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Topic:Enzymes
Enzymes are biological catalysts that facilitate biochemical reactions in horses by lowering the activation energy required for these processes. They are involved in various physiological functions, including digestion, metabolism, and cellular repair. Common enzymes in equine biology include amylase, lipase, and lactate dehydrogenase, each playing a specific role in the breakdown of nutrients and energy production. The activity and concentration of these enzymes can vary in response to different physiological and pathological conditions, serving as potential indicators in veterinary diagnostics. This page compiles peer-reviewed research studies and scholarly articles that explore the function, regulation, and clinical implications of enzymes in equine health.
[Histochemical and biochemical changes in skeletal muscles of rhabdomyolysis-sensitive racehorses following exertion. III: Elevated activity of various antioxidant enzymes]. In this communication, the results of a histochemical and biochemical enzyme study on gluteus medius muscle of horses, sensitive to exertional myopathy, during attacks of rhabdomyolysis are presented. For the biochemical study the biopsy specimens investigated were selected by means of histological and enzyme histochemical staining methods. Dissected specimens were used which contained groups of muscle fibres with a high or low activity of glucose-6-phosphate dehydrogenase. The activity of glucose-6-phosphate dehydrogenase, phosphogluconate dehydrogenase, glutathione reductase, glutathione per...
The carbohydrate side chains of the major plasma serpins of horse and wallaby: analyses of enzymatic and chemically treated (including ‘Smith degradation’) protein blots by lectin binding. The carbohydrate side chains of the major plasma serpins of the horse and wallaby have been characterized by lectin analyses of protein blots from two-dimensional gels using the major human plasma serpin, alpha 1-protease inhibitor, as a control. Eight lectins were used in the characterization in conjunction with enzymatic deglycosylation of complex and high mannose side chains, chemical desialylation and defucosylation, and one round of 'Smith degradation', all being performed on the nitrocellulose blots. Assuming a standard complex side chain structure, the results of the 21 lectin/treatment...
The isoelectric focusing properties of serum alkaline phosphatase in disease and following prednisolone and phenylbutazone administration in the horse. This study was undertaken to ascertain if the isoelectric focusing pattern of serum alkaline phosphatase (AP) from sick horses with high activity is useful for determining its tissue origin. The effect of oral prednisolone and phenylbutazone therapy on this enzyme in healthy horses was also investigated. The sick horses were divided into three groups: hepatic, intestinal and miscellaneous. All sera had approximately thirteen bands of AP activity when focused on agarose gels with a pH gradient of 3.5 to 9.5. All the horses in the liver disease group had greater than 65% of enzyme activity in ba...
Hypoxanthine phosphoribosyltransferase activity in tissues and hypoxanthine concentrations in plasma and CSF of the horse in comparison with other species. 1. Plasma hypoxanthine and xanthine concentrations are very low in the horse and low in rat, mouse and greyhound compared to concentrations in beagles, man, sheep and rabbit. 2. Activities in erythrocytes of the main enzyme metabolizing hypoxanthine, hypoxanthine phosphori-bosyltransferase, show a similar pattern (Tax et al., 1976, Comp. Biochem. Physiol. 54B, 209-212); thus low activities have been found where plasma concentrations were low. 3. Hypoxanthine phosphoribosyltransferase activities in horse tissue other than erythrocytes are similar to those in man and rabbit with high activities ...
Histochemical and molecular determination of fiber types in chemically skinned single equine skeletal muscle fibers. Until now, there has been no reliable method for histochemical determination of fiber types of single skinned muscle fibers. The major problem arises from the fact that most histochemical techniques use cross-sections of a large group of fibers and compare a given fiber with those surrounding it. This is not possible with a single skinned fiber which has been separated from a bundle to be used for mechanical analysis. A further problem is that the skinning procedure itself may alter the staining pattern. We have developed a procedure by which multiple cross-sections of single skinned fibers ca...
Androgen and 19-norandrogen aromatization by equine and human placental microsomes. The ability of equine and human placental microsomes to aromatize testosterone and 19-nortestosterone was studied. When 3 microM [1 beta,2 beta-3H]testosterone was used as substrate, the specific activity of equine placental microsomal aromatase was 2.5 times higher than that of the human microsomal enzyme. Although 19-nortestosterone was aromatized 67 times more rapidly by equine than by human aromatase, we found that equine aromatase exhibited a markedly weaker affinity for this substrate than did the human enzyme. Competitive inhibition of testosterone aromatization by 19-nortestosterone oc...
A new method for hydrolyzing sulfate and glucuronyl conjugates of steroids. A new method for hydrolyzing steroid conjugates (both sulfates and glucuronides conjugates) that is efficient, effective, and inexpensive is described. This method comprises incubation of the conjugates--after salting-out into ethyl acetate or elution from a C18 cartridge--with anhydrous methanolic hydrogen chloride (methanolysis) for 10 min. It has been successfully applied to our routine radioimmunoassay screening and GC/MS confirmation studies of steroids in prerace and postrace equine urine samples. Comparative GC/MS studies on entire (male horse) urine samples showed that methanolysis gav...
Comparison of the binding of Ca2+ and Mn2+ to bovine alpha-lactalbumin and equine lysozyme. The enthalpy change of the binding of Ca2+ and Mn2+ to equine lysozyme was measured at 25 degrees C and pH 7.5 by batch microcalorimetry: delta H degrees Ca2+ = -76 +/- 5 kJ mol-1, delta H degrees Mn2+ = -21 +/- 10 kJ mol-1. Binding constants, log KCa2+ = 6.5 +/- 0.2 and log KMn2+ = 4.1 +/- 0.5, were calculated from the calorimetric data. Therefore, delta S degrees Ca2+ = -131 +/- 20 JK-1 mol-1 and delta S degrees Mn2+ = 8 +/- 44 JK-1 mol-1. Removal of Ca2+ induces small but significant changes in the circular dichroism spectrum, indicating the existence of a partially unfolded apo-conformatio...
Histochemistry of complex carbohydrates in the horse duodenal gland. Complex carbohydrates were examined in glandular cells of the horse duodenal gland by using lectin histochemical techniques. In the horse, the duodenal gland was distributed in the area from the uppermost part of the small intestine to a point about 6m caudal to the pylorus. It consisted of two types of cells, mucous and serous cells. The former was found in glands distributed almost all over this part, but the latter was present in glands distributed restrictedly to the uppermost part of the small intestine at a point about 10 cm caudal to the pylorus. The cytoplasm of the mucous cell contain...
Regulation of ovarian function by catecholestrogens: current concepts. Development of the ovarian follicle(s) destined for ovulation appears to be a process in which antral follicles undergo a recruitment, selection and subsequent dominance phase. Several intraovarian or autocrine/paracrine regulatory mechanisms have been evoked to explain these processes. One of these potential autocrine/paracrine regulators is a catecholestrogen, 2-hydroxy-estradiol (2-OH-E2). Evidence implicating 2-OH-E2 as an autocrine/paracrine regulator of follicular function is reviewed. Studies have shown 2-OH-E2 to be present in nanomolar concentrations in fluid of human and equine folli...
Mössbauer spectroscopic study of the initial stages of iron-core formation in horse spleen apoferritin: evidence for both isolated Fe(III) atoms and oxo-bridged Fe(III) dimers as early intermediates. Ferritin stores iron within a hollow protein shell as a polynuclear Fe(III) hydrous oxide core. Although iron uptake into ferritin has been studied previously, the early stages in the creation of the core need to be clarified. These are dealt with in this paper by using Mössbauer spectroscopy, a technique that enables several types of Fe(II) and Fe(III) to be distinguished. Systematic Mössbauer studies were performed on samples prepared by adding 57Fe(II) atoms to apoferritin as a function of pH (5.6-7.0), n [the number of Fe/molecule (4-480)], and tf (the time the samples were held at room ...
Serum angiotensin converting enzyme activity and response to angiotensin I in horses. The activity of serum angiotensin converting enzyme (ACE) for healthy horses was 64 +/- 13 mUnits/ml. In vitro, equine serum ACE was sensitive to the following inhibitors (IC50): enalapril (570 nM or 215 ng/ml), captopril (190 nM or 41.3 ng/ml), and enalaprilat (6 nM or 2.1 ng/ml). The intravenous (i.v.) administration of angiotensin I to six healthy horses produced a dose proportional pressor response. The maximal increase in mean arterial pressure over baseline values was 65.6 mmHg at angiotensin I doses of 500 ng/kg bodyweight (bwt). The attenuation of this response to angiotensin I was fur...
The sequence-specific assignment of the 1H-NMR spectrum of an enzyme, horse-muscle acylphosphatase. A complete range of two-dimensional NMR experiments was used for the assignment of the 1H-NMR spectrum of horse muscle acylphosphatase. Firstly the spin systems of some easily identifiable amino acid side chains were assigned. These side chains involved all the aromatic residues and all the leucine, valine, isoleucine, threonine, alanine, proline as well as some of the glycine residues. Analysis of nuclear Overhauser enhancement spectra in our previous work had identified the sequential and long-range patterns characteristics for secondary structure elements. This result had also provided the ...
Identification and description of beta-structure in horse muscle acylphosphatase by nuclear magnetic resonance spectroscopy. Nuclear magnetic resonance spectra of acylphosphatase were searched for signs of beta-structure, i.e. characteristic nuclear Overhauser enhancement patterns displayed in the two-dimensional spectra, typical chemical shifts, coupling constants and slow 2H-H exchange. The results provided identification of the main-chain resonances of amino acid residues involved in the beta-structure. The full sequential assignment of this region was gained by identification of some amino acid spin systems and their alignment with the primary sequence. The assignment of the side-chains was virtually completed s...
Androgen synthesis and aromatization by equine corpus luteum microsomes. Whereas mare corpus luteum does not produce androgens or estrogens in vivo, the incubation of mare corpus luteum microsomes with progesterone and NADPH resulted in 17 alpha-hydroxyprogesterone and estrogen production with a small yield of androstenedione. In the presence of an aromatase inhibitor (4-hydroxyandrostenedione), 17 alpha-hydroxyprogesterone and androstenedione were accumulated. Aromatization of testosterone and androstenedione occurred via stereospecific loss of the 1 beta, 2 beta hydrogen atoms and was inhibited by MgCl2, KCl, and EDTA. The Km of estrogen synthetase from equine co...
Variations in serum sorbitol dehydrogenase, aspartate transaminase, and isoenzyme 5 of lactate dehydrogenase activities in horses given carbon tetrachloride. Seven horses were given 0.5 mg of carbon tetrachloride/kg of body weight via a nasogastric tube. Subsequent hepatocellular damage was monitored by serum enzyme determinations of sorbitol dehydrogenase, isoenzyme 5 of lactate dehydrogenase, and aspartate transaminase activities. Creatinine kinase activity was evaluated as an indicator of muscle cell damage. Sorbitol dehydrogenase, isoenzyme 5 of lactate dehydrogenase, and aspartate transaminase activities were significantly (P less than 0.05) increased by 24 hours after carbon tetrachloride administration. Isoenzyme 5 of lactate dehydrogenase a...
Observations on gamma-glutamyl transferase, 5′-nucleotidase and leucine aminopeptidase activities in the plasma of the horse. In 18 horses there was no effect of age or sex on plasma activities of gamma-glutamyl transferase (gamma-GT), 5'-nucleotidase (5'-NT) and leucine aminopeptidase (LAP). All the enzymes were equally stable after storage for one month at -20 degrees C and there was no significant difference between their activities in serum and plasma in clinically normal horses. The pattern of release of gamma-GT, 5'-NT and LAP into plasma was studied in 114 horses which had a variety of orthopaedic, gastrointestinal, cardiovascular and hepatic (necrosis, lipidosis, neoplasia and cirrhosis) conditions. A definit...
The preparation and biochemical characterization of intact capsids of equine infectious anemia virus. Capsids of equine infectious anemia virus have been isolated as cone-shaped particles 60 x 120 nm in size. Detergent treatment of whole virus followed by two cycles of rate-zonal centrifugation in Ficoll produces these capsids in a yield of approximately 10%. The major protein components are the gag-encoded p11 nucleocapsid protein and p26 capsid protein, which are present in equimolar amounts. Substantial cleavage of p11 to p6 and p4 can be observed under conditions where the viral protease packaged in the capsid is enzymatically active.
Nonequivalence of the two subunits of horse erythrocyte glutathione transferase in their reaction with sulfhydryl reagents. Glutathione transferase (EC 2.5.1.18) from horse erythrocytes has been purified and some molecular and kinetic properties have been investigated. It appears to be a dimeric protein composed of subunits of about 23 kDa, indistinguishable either in sodium dodecyl sulfate or in urea electrophoresis. Amino acid composition, substrate specificities, sensitivity to inhibitors, CD spectra, and immunological studies provide evidence that the horse enzyme is related to the pi class transferases. This enzyme has only two reactive thiol groups/dimer whose integrity appears to be essential for the activit...
Synthesis and aromatization of 19-norandrogens in the stallion testis. The results of the measurement of 19-nortestosterone in the testiscular artery and vein of the stallion, the very low levels of this steroid in the peripheral blood of geldings and the similar patterns of increase in the peripheral levels of 19-nortestosterone and testosterone after hCG stimulation, show that 19-nortestosterone, like testosterone, is essentially synthesized in the testis. This testicular origin was confirmed by the ability of testicular tissue to synthesize 19-norandrogens from [4-14C]androgens in vitro. 19-Nortestosterone was 50% conjugated in the peripheral blood and almost ...
Induction of equine postanesthetic myositis after halothane-induced hypotension. Wick catheters were used to measure intracompartmental pressures of the extensor carpi radialis muscles and long heads of the triceps brachii muscles of 7 horses maintained under halothane anesthesia during controlled ventilation. Horses were positioned in left lateral recumbency on a water bed for 4 hours. Using a crossover design, 6 of the 7 horses were subjected to normotensive and hypotensive anesthesia on separate occasions. Hypotension was achieved by increasing the inspired halothane concentration. Hematologic and biochemical measurements were determined at designated intervals before, ...
Analysis of N-acetyl-4-O-acetylneuraminic-acid-containing N-linked carbohydrate chains released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. Application to the structure determination of the carbohydrate chains of equine fibrinogen. The carbohydrate chains of equine fibrinogen were enzymatically released by peptide-N4-(N-acetyl-beta-glucosaminyl)asparagine amidase F. The oligosaccharides obtained were fractionated by a combination of FPLC and HPLC and analyzed by 500-MHz 1H-NMR spectroscopy. Four monosialo and four disialo diantennary N-acetyllactosamine type of carbohydrate chains occur: (formula; see text)
A monoclonal antibody against horse kidney (Na+ + K+)-ATPase inhibits sodium pump and E2K to E1 conversion of (Na+ + K+)-ATPase from outside of the cell membrane. Monoclonal antibodies against horse kidney outer medulla (Na+ + K+)-ATPase were prepared. One of these antibodies (M45-80), was identified as an IgM, recognized the alpha subunit of the enzyme. M45-80 had the following effects on horse kidney (Na+ + K+)-ATPase: (1) it inhibited the enzyme activity by 50% in 140 mM Na+ and by 80% in 8.3 mM Na+; (2) it increased the Na+ concentration necessary for half-maximal activation (K0.5 for Na+) from 12.0 to 57.6 mM, but did not affect K0.5 for K+; (3) it slightly increased the K+-dependent p-nitrophenylphosphatase (K-pNPPase) activity; (4) it inhibited p...
Studies on serum selenium and tocopherol in white muscle disease of foal. In order to clarify the cause of white muscle disease (W.M.D.) in foals, tocopherol and selenium concentrations in serum and glutathione peroxidase activities in blood were measured. Examination was made on the samples from horses affected with W.M.D., the foal kept with them in the same stable, the foals kept in the stables without affected foals, and respective mares. The heavy-breed horses in Fukuoka prefecture and Tokachi district were also examined for comparison. Serum tocopherol levels of these foals were normal because after intake of colostrum. Mares of affected foals showed lower toc...
Steroids in equine testes: the identification of endogenous 19-hydroxy and 19-nor neutral steroids by gas chromatography–mass spectrometry. After homogenization of testicular tissue from stallions aged 1, 2 and 5 years, the unconjugated and conjugated steroids were isolated by a combined solvent-solid extraction procedure. The conjugates were further separated into glucuronides and sulphates by chromatography using Sephadex LH-20. After enzyme hydrolysis and solvolysis of the respective conjugate classes, the three extracts, unconjugated steroids, aglycones and solvolysed sulphates, were purified by chromatography using Kieselgel 60H columns. Five fractions were resolved from each extract; an aliquot of each fraction was derivatiz...
Crystallization and preliminary X-ray study of horse pancreatic lipase. Horse (Equus caballus) pancreatic lipase (EC 3.1.1.3) has been crystallized using the hanging drop method of vapour diffusion at 20 degrees C. The best crystals were grown from an 8 mg/ml solution in 10 to 20% (w/v) polyethylene glycol 8000, 10 mM-MgCl2, 0.1 M-NaCl, 0.1 M-Mes buffer (pH 5.6). They reach dimensions of 0.8 mm x 0.4 mm x 0.6 mm. X-ray examination of the lipase crystals shows that they are orthorombic with a space group P2(1)2(1)2(1). Their cell dimensions are a = 79.8 A, b = 97.2 A c = 145.3 A. Two molecules per asymmetric unit give a Vm value of 2.82 A3/dalton (56% water content...
The structure and properties of horse muscle acylphosphatase in solution. Mobility of antigenic and active site regions. The solution structure of acylphosphatase determined by proton nuclear magnetic resonance spectroscopy is described. The results allow us to discuss the fold of the protein (101 amino acids), to correlate the exposure and the mobility of the backbone with the antigenicity, and to locate the active site.
Immunocytochemical localization of carbonic anhydrase isozyme III in equine thymus. Sections of equine thymus were examined for the presence of carbonic anhydrase (CA) isozymes by an immunohistochemical method. Carbonic anhydrase III, a major enzyme of skeletal muscle, was localized in some of the epithelial-reticular cells of the equine thymus. This finding suggests the presence of a new type of cell in the thymic cortex. The concentration of CA-III in the thymus was 17 micrograms/g wet tissue. CA-I and CA-II were not found in equine thymus.
