Effect of probiotic-, bacteriophage-, or organic acid-supplemented feeds or fermented soybean meal on the growth performance, acute-phase response, and bacterial shedding of grower pigs challenged with Salmonella enterica serotype Typhimurium.

A 28-d experiment evaluated the growth performance, acute-phase response, and bacterial shedding patterns in pigs (n = 108; initially, 38.7 ± 6.7 kg) fed 6 treatment diets, including a control diet with no antimicrobial agents (CON), a positive control diet containing chlortetracycline, 100 mg/kg (CT), a diet containing anti-Salmonella Typhimurium bacteriophage, 3 × 10(9) plaque-forming units/kg of feed (ASB), Lactobacillus plantarum CJLP56, 6.5 × 10(8) cfu/kg of feed (LP), 0.2% microencapsulated organic acids (MOA), or 5% fermented soybean meal (FSM). Pigs were fed the diets for 2 wk before and 2 wk after challenging orally with Salmonella enterica serotype Typhimurium (SalT). Before bacterial challenge, ADFI was similar in all groups. After SalT challenge, ADFI of CON pigs was less (P < 0.05) than all other groups. Before challenge, pigs on MOA, FSM, and CT diets had greater (P < 0.05) ADG and G:F than CON pigs. After challenge (wk 3 to 4) and during the overall experimental period (wk 1 to 4), ADG of all treatment groups and G:F of all treatment groups except the LP group were greater (P < 0.05) than those of CON pigs. Relative to all other treatments, CON and LP pigs had greater (P < 0.05) bacterial shedding scores on d 7 after SalT challenge. At d 14 postchallenge, shedding scores declined (P < 0.05) in all treatment groups compared with CON pigs. Serum haptoglobin for all treatment groups increased from d 0 concentrations on d 6 postchallenge and declined to prechallenge concentrations on d 13 (P < 0.05). Circulating IGF-I concentrations declined from 2 to 6 d postchallenge and increased again by d 13 in ASB and LP groups, did not decline in FSM and CT groups, and continuously declined through d 13 in CON and LP groups (P < 0.05). However, in MOA group, IGF-I concentrations declined from preinfection concentrations on d 2, increased on d 4, and declined again until d 13 (P < 0.05). The serum concentrations of the cytokines IL-6 and IL-1ß were not generally affected by SalT challenge. In conclusion, acute infection of growing pigs with SalT was associated with short-term febrile responses in most pigs and reductions in ADFI and ADG of CON pigs. Compared with the CON diet, ASB, FSM, and MOA diets had a similar benefit to the antibiotic-supplemented diet in improving the performance of growing pigs, especially after bacterial challenge. However, further work needs to be done to better understand their mode of action in this class of pigs.

Glucose analogue inhibitors of glycogen phosphorylase: from crystallographic analysis to drug prediction using GRID force-field and GOLPE variable selection.

Several inhibitors of the large regulatory enzyme glycogen phosphorylase (GP) have been studied in crystallographic and kinetic experiments. GP catalyses the first step in the phosphorylysis of glycogen to glucose-l-phosphate, which is utilized via glycolysis to provide energy to sustain muscle contraction and in the liver is converted to glucose. alpha-D-Glucose is a weak inhibitor of glycogen phosphorylase form b (GPb, K(i) = 1.7 mM) and acts as a physiological regulator of hepatic glycogen metabolism. Glucose binds to phosphorylase at the catalytic site and results in a conformational change that stabilizes the inactive T state of the enzyme, promoting the action of protein phosphatase 1 and stimulating glycogen synthase. It has been suggested that in the liver, glucose analogues with greater affinity for glycogen phosphorylase may result in a more effective regulatory agent. Several N-acetyl glucopyranosylamine derivatives have been synthesized and tested in a series of crystallographic and kinetic binding studies with GPb. The structural results of the bound enzyme-ligand complexes have been analysed together with the resulting affinities in an effort to understand and exploit the molecular interactions that might give rise to a better inhibitor. Comparison of the N-methylacetyl glucopyranosylamine (N-methylamide, K(i) = 0.032 mM) with the analogous beta-methylamide derivative (C-methylamide, K(i) = 0.16 mM) illustrate the importance of forming good hydrogen bonds and obtaining complementarity of van der Waals interactions. These studies also have shown that the binding modes can be unpredictable but may be rationalized with the benefit of structural data and that a buried and mixed polar/non-polar catalytic site poses problems for the systematic addition of functional groups. Together with previous studies of glucose analogue inhibitors of GPb, this work forms the basis of a training set suitable for three-dimensional quantitative structure-activity relationship studies. The molecules in the training set are void of problems and potential errors arising from the alignment and bound conformations of each of the ligands since the coordinates were those determined experimentally from the X-ray crystallographic refined ligand-enzyme complexes. The computational procedure described in this work involves the use of the program GRID to describe the molecular structures and the progam GOLPE to obtain the partial least squares regression model with the highest prediction ability. The GRID/GOLPE procedure performed using 51 glucose analogue inhibitors of GPb has good overall predictivity [standard deviation of error predictions (SDEP) = 0.98 and Q(2) = 0.76] and has shown good agreement with the crystallographic and kinetic results by reliably selecting regions that are known to affect the binding affinity.

The design of potential antidiabetic drugs: experimental investigation of a number of beta-D-glucose analogue inhibitors of glycogen phosphorylase.

alpha-D-glucose is a weak inhibitor (Ki = 1.7 mM) of glycogen phosphorylase (GP) and acts as physiological regulator of hepatic glycogen metabolism; it binds to GP at the catalytic site and stabilizes the inactive T state of the enzyme promoting the action of protein phosphatase 1 and stimulating glycogen synthase. The three-dimensional structures of T state rabbit muscle GPb and the GPb-alpha-D-glucose complex have been exploited in the design of better regulators of GP that could shift the balance between glycogen synthesis and glycogen degradation in favour of the former. Close examination of the catalytic site with alpha-D-glucose bound shows that there is an empty pocket adjacent to the beta-1-C position. beta-D-glucose is a poorer inhibitor (Ki = 7.4 mM) than alpha-D-glucose, but mutarotation has prevented the binding of beta-D-glucose in T state GP crystals. A series of beta-D-glucose analogues has been designed and tested in kinetic and crystallographic experiments. Several compounds have been discovered that have an increased affinity for GP than the parent compound.

Design of inhibitors of glycogen phosphorylase: a study of alpha- and beta-C-glucosides and 1-thio-beta-D-glucose compounds.

alpha-D-Glucose is a weak inhibitor of glycogen phosphorylase b (Ki = 1.7 mM) and acts as a physiological regulator of hepatic glycogen metabolism. Glucose binds to phosphorylase at the catalytic site and results in a conformational change that stabilizes the inactive T state of the enzyme, promoting the action of protein phosphatase 1 and stimulating glycogen synthase. It has been suggested that, in the liver, glucose analogues with greater affinity for glycogen phosphorylase may result in a more effective regulatory agent. Several alpha- and beta-anhydroglucoheptonic acid derivatives and 1-deoxy-1-thio-beta-D-glucose analogues have been synthesized and tested in a series of crystallographic and kinetic binding studies with glycogen phosphorylase. The structural results of the bound enzyme-ligand complexes have been analyzed, together with the resulting affinities, in an effort to understand and exploit the molecular interactions that might give rise to a better inhibitor. This work has shown the following: (i) Similar affinities may be obtained through different sets of interactions. Specifically, in the case of the alpha- and beta-glucose-C-amides, similar Ki's (0.37 and 0.44 mM, respectively) are obtained with the alpha-anomer through interactions from the ligand via water molecules to the protein and with the beta-anomer through direct interaction from the ligand to the protein. Thus, hydrogen bonds through water can contribute binding energy similar to that of hydrogen bonds directly to the protein. (ii) Attempts to improve the inhibition by additional groups did not always lead to the expected result. The addition of nonpolar groups to the alpha-carboxamide resulted in a change in conformation of the pyranose ring from a chair to a skew boat and the consequent loss of favorable hydrogen bonds and increase in the Ki. (iii) The addition of polar groups to the alpha-carboxamide led to compounds with the chair conformation, and in the examples studied, it appears that hydration by a water molecule may provide sufficient stabilization to retain the chair conformation. (iv) The best inhibitor was N-methyl-beta-glucose-C-carboxamide (Ki = 0.16 mM), which showed a 46-fold improvement in Ki from the parent beta-D-glucose. The decrease in Ki may be accounted for by a single hydrogen bond from the amide nitrogen to a main-chain carbonyl oxygen, an increase in entropy through displacement of a water molecule, and favorable van der Waals contacts between the methyl substituent and nonpolar protein residues.(ABSTRACT TRUNCATED AT 250 WORDS)

Inhibition of HIV replication by amino-sugar derivatives.

The plant alkaloids castanospermine, dihydroxymethyldihydroxypyrrolidine and deoxynojirimycin have recently been shown to have potential anti-HIV activity [(1987) Proc. Natl. Acad. Sci. USA 84, 8120-8124; (1987) Nature 330, 74-77; (1987) Lancet i, 1025-1026]. They are thought to act by inhibiting alpha-glucosidase I, an enzyme involved in the processing of N-linked oligosaccharides on glycoproteins. We report here the relative efficacy of a spectrum of amino-sugar derivatives as inhibition of HIV cytopathicity. Several alpha-glucosidase inhibitors and alpha-fucosidase inhibitors were found to be active at concentrations which were non-cytotoxic.

A modified electrode for the electrochemical reduction of isoflurane.

The popular anaesthetic gas isoflurane is found to be electrochemically inert on a wide range of conventional electrode materials in both aqueous and non-aqueous solvents. However the chemically modified electrode produced by depositing the novel electroactive polymer, poly(vinylfluoranthene), onto a platinum electrode is shown to mediate electron transfer to the anaesthetic thus bringing about its electro-reduction and so offers the promise of amperometric electrochemical sensors based on this electrode.