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Biomedical subjects

N C Price

Publications and source records attributed to N C Price.

At least 127 records · Page 7Linked to original sources
PubMed1986

The susceptibility towards proteolysis of intermediates during the renaturation of yeast phosphoglycerate mutase.

The renaturation of the tetrameric enzyme phosphoglycerate mutase from baker's yeast after denaturation in guanidinium chloride was studied. Three proteinases (trypsin, chymotrypsin and thermolysin) cause extensive loss of activity of samples taken during the early stages of refolding. As judged by SDS/polyacrylamide-gel electrophoresis, the proteinases cause substantial degradation of the polypeptide chain with no evidence for large quantities of fragments of Mr greater than 6500. These data suggest that the early intermediates in the refolding, especially the folded monomer, possess a number of sites that are susceptible to proteolysis.

Binding Sites↗
PubMed1986

The in vitro metabolism of Eulan WA New by liver homogenates from freshwater fish.

The rate of formation of pentachloro-2-aminodiphenylether (5-PAD) from pentachloro-2-(chloromethylsulphonamido) diphenylether (6-PCSD) by goldfish liver homogenates was studied in a variety of buffers. The highest rate was observed when the homogenate was prepared and assayed in phosphate buffer containing dithiothreitol. The 5-PAD forming activity of liver homogenate preparations from a variety of freshwater fish (goldfish, carp, pike, rainbow trout, perch and eel) showed contrasting species differences. These results could be correlated with information available on the toxicity of the mothproofing agent Eulan WA New (which contains PCSDs) to these fish.

Animals↗
PubMed1986

The role of vitrectomy following accidental intraocular injection of deposteroid preparations.

Four cases of accidental intraocular injection of deposteroid preparation treated by early vitrectomy are reported. Previous reports indicate that with conservative management loss of vision is due to retinal detachment or optic neuropathy. Early vitrectomy is advocated to allow identification and treatment of retinal breaks to prevent retinal detachment and the removal of potentially retino-toxic preservatives and carriers contained in the steroid preparations. Immediate paracentesis may prevent ischaemic optic neuropathy.

Adult↗
PubMed1985

Inactivation of rabbit muscle phosphoglycerate mutase by limited proteolysis with thermolysin.

Rabbit muscle phosphoglycerate mutase is inactivated by proteolysis with thermolysin. Inactivation is correlated with the breakage of one (or a few) bond(s) near one end of the polypeptide chain. There is no change in the overall conformation, quaternary structure or binding to Cibacron Blue on proteolysis. The possible analogy with the existence of a flexible tail in the yeast enzyme is discussed.

Animals↗
PubMed1985

Evidence for active intermediates during the reconstitution of yeast phosphoglycerate mutase.

The reconstitution of the tetrameric phosphoglycerate mutase from bakers' yeast after denaturation in guanidine hydrochloride has been studied. When assays are performed in the presence of trypsin, it is found that reactivation parallels the regain of tetrameric structure. However, in the absence of trypsin, the regain of activity is more rapid, suggesting that monomeric and dimeric intermediates possess partial activity (35% of the value of native enzyme) which is sensitive to trypsin. When reconstitution is studied in the presence of substrates, it is again found that monomeric and dimeric intermediates possess 35% activity. Under these latter conditions, the activity of the monomer but not of the dimer is sensitive to trypsin.

Bisphosphoglycerate Mutase↗
PubMed1985

Purification and preliminary characterization of phosphoglycerate mutase from Schizosaccharomyces pombe.

Phosphoglycerate mutase could be purified to over 95% homogeneity by a single step procedure involving elution from Cibacron Blue-Sepharose by a pulse of cofactor 2,3-bisphosphoglycerate. Although the enzyme has been isolated in only small quantities (c. 100 micrograms), gel filtration and sodium dodecylsulphate polyacrylamide gel electrophoresis indicated that it is monomeric with Mr approximately 23,000, an extremely low value for this enzyme. Preliminary investigations of the kinetic characteristics and the nature of important amino acid side chains have been undertaken.

Diacetyl↗
PubMed1984

The in vitro metabolism by homogenates of goldfish (Carassius auratus) liver of pentachloro-2-(chloromethyl sulphonamido) diphenyl ether, the major component and active ingredient of the mothproofing agent Eulan WA New.

After dosing goldfish (Carassius auratus) for 2 hr with pentachloro-2-(chloromethyl sulphonamido) diphenyl ether, 6-PCSD, (the major component of the active ingredients of the mothproofing agent Eulan WA New) in the surrounding water, high concentrations of the corresponding amine pentachloro-2-aminodiphenyl ether, 5-PAD, were found in the liver. This suggests that the liver is a primary site of 5-PAD formation in vivo. Homogenates of goldfish liver showed a 5-PAD forming activity when incubated with 6-PCSD; this activity was destroyed by boiling the homogenate or by treatment with trypsin. The activity was greater at pH 7.4 than at pH 3, 5 or 9. There was no detectable corresponding activity in homogenates of goldfish spleen or digestive tissue or in the digestive tract contents.

Animals↗
PubMed1983

The reconstitution of denatured phosphoglycerate mutase.

The reconstitution of the tetrameric enzyme yeast phosphoglycerate mutase after denaturation in guanidine hydrochloride has been studied. Denaturation is almost completely reversible at enzyme concentrations greater than 10 micrograms/ml. Cross-linking by glutaraldehyde has been used to monitor the reassociation of the subunits; the kinetics of this process has been analyzed in terms of a model involving an equilibrium between monomer and dimer followed by a bimolecular association of two dimers to give a tetramer. Reactivation is found to parallel the appearance of tetramer. Structural changes during reconstitution have been measured by circular dichroism and fluorescence. Both methods reveal complex kinetics indicating the rapid formation of structured monomers (half-time less than 10 s), followed by slow subunit association. For comparison, preliminary reconstitution experiments were performed on the dimeric phosphoglycerate mutase from rabbit muscle.

Animals↗
PubMed1983

The denaturation of rabbit muscle phosphorylase b by guanidinium chloride.

The denaturation of phosphorylase b by guanidinium chloride (GdnHCl) was studied. The enzyme is unusually sensitive to the denaturing agent, being more than 50% inactivated after incubation for 15 min in 0.1 M-GdnHCl. Full activity can be regained on dilution of the GdnHCl to 0.01 M, provided that the initial concentration of GdnHCl is less than 0.5 M. Studies of protein fluorescence, thiol-group reactivity, circular dichroism and absorption spectroscopy indicate that phosphorylase b undergoes slow structural changes in the range of GdnHCl concentrations from 0.5 to 0.8 M. The enzyme retains considerable folded structure even after 15 min incubation in 1 M-GdnHCl, but is rapidly and completely unfolded in 3 M-GdnHCl.

Animals↗
PubMed1983

The refolding of denatured rabbit muscle pyruvate kinase.

The refolding of rabbit muscle pyruvate kinase after denaturation by guanidine hydrochloride was studied. On dilution of the denaturing agent, enzyme activity is only partially regained. The extent of regain of activity is dependent on protein concentration, showing a marked decrease at higher concentrations. The failure to regain complete activity appears to be related to the formation of inactive aggregates, which can be separated from active enzyme by gel filtration. Insoluble aggregates can be partially re-activated after solubilization in guanidine hydrochloride. Changes in the circular-dichroism and fluorescence spectra during refolding suggest that a partially folded, inactive species is formed rapidly; this differs from native enzyme in being more susceptible to proteolysis by trypsin.

Animals↗
PubMed1983

Distinction between cofactor-dependent and -independent phosphoglycerate mutases by chromatography on Cibacron Blue-Sepharose.

The binding of phosphoglycerate mutases from a variety of sources to Cibacron Blue-Sepharose has been examined. Those enzymes which are dependent on 2,3-bisphosphoglycerate (BPG) for activity bind to the immobilized dye and can be eluted by BPG. Those enzymes which are independent of BPG do not bind to the immobilized dye. The possible structural significance of this distinction is discussed.

2,3-Diphosphoglycerate↗
PubMed1982

The refolding of denatured rabbit muscle creatine kinase. Search for intermediates in the refolding process and effect of modification at the reactive thiol group on refolding.

A number of aspects of the refolding of denatured rabbit muscle creatine kinase have been studied. Addition of substrates has no effect on the rate or extent of regain of activity. The changes in protein fluorescence during refolding broadly parallel the regain of activity. A study of the susceptibility of the enzyme to proteolysis during refolding indicates that there is no significant accumulation of folded, but inactive, intermediates in the folding process. Modification of the reactive thiol group on each subunit of the enzyme by small reagents such as iodoacetate or iodoacetamide prior to denaturation has only a small effect on the rate of subsequent refolding. However, modification by the bulky reagent 6-(4-iodoacetamidophenyl)aminonaphthalene-2-sulphonate has a very large effect on the ability of the enzyme to refold after denaturation.

Animals↗
PubMed1981

The effect of limited proteolysis on rabbit muscle creatine kinase.

Creatine kinase from rabbit muscle is inactivated by limited proteolysis with proteinase K from Tritirachium album. Gel-filtration and cross-linking studies showed that the limited proteolysis did not affect the molecular weight of the enzyme under non-denaturing conditions, but did cause changes in the reactivity of the reactive thiol group on each subunit and in the ability of the enzyme to form a 'transition-state analogue' complex in the presence of magnesium acetate plus ADP plus creatinine plus NaNO3.

Adenosine Diphosphate↗