., Nutley, M., Jumel, K., and Harding, S. E. (2003) Electron transfer complexes of cytochrome c peroxidase from Paracoccus denitrificans, Biochemistry 42, 2046-2055], cytochrome c peroxidase of Paracoccus denitrificans can accommodate horse cytochrome c and Paracoccus cytochrome c(550) at different sites on its molecular surface. Here we use (1)H NMR spectroscopy, analytical ultracentrifugation, molecular docking simulation, and microcalorimetry to investigate whether these small cytochromes can be accommodated simultaneously in the formation of a ternary complex. The pattern of perturbation of heme methyl and methionine methyl resonances in binary and ternary solutions shows that a ternary complex can be formed, and this is confirmed by the increase in the sedimentation coefficient upon addition of horse cytochrome c to a solution in which cytochrome c(550) fully occupies its binding site on cytochrome c peroxidase. Docking experiments in which favored binary solutions of cytochrome c(550) bound to cytochrome c peroxidase act as targets for horse cytochrome c and the reciprocal experiments in which favored binary solutions of horse cytochrome c bound to cytochrome c peroxidase act as targets for cytochrome c(550) show that the enzyme can accommodate both cytochromes at the same time on adjacent sites. Microcalorimetric titrations are difficult to interpret but are consistent with a weakened binding of horse cytochrome c to a binary complex of cytochrome c peroxidase and cytochrome c(550) and binding of cytochrome c(550) to the cytochrome c peroxidase that is affected little by the presence of horse cytochrome c in the other site. The presence of a substantial capture surface for small cytochromes on the cytochrome c peroxidase has implications for rate enhancement mechanisms which ensure that the two electrons required for re-reduction of the enzyme after reaction with hydrogen peroxide are delivered efficiently.     

A left ventricle model to predict post-revascularization ejection fraction based on cine magnetic resonance images

The left ventricular ejection fraction (LVEF) is an important clinical indicator of the cardiac function and long-term outcome for patients with coronary artery disease. A biomechanical model of the left ventricle was developed to quantitatively predict post-revascularization LVEF based on noninvasive magnetic resonance imaging. The myocardium was categorized into normal, hibernating, and infarcted regions from the ventricular short-axis images. Assuming that hibernating tissue would potentially regain contractility after revascularization, the expected maximum post-revascularization LVEF was calculated for four patients with chronic left ventricular dysfunction. The predictions were within three ejection fraction points of the follow-up LVEFs. This model may be useful to estimate the outcome and efficacy of revascularization plans.     

Females of the sea urchin Strongylocentrotus purpuratus differ in the structures of their egg jelly sulfated fucans

The egg jelly coats of sea urchins contain sulfated fucans which bind to a sperm surface receptor glycoprotein to initiate the signal transduction events resulting in the sperm acrosome reaction. The acrosome reaction is an ion channel regulated exocytosis which is an obligatory event for sperm binding to, and fusion with, the egg. Approximately 90% of individual females of the sea urchin Strongylocentrotus purpuratus spawned eggs having only one of two possible sulfated fucan electrophoretic isotypes, a slow migrating (sulfated fucan I), or a fast migrating (sulfated fucan II) isotype. The remaining 10% of females spawned eggs having both sulfated fucan isotypes. The two sulfated fucan isotypes were purified from egg jelly coats and their structures determined by NMR spectroscopy and methylation analysis. Both sulfated fucans are linear polysaccharides composed of 1-->3-linked alpha-L-fucopyranosyl units. Sulfated fucan I is entirely sulfated at the O -2 position but with a heterogeneous sulfation pattern at O -4 position. Sulfated fucan II is composed of a regular repeating sequence of 3 residues, as follows: [3-alpha-L-Fuc p - 2,4(OSO3)-1-->3-alpha-L-Fuc p -4(OSO3)-1-->3-alpha-L-Fuc p -4(OSO3)- 1]n. Both purified sulfated fucans have approximately equal potency in inducing the sperm acrosome reaction. The significance of two structurally different sulfated fucans in the egg jelly coat of this species could relate to the finding that the sperm receptor protein which binds sulfated fucan contains two carbohydrate recognition modules of the C-type lectin variety which differ by 50% in their primary structure.      

Base-compositional biases and the bat problem. I. DNA-hybridization melting curves based on AT- and GC-enriched tracers.

We explored the interordinal relationships of mammals using DNA-DNA hybridization, with particular reference to the much-debated problem of whether the megabats and microbats are more closely related to each other than the megabats are to primates. To try to improve resolution when taxa are distantly related and the melting points of hybrids are low and difficult to distinguish, we increased the GC content of DNA by a fractionation method that used the same melting-point apparatus used in the hybridization studies. When we used GC-rich DNA as the tracer to make hybrids, the melting point of the self-hybrid shifted to a higher temperature as expected, but the behaviour of heterologous hybrids varied with the taxa being compared. When the melting point of the heterologous hybrid also shifted to a higher temperature so that the two compared taxa maintained the same or proportional distance, we called this ''following behaviour'', because the heterologous hybrid made with GC-rich tracer ''followed'' the GC-rich self-hybrid to higher temperatures. We also commonly saw anomalous behaviour, where the melting point of the heterologous hybrid shifted to a lower temperature when compared with an AT-rich hybrid. In these anomalous cases, the distance measured between the taxa increased markedly as a result of GC-enrichment, indicating that an underestimate of distance may have resulted from AT bias in DNA. This inference was supported by the finding that it was rare to observe a decrease in measured distance between taxa using GC-rich DNA, but very common to find an increase as would be expected from the generally higher AT contents of eutherian DNAs. Moreover, the most extreme cases, where distances changed most using GC-rich DNA, were usually those involving comparisons between taxa known to have the most extreme AT-biases among mammals, such as the megabats and rhinolophoid (including megadermatid) microbats. Our results show consistent underestimates of measured differences between eutherian taxa with extreme AT-biases.     

Cleavage of native type III collagen in the collagenase susceptible region by thermolysin.

Viscometric assays were used to demonstrate the activity of thermolysin (EC 3.4.24.4) on native type III collagen in solution. Analysis of the reaction products by sodium dodecyl sulphate-polyacrylamide gel electrophoresis and electron microscopic visualisation of segment long spacing aggregates demonstrated localised cleavage of the collagen in the collagenase susceptible region.