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Wilson, M. T., Ranson, R. J., Masiakowski, P., Czarnecka, E., & Brunori, M. (1977). A kinetic study of the pH-dependent properties of the ferric undecapeptide of cytochrome c (microperoxidase). Eur J Biochem, 77(1), 193–199.
Abstract: The ferric form of the haem undecapeptide, derived from horse cytochrome c by peptic digestion, undergoes at least three pH-induced transitions with pK values of 3.4, 5.8 and 7.6. Temperature-jump experiments suggest that the first of these is due to the binding of a deprotonated imidazole group to the feric iron while the second and third arise from the binding of the two available amino groups present (the alpha-NH2 of valine and the epsilon-NH2 of lysine). Molecular models indicate that steric retraints on the peptide dictate that these amino groups may only coordinate to iron atoms via intermolecular bonds, thus leading to the polymerization of the peptide. Cyanide binding studies are in agreement with these conclusions and also yield a value of 3.6 X 10(6) M-1 s-1 for the intrinsic combination constant of CN- anion with the haem. A model is proposed which describes the pH-dependent properties of the ferric undecapeptide.
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Yarnell, K., Hall, C., & Billett, E. (2013). An assessment of the aversive nature of an animal management procedure (clipping) using behavioral and physiological measures. Physiol. Behav., 118, 32–39.
Abstract: Animal management often involves procedures that, while unlikely to cause physical pain, still cause aversive responses. The domestic horse (Equus caballus) regularly has excessive hair clipped off to facilitate its use as a riding/driving animal and this procedure causes adverse behavioral responses in some animals. The aim of this study was to compare behavioral and physiological measures to assess the aversive effect of this procedure. Ten horses were selected on the basis of being either compliant (C: n=5) or non-compliant (NC: n=5) during this procedure. The horses were subjected to a sham clipping procedure (SC: where the blades had been removed from the clippers) for a period of ten minutes. Measures were taken pre, during and post SC (-10min to +30min) and mean values calculated for ALL horses and for C and NC separately. Behavioral activity was scored (scale 1-5) by twenty students from video footage in (phase/group-blind scoring). Heart rate (HR), salivary cortisol and eye temperature were monitored throughout the procedure. The NC horses were found to be significantly more behaviorally active/less relaxed throughout the trial than C horses (p<0.05) with the greatest difference occurring during the SC procedure (p<0.01). NC horses were more active/less relaxed during, compared with pre or post SC (p<0.05), but showed no behavioral difference pre and post SC. HR of the NC horses was higher than that of the C horses throughout the trial but only significantly so after 10min of SC (p<0.01). ALL horses showed a significant increase in HR between +5 and +10min into the procedure (p<0.05). There was a significant increase in salivary cortisol concentration in ALL horses post procedure (p<0.01) with levels peaking at 20minute post SC. No significant differences in salivary cortisol concentration between C and NC were found at any stage of the trial. Eye temperature increased significantly in ALL horses during SC, peaking at +10min into the procedure (p<0.05) and then decreased substantially when SC had ceased (p<0.01). Although no significant differences were found between C and NC per se, there was a significant interaction between group and phase of trial (p<0.05) with the NC group showing a greater decrease in eye temperature post SC. There was a significant positive correlation between changes in salivary cortisol concentration and eye temperature (p<0.01) but no correlation between any of the other measures. Although the behavioral response of C and NC to this procedure was significantly different the physiological responses indicated that ALL horses found the procedure aversive. Eye temperature could be used as an objective and immediate measure of how an animal is responding to a specific situation in order to evaluate management procedures and adapt them where appropriate to reduce the negative impact on animal health and welfare.
Keywords: Thermography; Eye temperature; Cortisol; Horse; Welfare; Clipping
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Shaw, E. B., Houpt, K. A., & Holmes, D. F. (1988). Body temperature and behaviour of mares during the last two weeks of pregnancy. Equine Vet J, 20(3), 199–202.
Abstract: Average daily core body temperature and behavioural patterns of pregnant mares were studied, in search of definitive signs of parturition within 24 h of the event. Nineteen pony mares were sampled twice daily for core body temperature. A significant temperature drop, averaging 0.1 degrees C (0.2 degrees F) was observed during the day prior to parturition. Between 18.00 h and 06.00 h, during the two weeks before parturition, Thoroughbred and Standardbred mares (n = 52) spent an average 66.8 per cent of their time standing, 27.0 per cent eating, 4.9 per cent lying in sternal recumbency, 1.0 per cent lying in lateral recumbency, and 0.3 per cent walking. On the night before parturition, mares spent significantly less time lying in sternal recumbency than on previous nights and on the night of parturition all behaviour patterns except eating were significantly different from the nights of the two weeks before parturition. There was an increase in walking (5.3 per cent), lying in sternal recumbency (8 per cent) and lying in lateral recumbency (5.3 per cent) whereas standing (53.3 per cent) was decreased. In 58 observed pregnancies, 54 mares (97 per cent) foaled in a recumbent position and 50 mares (86 per cent) foaled between 18.00 h and 06.00 h.
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Permyakov, S. E., Khokhlova, T. I., Nazipova, A. A., Zhadan, A. P., Morozova-Roche, L. A., & Permyakov, E. A. (2006). Calcium-binding and temperature induced transitions in equine lysozyme: new insights from the pCa-temperature “phase diagrams”. Proteins, 65(4), 984–998.
Abstract: The most universal approach to the studies of metal binding properties of single-site metal binding proteins, i.e., construction of a “phase diagram” in coordinates of free metal ion concentration-temperature, has been applied to equine lysozyme (EQL). EQL has one relatively strong calcium binding site and shows two thermal transitions, but only one of them is Ca(2+)-dependent. It has been found that the Ca(2+)-dependent behavior of the low temperature thermal transition (I) of EQL can be adequately described based upon the simplest four-states scheme of metal- and temperature-induced structural changes in a protein. All thermodynamic parameters of this scheme were determined experimentally and used for construction of the EQL phase diagram in the pCa-temperature space. Comparison of the phase diagram with that for alpha-lactalbumin (alpha-LA), a close homologue of lysozyme, allows visualization of the differences in thermodynamic behavior of the two proteins. The thermal stability of apo-EQL (transition I) closely resembles that for apo-alpha-LA (mid-temperature 25 degrees C), while the thermal stabilities of their Ca(2+)-bound forms are almost indistinguishable. The native state of EQL has three orders of magnitude lower affinity for Ca(2+) in comparison with alpha-LA while its thermally unfolded state (after the I transition) has about one order lower (K = 15M(-1)) affinity for calcium. Circular dichroism studies of the apo-lysozyme state after the first thermal transition show that it shares common features with the molten globule state of alpha-LA.
Keywords: Animals; Apoproteins/chemistry/metabolism; Binding Sites; Calcium/chemistry/*metabolism; Cattle; Edetic Acid/metabolism; Horses/metabolism; Hydrogen-Ion Concentration; Lactalbumin/chemistry/metabolism; Muramidase/*chemistry/*metabolism; Protein Denaturation; Spectrometry, Fluorescence; *Temperature; Thermodynamics; Tryptophan/chemistry/metabolism
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Czerlinski, G. H., Erickson, J. O., & Theorell, H. (1979). Chemical relaxation studies on the horse liver alcohol dehydrogenase system. Physiol Chem Phys, 11(6), 537–569.
Abstract: Chemical relaxation studies on the system horse liver alcohol dehydrogenase, nicotinamide adenine dinucleotide, and ethanol were conducted observing fluorescence changes between 400 and 500 nm. Temperature-jump experiments were performed at pH 6.5, 7.0, 8.0, and 9.0; concentration-jump experiments at pH 9.0. The reciprocal of the slowest relaxation time was found to be linearly dependent upon the enzyme concentration for relatively low enzyme concentrations, as predicted earlier. Use of the wide pH-range necessitated expression of the four apparent dissociation constants of the catalytic reaction cycle in terms of pH-independent constants. The system was described in terms of only one (or two) catalysis-linked protons not associated with the electron transfer. Protonic steps in a buffered system are in rapid equilibrium, too fast to be measured with the equipment available. Assuming only two of the four bimolecular reaction steps in the four-step cycle are fast compared to the remaining two, six cases may be considered with six expressions for the reciprocal of the slowest relaxation time. Comparison with the experimental data revealed that the bimolecular reaction steps governing the slowest relaxation time change with pH. Above the effective time resolution of the temperature-lump instrument with fluorescence detection (0.1 msec) only one other relaxation time was detectable and only at pH 9. This relaxation time, found to be independent of the concentration of all reactants within experimental error (r = 10 +/- 5 msec), is most likely due to an interconversion among ternary complexes.
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Kihara, H. (1981). Comparison of the redox reactions of various types of cytochrome c with iron hexacyanides. Biochimica et Biophysica Acta (BBA) – Bioenergetics, 634, 93–104.
Abstract: The dynamic behavior of various types of cytochromes c in the redox reaction with iron hexacyanides was studied using a temperature-jump method in order to elucidate the molecular mechanism of the redox reaction of cytochromes with their oxidoreductants. Transmittance after the temperature jump changed through a single exponential decay for all cytochromes investigated. Under a constant concentration of anion, the redox reaction of various types of cytochrome c with iron hexacyanides was analyzed according to the scheme: Ki=kt/k-i (i=1,2,3) where C(III) and C(II) are ferric and ferrous cytochromes, respectively, Fe(III) and Fe(II) are ferri- and ferrocyanides, respectively, C(III) [middle dot] Fe(II) is the ferricytochrome-ferrocyanide complex and C(II) [middle dot] Fe(III) is the ferrocytochrome-ferricyanide complex. When step B is slower than the other two steps A and C, τ-1 can be represented approximately as where the bar over the variables denotes the equilibrium value. In a large excess of ferrocyanide against cytochrome, we can estimate k2, k-2, K1 and K3 independently. In the case of horse cytochrome c at 18[degree sign]C in 0.1 M phosphate buffer at pH 7 with 0.3 M KNO3, the estimated parameters are k2 = 100 +/- 50 s-1, k-2 = (3.5 +/- 1.0) [middle dot] 103 s-1, K1 = 15 +/- 7 M-1 and K3 = (8.5 +/- 1.5) [middle dot] 10-4 M. From the same experiments for seven cytochromes (cytochrome c from horse, tuna, Candida krusei, Saccharomyces oviformis, Rhodospirillum rubrum cytochrome c2, Spirulina platensis cytochrome c-554 and Thermus thermophilus cytochrome c-552), the following results can be deduced. (1) Each parameter defined in the scheme above (k2, k-2, K1, K3) diverged beyond the error range. Above all, k2 values of cytochromes c-554 and c-552 are as large as 1 [middle dot] 104 s-1 and much larger than those for the other cytochromes (to 50 approx. 700 S-1). (2) The variance of k2K1 and k-2/K3 are relatively less than the variances of individual parameters (k2, k-2, K1 and K3), which suggests that the values of k2K1 and k-2/K3 have been conserved during the course of evolution.
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Steinhoff, H. J., Lieutenant, K., & Redhardt, A. (1989). Conformational transition of aquomethemoglobin: intramolecular histidine E7 binding reaction to the heme iron in the temperature range between 220 K and 295 K as seen by EPR and temperature-jump measurements. Biochim Biophys Acta, 996(1-2), 49–56.
Abstract: Temperature-dependent EPR and temperature-jump measurements have been carried out, in order to examine the high-spin to low-spin transition of aquomethemogobin (pH 6.0). Relaxation rates and equilibrium constants could be determined as a function of temperature. As a reaction mechanism for the high-spin to low-spin transition, the binding of N epsilon of His E7 to the heme iron had been proposed; the same mechanism had been suggested for the ms-effect, found in temperature-jump experiments on aquomethemoglobin. A comparison of the thermodynamic quantities, deduced form the measurements in this paper, gives evidence that indeed the same reaction is investigated in both cases. Our results and most of the findings of earlier studies on the spin-state transitions of aquomethemoglobin, using susceptibility, optical, or EPR measurements, can be explained by the transition of methemoglobin with H2O as ligand (with high-spin state at all temperatures) and methemoglobin with ligand N epsilon of His E7 (with a low-spin ground state). Thermal fluctuations of large amplitude have to be postulated for the reaction to take place, so this reaction may be understood as a probe for the study of protein dynamics.
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Ballew, R. M., Sabelko, J., & Gruebele, M. (1996). Direct observation of fast protein folding: the initial collapse of apomyoglobin. Proc. Natl. Acad. Sci. U.S.A., 93(12), 5759–5764.
Abstract: The rapid refolding dynamics of apomyoglobin are followed by a new temperature-jump fluorescence technique on a 15-ns to 0.5-ms time scale in vitro. The apparatus measures the protein-folding history in a single sweep in standard aqueous buffers. The earliest steps during folding to a compact state are observed and are complete in under 20 micros. Experiments on mutants and consideration of steady-state CD and fluorescence spectra indicate that the observed microsecond phase monitors assembly of an A x (H x G) helix subunit. Measurements at different viscosities indicate diffusive behavior even at low viscosities, in agreement with motions of a solvent-exposed protein during the initial collapse.
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Fazio, E., Medica, P., Cravana, C., Giacoppo, E., & Ferlazzo, A. (2008). Effect of Short-Distance Road Transport on Thyroid Function, Rectal Temperature, Body Weight and Heart Rate of Stallions. In IESM 2008.
Abstract: Aim of study was to investigate the effects of transport stress on thyroid response, body weight, rectal temperature and heart rate changes in one hundred twenty-six healthy stallions in basal conditions, before and after short road transport. One hundred twenty-six Thoroughbreds and crossbreds stallions with previous travelling experience, aged 4 to 15 yr, were transported by road in a commercial trailer for a period of 3 h (distance <300 km). Blood samples and physiological parameters were collected at 0800 (basal I) and at 1100 (basal II), in each horse“s box, one week before the loading and transport in basal conditions, and one week later, at 0800 immediately before loading (pre-transport), and after 3 h period of transport and unloading, on their arrival at the breeding stations (post-transport), in each new horse”s box, within 30 min. Increases in circulating T3, T4 and fT4 levels (P < 0.01), but not for fT3 levels, were observed after transport, as compared to before loading values, irrespective of different breed. Lower T4 and fT4 levels were observed in basal II (P < 0.01) than basal I and before loading values (pre-transport). After transport Thoroughbreds showed higher fT3 (P < 0.05) and fT4 (P < 0.01) levels than crossbred stallions. No significant differences for T3 and T4 changes were observed. A significant increase in rectal temperature (P < 0.01) and heart rate (P < 0.05) was observed after transport, as compared to before loading values (pre-transport). No differences between basal I, basal II and before loading values (pre-transport) for physiological parameters were observed.
The highest T3, T4 and fT4 levels recorded after short transport seem to suggest a preferential release from the thyroid gland. The results indicate that short road transport stress contributes significantly to thyroid hormone changes, according to different breed, and to the increase in rectal temperature and heart rate. No differences related to different age were observed. |
Natalini, C. C., & Robinson, E. P. (2003). Effects of epidural opioid analgesics on heart rate, arterial blood pressure, respiratory rate, body temperature, and behavior in horses. Vet Ther, 4(4), 364–375.
Abstract: Heart rate, arterial blood pressures, respiratory rate, body temperature, and central nervous system excitement were compared before and after epidural administration of morphine (0.1 mg/kg), butorphanol (0.08 mg/kg), alfentanil (0.02 mg/kg), tramadol (1.0 mg/kg), the k-opioid agonist U50488H (0.08 mg/kg), or sterile water using an incomplete Latin square crossover design in five conscious adult horses. Treatments were administered into the first intercoccygeal epidural space. Significant (P <.05) reductions in respiratory rate were detected after epidural administration of morphine, alfentanil, U50488H, and sterile water. Additionally, significant (P <.05) head ptosis was observed within the first hour after administration of morphine, U50488H, and tramadol, but neither of these changes appeared to be of clinical significance. No treatment-related changes in motor activity or behavior were observed.
Keywords: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer/administration & dosage/pharmacology; Alfentanil/administration & dosage/pharmacology; Analgesics, Opioid/administration & dosage/*pharmacology; Anesthesia, Epidural/*veterinary; Animals; Behavior, Animal/drug effects; Blood Pressure/drug effects; Body Temperature/drug effects; Butorphanol/administration & dosage/pharmacology; Cross-Over Studies; Female; Heart Rate/drug effects; Horses/*physiology; Injections, Epidural/veterinary; Male; Morphine/administration & dosage/pharmacology; Respiration/drug effects; Tramadol/administration & dosage/pharmacology
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