Substitution of both histidines in the active site of yeast alcohol dehydrogenase 1 exposes underlying pH dependencies.

Histidine residues 44 and 48 in yeast alcohol dehydrogenase (ADH) bind to the coenzymes NAD(H) and contribute to catalysis. The individual H44R and H48Q substitutions alter the kinetics and pH dependencies, and now the roles of other ionizable groups in the enzyme were studied in the doubly substituted H44R/H48Q ADH. The substitutions make the enzyme more resistant to inactivation by diethyl pyrocarbonate, modestly improve affinity for coenzymes, and substantially decrease catalytic efficiencies for ethanol oxidation and acetaldehyde reduction.

Specific base catalysis by yeast alcohol dehydrogenase I with substitutions of histidine-48 by glutamate or serine residues in the proton relay system.

His-48 in yeast alcohol dehydrogenase I (His 51 in horse liver alcohol dehydrogenase) is a highly conserved residue in the active sites of many alcohol dehydrogenases. The imidazole group of His-48 may participate in base catalysis of proton transfer as it is linked by hydrogen bonds through the 2'-hydroxyl group of the nicotinamide ribose and the hydroxyl group of Thr-45 to the hydroxyl group of the alcohol bound to the catalytic zinc. In this study, His-48 was substituted with a glutamic acid residue to determine if a carboxylate could replace imidazole or to a serine residue to determine if the exposure of the 2'-hydroxyl group of the ribose to solvent would allow proton transfer to water without base catalysis.

Potential of electrospun cationic BSA fibers to guide osteogenic MSC differentiation via surface charge and fibrous topography.

Large or complex bone fractures often need clinical treatments for sufficient bone repair. New treatment strategies have pursued the idea of using mesenchymal stromal cells (MSCs) in combination with osteoinductive materials to guide differentiation of MSCs into bone cells ensuring complete bone regeneration. To overcome the challenge of developing such materials, fundamental studies are needed to analyze and understand the MSC behavior on modified surfaces of applicable materials for bone healing.

Carbohydrate-Based Polymer Brushes Prevent Viral Adsorption on Electrostatically Heterogeneous Interfaces.

Chemical heterogeneity on biomaterial surfaces can transform its interfacial properties, rendering nanoscale heterogeneity profoundly consequential during bioadhesion. To examine the role played by chemical heterogeneity in the adsorption of viruses on synthetic surfaces, a range of novel coatings is developed wherein a tunable mixture of electrostatic tethers for viral binding, and carbohydrate brushes, bearing pendant α-mannose, β-galactose, or β-glucose groups, is incorporated. The effects of binding site density, brush composition, and brush architecture on viral adsorption, with the goal of formulating design specifications for virus-resistant coatings are experimentally evaluated.

Bioinstructive Coatings for Hematopoietic Stem Cell Expansion Based on Chemical Vapor Deposition Copolymerization.

We report the chemical vapor deposition (CVD) of dual-functional polymer films for the specific and orthogonal immobilization of two biomolecules (notch ligand delta-like 1 (DLL1) and an RGD-peptide) that govern the fate of hematopoietic stem and progenitor cells. The composition of the CVD polymer and thus the biomolecule ratio can be tailored to investigate and optimize the influence of the relative surface concentrations of biomolecules on stem cell behavior. Prior to cell experiments, all surfaces were characterized by infrared reflection adsorption spectroscopy, time-of-flight secondary ion mass spectrometry, and X-ray photoelectron spectroscopy to confirm the presence of both biomolecules.

Backbone-Degradable Polymers Prepared by Chemical Vapor Deposition.

Polymers prepared by chemical vapor deposition (CVD) polymerization have found broad acceptance in research and industrial applications. However, their intrinsic lack of degradability has limited wider applicability in many areas, such as biomedical devices or regenerative medicine. Herein, we demonstrate, for the first time, a backbone-degradable polymer directly synthesized via CVD.

Readily available hydrogen bond catalysts for the asymmetric transfer hydrogenation of nitroolefins.

This paper focuses on readily accessible thiourea hydrogen bond catalysts derived from amino acids, whose steric and electronic features are modulated by their degree of substitution at the carbinol carbon center. These catalysts were applied in the asymmetric transfer hydrogenation of nitroolefins furnishing the chiral products in up to 99% yield and 86% enantiomeric excess. The proposed catalyst's mode of action is supported by mechanistic investigations.

Appropriate statistical methods to compare dose responses of methionine sources.

Two sources of methionine (Met) activity are frequently used in commercial feed formulation: DL-2-hydroxy-4-(methylthio) butanoic acid (HMTBA), most commonly available as an 88% solution with 12% water; and DL-methionine (DLM, 99% powder). Despite the fact that both compounds have been in commercial use for over 50 yr, controversy and confusion remain with respect to their relative bioefficacy (RBE). This paper presents a review of the use of a nonlinear common plateau asymptotic regression technique (NLCPAR) that has been used to compare the 2 Met sources with particular emphasis on the validity of the basic assumptions of that model.

A multiple regression model approach to contrast the performance of 2-hydroxy-4-methylthio butanoic acid and DL-methionine supplementation tested in broiler experiments and reported in the literature.

The purpose of this paper was to compile all available literature comparing the relative performance of 2-hydroxy-4-methylthio butanoic acid (HMTBA) with DL-methionine (DLM) in broiler chickens and using multiple regression techniques, to estimate the predicted dose responses and relative performance of the 2 Met compounds for gain and feed conversion (FC). A database was developed that contained all available broiler studies in which HMTBA and DLM were both present in the same study; weight gain was recorded; Met addition, age of birds, and duration of study were defined; and an unsupplemented control treatment was present. Sixty-two references complied with these criteria and included 100 experiments with 427 observations for HMTBA and 411 for DLM.

Therapeutic efficacy of water-soluble lincomycin-spectinomycin powder against porcine proliferation enteropathy in a European field study.

Controlled clinical trials to a standardised protocol were conducted into the effect of a water-soluble antibiotic on proliferative enteropathy and its causative agent (Lawsonia intracellularis) on commercial pig farms at six sites in four European countries. Clinical signs of the disease and L intracellularis-specific polymerase chain reaction (PCR)-positive pigs were detected in pens of six- to 12-week-old pigs (weighing 5 to 55 kg) immediately before each trial. Matched pens of randomised pigs were either left unmedicated (32 to 59 pigs per trial), or medicated orally with 10 mg/kg of a water-soluble combination of lincomycin and spectinomycin powder (21 and 42 mg, respectively, of antibiotic activity per litre) for either seven days (33 to 61 pigs per trial), or 14 days (33 to 61 pigs per trial), delivered via the drinking water.

Estimating the maximum effective dose in a quantitative dose-response experiment.

A simulation study was conducted to compare several procedures for estimating the maximum effective dose in a quantitative dose-response experiment. Using four equally spaced dose levels, data were generated from four different model types: the quadratic growth curve, the Mitcherlich growth curve, the linear-linear plateau spline model, and the quadratic-linear plateau spline model. Each model type was parameterized to create three different model ranges, and for each range, data were generated from populations with three different standard deviations.

Production performance of dairy cattle administered recombinantly derived bovine somatotropin (USAN, Somavubove) daily: a dose range study.

Herds (n = 9) were used on which cattle (n = 598) received 0, 4.3, 8.6, 12.

Crystallization and preliminary crystallographic studies of Saccharomyces cerevisiae alcohol dehydrogenase I.

The cytoplasmic yeast alcohol dehydrogenase I crystallized at 5 degrees C as hexagonal plates or short columns in the presence of NAD+ and 2,2,2-trifluoroethanol, in sodium N-tris(hydroxymethyl)methyl-3-aminopropanesulfonate buffer at pH 8.2 to 8.6, using polyethylene glycol 4000 as precipitant.

Alpha-isoenzyme of alcohol dehydrogenase from monkey liver. Cloning, expression, mechanism, coenzyme, and substrate specificity.

The cDNA for the alpha-isoenzyme from rhesus monkey (Macaca mulatta) liver was cloned and expressed in yeast. The alpha-isoenzymes of human and monkey liver alcohol dehydrogenase differ from the other human and horse liver enzymes in having Met57, Ala93, and Val116 instead of Leu57, Phe93, and Leu116 in the substrate binding pocket and Gly47 instead of Arg47 near the pyrophosphate moiety of the coenzyme. The effects of these differences on the kinetic mechanism, substrate specificity, and coenzyme binding were studied with the purified, recombinant monkey alpha-isoenzyme (MmADH alpha) and mutated enzymes with Gly47 substituted with His or Arg.

Postparturient metabolic and production responses in cows previously exposed to long-term treatment with somatotropin.

Postparturient metabolic and production responses were studied for 85 multiparous cows consisting of controls and animals that had been previously exposed to long-term treatment with bST. Older cows previously exposed to bST at high doses (51.6 mg/d) had lower milk yields than controls.

Catalysis by yeast alcohol dehydrogenase.

Table 7 presents a brief summary of the effects of various mutations on some of the relevant kinetic constants. The results illustrate several important features of the use of site-directed mutagenesis in exploring structure and function of enzymes. Note that most of the mutations affect a given step or kinetic parameter in the mechanism, such as the binding of NAD+ or the turnover number with ethanol.

LH and FSH response of Holstein heifers to fertirelin acetate, gonadorelin and buserelin.

This study was conducted to describe the changes in serum LH and FSH concentrations in Holstein heifers following intramuscular (i.m.) injection of various dosages of fertirelin acetate and other commercially available GnRH products at their labeled dosages.

Efficacy of ceftiofur hydrochloride for treatment of experimentally induced colibacillosis in neonatal swine.

Ceftiofur hydrochloride was tested for effectiveness against induced colibacillosis in neonatal swine. In this model, pigs less than 12 hours old were inoculated via stomach tube with a virulent, K99+, nalidixic acid-resistant strain of Escherichia coli. Six hours after challenge exposure, 1 dose of ceftiofur was administered either IM or orally in experiment 1 and orally only in experiment 2.

Microbiological determination of neomycin in feeds: collaborative study.

A modification of the AOAC microbiological determination of neomycin in feeds was collaboratively studied by 12 laboratories. The official method was modified by substituting a constant salt concentration diluent for the feed extract diluent, preparing the agar medium in tris buffer, and performing the test with a monolayer plating system. Each laboratory performed single assays on 8 samples in a randomized sequence.

Effects of PGE1 or PGE2 and/or acetazolamide on expulsion of Nippostrongylus brasiliensis from rats.

PGE1 and PGE2 have been reported to enhance natural expulsion of Nippostrongylus brasiliensis, a nematode parasite, from the intestine of the rat. Mucus production may also be a key element of worm rejection. Our study attempts to determine if 1) PGE1 or PGE2 alter the normal course of infection with N.

Determining animal drug combinations based on efficacy and safety.

A procedure for deriving drug combinations for animal health is used to derive an optimal combination of 200 mg of novobiocin and 650,000 IU of penicillin for nonlactating cow mastitis treatment. The procedure starts with an estimated second order polynomial response surface equation. That surface is translated into a probability surface with contours called isoprobs.

Evaluation of a topical treatment, alone and in combination with a detergent, for generalized demodicosis.

Thirty dogs (20 treated, 10 controls) with naturally-acquired generalized demodicosis were utilized to evaluate the bio-activity and safety of a liquid concentrate formulation of amitraz, with or without the addition of a nonionic detergent. The detergent was added to the treatment mixture to enhance wetting and thereby reduce the number of treatments required to return diseased animals to a normal state. Three--six miticide treatments were topically applied to dogs at 14-day intervals, at a concentration of 250 parts per million active drug.

Microbiological determination of neomycin in feeds and formulated products.

An AOAC modified method is described for the microbiological assay of neomycin, which has been adapted to include complete feeds, supplements, premixes, liquids, oil suspensions, boluses, and antibiotic-impregnated paper. The method features a more sensitive standard response line with a monolayer plating system. The use of a buffered plating medium in place of the water-prepared medium results in a curve with less degree of slope, which allows for more accurate interpretation of the standard response.

Evaluation of a sponge-on therapy for canine scabies.

Forty dogs (20 treated, 20 controls) were utilized to evaluate a new treatment for naturally acquired canine scabies. A liquid concentrate formulation of amitraz was diluted and applied as a sponge-on therapy. Ninety-four percent of the dogs treated with the scabicide were cleared of mites and returned to clinical normality with a single topical treatment; one dog was retreated, cleared of mites and was also returned to normality.

Chemotherapeutic treatment of naturally acquired generalized demodicosis.

Fifty-two dogs naturally parasitized with Demodex canis and having the generalized form of the disease were utilized to evaluate the efficacy and safety of single or multiple topical treatments with a liquid concentrate formulation of amitraz. Ten dogs (5 treated, 5 controls) were utilized to evaluate a single treatment. A single topical treatment with the miticide did not significantly reduce the incidence of dogs with mites, however, significant clinical improvement resulted.