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Biomedical subjects

J F Morrison

Publications and source records attributed to J F Morrison.

At least 73 records · Page 4Linked to original sources
PubMed1991

pH dependency of the reactions catalyzed by chorismate mutase-prephenate dehydrogenase from Escherichia coli.

The variation with pH of the kinetic parameters associated with the mutase and dehydrogenase reactions catalyzed by chorismate mutase-prephenate dehydrogenase has been determined with the aim of elucidating the role that ionizing amino acid residues play in binding and catalysis. The pH dependency of log V for the dehydrogenase reaction shows that the enzyme possesses a single ionizing group with a pK value of 6.5 that must be unprotonated for catalysis. This same group is observed in the V/Kprephenate, as well as in the V/KNAD, profile. The V/Kprephenate profile exhibits a second ionizing residue with a pK value of 8.4 that must be protonated for the binding of prephenate to the enzyme. For the mutase reaction, the V/Kchorismate profile indicates the presence of three ionizing residues at the active site. Two of these residues, with similar pK values of about 7, must be protonated, while the third, with a pK value of 6.3, must be unprotonated. It can be concluded that all three groups are concerned with the binding of chorismate to the enzyme since the maximum velocity of the mutase reaction is essentially independent of pH. This conclusion is confirmed by the finding that the Ki profile for the competitive inhibitor, (3-endo,8-exo)-8-hydroxy-2-oxabicyclo[3.3]non-6-ene-3,5-dicarboxylic acid, shows the same three ionizing groups as observed in the V/Kchorismate profile. By contrast, the Ki profile for carboxyethyldihydrobenzoate shows only one residue, with a pK value of 7.3, that must be protonated for binding of the inhibitor.(ABSTRACT TRUNCATED AT 250 WORDS)

Binding Sites↗
PubMed1991

Kinetic studies on chorismate mutase-prephenate dehydrogenase from Escherichia coli: models for the feedback inhibition of prephenate dehydrogenase by L-tyrosine.

Kinetic studies have been undertaken to elucidate the mechanism of the allosteric inhibition by tyrosine of the prephenate dehydrogenase activity of the bifunctional dimeric enzyme chorismate mutase-prephenate dehydrogenase. The effect of tyrosine on the initial velocity of the reactions in the presence of both prephenate and the alternative substrate, 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate, have been determined. In addition, investigations have been made of the effect of tyrosine on the inhibition of the reaction by the inhibitory analogues of prephenate, (4-hydroxyphenyl)pyruvate, and (carboxyethyl)-1,4-dihydrobenzoate. The results of the double inhibition experiments indicate clearly that the enzyme possesses a distinct allosteric site for the binding of tyrosine. The initial velocity data obtained with both substrates have been fitted to the rate equations that describe a wide range of models. From a comparison of the results obtained from studies with the two substrates, and with a knowledge of the value for the dissociation constant of the tyrosine-enzyme complex, definitive conclusions have been reached about the mechanism of the allosteric inhibition. It is concluded that tyrosine combines twice at allosteric sites and in an antisynergistic fashion, while prephenate reacts at both active sites of the dimeric enzyme as well as weakly at one of the allosteric sites. It appears that the latter is simple competition reaction that affects neither the binding of prephenate at the active site nor the rate of product formation. The model also predicts the formation of an active tyrosine-enzyme-prephenate complex that yields product at a much slower rate than does the enzyme-prephenate complex.(ABSTRACT TRUNCATED AT 250 WORDS)

Allosteric Regulation↗
PubMed1991

Further evidence for the absence of a descending cholinergic projection from the brainstem to the spinal cord in the rat.

Serotonergic and catecholaminergic neurons are known to project from the brainstem to the spinal cord. However, evidence for a bulbo-spinal projection that is cholinergic is sparse despite immunocytochemical and physiological evidence for a cholinergic influence on the cord. In this study we examined the possibility of a direct cholinergic bulbo-spinal projection in the rat using a combination of retrograde axonal tracing techniques and choline acetyltransferase immunocytochemistry. Although many cells were found to project to the cord from the brainstem, none were identified as being cholinergic, confirming previous evidence that the cholinergic innervation of the cord is intrinsic.

Animals↗
PubMed1991

Differential neuropeptide expression after visceral and somatic nerve injury in the cat and rat.

The expression of neuropeptides galanin, vasoactive intestinal polypeptide (VIP) and substance P was compared after injury to somatic (sciatic, pudendal) and visceral (pelvic) nerves. Studies in normal rats and the mutant rat 'mutilated foot' suggested that galanin increases in sensory but not sympathetic fibres after sciatic nerve injury, while VIP appears to increase in both sensory and sympathetic fibres, and substance P to decrease in sensory fibres. A direct comparison of neuropeptide changes after somatic and visceral nerve injury was made in the cat dorsal sacral spinal cord, where both pudenal (somatic) and pelvic (visceral) afferents terminate. Four weeks after pudendal nerve transection in the cat there was an increase of VIP and galanin but decrease of substance P in the dorsal sacral cord, similar to the changes in lumbar dorsal cord after sciatic nerve section in the rat. In contrast, 4 weeks after pelvic nerve transection in the cat, galanin was unchanged in the ipsilateral dorsal sacral spinal cord, whereas VIP is known to decrease markedly and substance P to remain unchanged. There is thus differential peptide expression before and after injury in somatic and visceral systems, which may be regulated in part by the target organ. We have proposed that the neuropeptide changes occur in neurons that regulate development, maintenance and repair after injury, processes that may differ in somatic and visceral systems.

Animals↗
PubMed1991

Characterization of tightly bound substrates in pure preparations of dihydrofolate reductase: implications for studies on enzymes.

Investigations have been made to determine the identity and binding characteristics of the pterins that are bound tightly to dihydrofolate reductases which are isolated from vertebrate sources by a well established procedure. This procedure involves the binding of enzyme to a Sepharose-methotrexate column, elution with dihydrofolate and removal of free dihydrofolate by dialysis or by size-exclusion chromatography. Addition to such preparations of NADPH results in oxidation of the nucleotide and from the progress curves so obtained, it is possible to identify the bound pterin and to calculate the stoichiometry of binding. The data indicated that stoichiometric amounts of dihydrofolate or folate plus dihydrofolate were bound to the dihydrofolate reductases. It can also be concluded that binding occurs at the active sites of the enzymes. Enzyme preparations from which bound pterin had been removed by isoelectric focussing reacted with folate or dihydrofolate to form 1:1 enzyme-pterin complexes from which the pterin could not be removed by dialysis or by size-exclusion chromatography. From the magnitude of the dissociation constants for the enzyme-pterin complexes and the concentration of enzyme present in fractions after the step involving affinity chromatography on Sepharose-methotrexate, it could be concluded that the presence of bound folate and/or dihydrofolate in pure preparations of dihydrofolate reductase is simply a consequence of an association-dissociation reaction. Preparations of the enzyme from bacterial sources were also found to contain bound pterins. The findings have implications with respect to kinetic and thermodynamic studies on dihydrofolate reductase and other enzymes which are isolated by affinity chromatography techniques.

Chromatography↗
PubMed1991

Reversibility tests in chronic obstructive airways disease: their predictive value with reference to benefit from domiciliary nebuliser therapy.

The role of short-term tests of reversibility in selecting patients with COAD for long-term nebuliser therapy is uncertain. In a double-blind placebo-controlled crossover study we have examined the correlation between short-term reversibility and response to a home nebuliser. We studied 20 patients with severe COAD (mean age 66, mean FEV1 0.81 l) and little reversibility (less than 20% increase in FEV1 post-inhaled salbutamol 200 micrograms and less than 25% increase in peak expiratory flow rate, PEFR, on oral steroids). PEFR, spirometry, lung volumes and airways conductance were recorded before and 1 h after a mixture of nebulised ipratropium 0.5 mg and fenoterol 1.25 mg. Patients then recorded twice-daily PEFR at home while they received nebulised ipratropium plus fenoterol, or saline placebo, four times a day for three week blocks using a double-blind cross over protocol. Mean PEFR on home nebuliser rose from 164 l m-1 (placebo) to 196 l m-1 (ipratropium plus fenoterol), paired t-test P = 0.0001. Correlation coefficients between short-term response for PEFR, spirometry and lung volumes, and improvement in home PEFR on nebulised ipratropium plus fenoterol, were all poor (R = -0.37-0.35, P = 0.83-0.11). We conclude that in severe COAD, reversibility tests of PEFR, spirometry and lung volumes do not correlate with response to a home nebuliser. Home measurements of PEFR are probably the best objective method of assessing response to a home nebuliser in such patients.

Aged↗
PubMed1991

The parasympathetic nervous system and the diurnal variation of lung mechanics in asthma.

In a placebo-controlled double-blind experiment the effects of cholinergic blockade with 30 micrograms kg-1 atropine administered by intravenous injection have been studied in nocturnal asthma. Cholinergic blockade at night reversed the changes in indices of airflow in small airways and of gas trapping to the values seen after administration of atropine during the daytime. However, the improvement in indices of airflow in large airways was not complete, reaching only the daytime placebo levels. No changes were seen in any aspect of breathing pattern. The results are discussed in relation to the literature.

Adult↗
PubMed1991

Effects of H1-receptor blockade with terfenadine in nocturnal asthma.

1. In a single-blind placebo controlled study we have measured peak flow (PEFR) at 04.00 h and 16.00 h in eight asthmatics 6 h after placebo or terfenadine 120 mg, to determine if diurnal variation in histamine mediated effects contribute to nocturnal bronchoconstriction in asthma. 2. On placebo there was a significant diurnal variation in mean PEFR of 41 l min-1 (P less than 0.05). Terfenadine improved 04.00 h baseline mean PEFR from 242 to 278 l min-1 (P less than 0.05) but a 38 l min-1 diurnal variation in mean PEFR persisted (P less than 0.05). 3. We conclude that H1-receptor blockade with terfenadine may produce modest nocturnal bronchodilatation but does not influence the diurnal variation in PEFR in asthma suggesting that H1-receptor mediated effects are not important in the pathogenesis of nocturnal asthma.

Adult↗
PubMed1991

Dose response to inhaled salbutamol in chronic obstructive airways disease.

High dose inhaled salbutamol is increasingly used in the management of chronic obstructive airways disease. To determine the range of doses to achieve optimal bronchodilatation and the proportion of patients requiring high dose therapy we have studied 23 patients with chronic obstructive airways disease. Cumulative dose responses were measured to six incremental doses of salbutamol (0.2 to 1.2 mg) delivered by metered dose inhaler. Results were analysed by polynomial regression to calculate the smallest dose required to produce 90% maximal bronchodilatation in each patient. While 5/23 (22%) required greater than 1 mg the majority, 14/23 (61%), achieved 90% maximal bronchodilation with salbutamol 0.6 mg or less. The 8 patients with severe airflow limitation (FEV1 less than or equal to 1 litre) showed a similar pattern of response. We conclude that in chronic obstructive airways disease there are wide individual variations in the dose of inhaled salbutamol producing 90% maximal bronchodilatation with only a minority requiring high dose therapy.

Administration, Inhalation↗
PubMed1991

Platelet activation in nocturnal asthma.

Platelet activation may be a factor in the bronchial hyperresponsiveness that characterises asthma. As hyperresponsiveness is increased at night, changes in platelet activation over 24 hours were related to the diurnal changes in peak expiratory flow and plasma catecholamine concentrations in five subjects with asthma and five normal subjects. The effect of muscarinic receptor blockade with intravenous atropine at 0400 hours on these measurements was also studied. Platelet activation, assessed as the ration of beta thromboglobulin to platelet factor 4, was highest when the peak expiratory flow rate was at its lowest in the asthmatic subjects. There was no correlation between platelet activation and plasma catecholamine concentrations. Intravenous atropine did not alter the ratio of beta thromboglobulin to platelet factor 4, suggesting that parasympathetic activity is not the cause of the increased platelet activation at night.

Adult↗
PubMed1990

Chorismate mutase-prephenate dehydrogenase from Escherichia coli. 1. Kinetic characterization of the dehydrogenase reaction by use of alternative substrates.

The bifunctional enzyme involved in tyrosine biosynthesis, chorismate mutase-prephenate dehydrogenase, has been isolated from extracts of a plasmid-containing strain of Escherichia coli K12 and purified to homogeneity by a modified procedure that involves chromatography on both Matrex Blue A and Sepharose-AMP. Detailed studies of the dehydrogenase reaction have been undertaken with analogues of prephenate that act as substrates. The analogues, which included two of the four possible diastereoisomers of 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate (deoxodihydroprephenate) as well as D- and L-arogenate, were synthesized chemically. As judged by their V/K values, all analogues were poorer substrates than prephenate. The order of their effectiveness as substrates is prephenate greater than one isomer of 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate greater than L-arogenate greater than other isomer of 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate greater than D-arogenate. Thus the dehydrogenase activity is dependent on the degree and position of unsaturation in the ring structure of prephenate as well as on the type of substitution on the pyruvyl side chain. With prephenate as a substrate, the reaction is irreversible because it involves oxidative decarboxylation. By contrast, 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate undergoes only a simple oxidation, and thus, with this substrate, the reaction is reversible. Steady-state velocity data, obtained by varying substrates over a range of higher concentrations, suggest that the dehydrogenase reaction conforms to a rapid equilibrium, random mechanism with 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate as a substrate in the forward reaction or with the corresponding ketone derivative as a substrate in the reverse direction. The initial velocity patterns obtained by varying prephenate or 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate over a range of lower concentrations, at different fixed concentrations of NAD, were nonlinear and consistent with a unique model that is described by a velocity equation which is the ratio of quadratic polynomials. An equilibrium constant of 1.4 x 10(-7) M for the reaction in the presence of 1-carboxy-4-hydroxy-2-cyclohexene-1-propanoate indicates that the equilibrium lies very much in favor of ketone production.

Bacterial Proteins↗
PubMed1990

Chorismate mutase-prephenate dehydrogenase from Escherichia coli. 2. Evidence for two different active sites.

The inhibition of the bifunctional enzyme chorismate mutase-prephenate dehydrogenase by substrate analogues, by the end product, tyrosine, and by the protein modifying agent iodoacetate has been investigated. The purpose of the investigations was to determine if the two reactions catalyzed by the enzyme occur at a single active site or at two separate active sites. Evidence in support of the conclusion that the mutase and dehydrogenase reactions are catalyzed at two similar but distinct active sites comes from the following results: (1) A substrate analogue (endo-oxabicyclic diacid) that inhibits competitively the mutase reaction has no effect on the dehydrogenase reaction. (2) Malonic acid and several of its derivatives act as inhibitory analogues of chorismate in the mutase reaction and of prephenate in the dehydrogenase reaction. However, different dissociation constants for their interaction with the free enzyme are obtained from studies on the mutase and dehydrogenase reactions. (3) The kinetics of the inhibition by tyrosine of the mutase reaction in the presence of NAD differ from those of the dehydrogenase reaction. The results confirm that carboxymethylation with iodoacetate of one cysteine residue per subunit eliminates both mutase and dehydrogenase activities and show that the inactivation of the enzyme activities is due to iodoacetate functioning as an active site directed inhibitor.

Bacterial Proteins↗
PubMed1990

Adrenaline and nocturnal asthma.

OBJECTIVE: To determine whether the nocturnal fall in plasma adrenaline is a cause of nocturnal asthma. DESIGN: Double blind placebo controlled cross-over study. In the first experiment the nocturnal fall in plasma adrenaline at 4 am was corrected in 10 asthmatic subjects with an infusion of adrenaline after parasympathetic blockade with 30 micrograms/kg intravenous atropine. In the second experiment 11 asthmatic subjects showing similar variations in peak expiratory flow rate had the nocturnal fall in plasma adrenaline corrected by infusion before atropine was given. PATIENTS: Asthmatic subjects with a diurnal variation in home peak expiratory flow rate of greater than 20% for at least 75% of the time in the two weeks before the study. MAIN OUTCOME MEASURES: Peak expiratory flow rate and plasma adrenaline. RESULTS: Correction of the nocturnal fall in plasma adrenaline at 4 am to resting 4 pm levels did not alter peak expiratory flow rate either before or after parasympathetic blockade with atropine. CONCLUSION: A nighttime fall in plasma adrenaline is not a cause of nocturnal asthma.

Adult↗
PubMed1990

A system for ambulatory monitoring and computerised analysis of oesophageal motility and luminal pH.

It is often desirable to monitor physiological events from subjects free to participate in their normal daily activities. This is especially so where clinical symptoms are elusive or precipitated by specific activities. Through ambulatory monitoring, evidence of a causal relationship between recorded parameters and the spontaneous onset of symptoms may be revealed. We describe an ambulatory monitoring system which permits continuous 24-hour recording of oesophageal motility from three levels within the oesophagus together with luminal pH. Computerised analysis of the recorded data addresses many of the problems associated with processing the information accrued through long-term monitoring by providing an automatic comparison of patient data against either preselected values or those of the program's algorithm. Motility is quantified in terms of wave frequency, amplitude, duration and temporal relationship together with an integration of the wave area and mean pressure baseline. The corresponding pH data is quantified as frequency, duration and integrated area of pH excursions throughout a range of pH1-8. This system provides access to the entire patient record which may either be viewed on screen, analysed or output to a printer. Analysis yields a tabulated breakdown of motility and pH events from any section of the patient's record. Selective analysis readily permits statistical comparison of discrete time-matched samples corresponding to different patient circumstances.

Algorithms↗
PubMed1990

The distribution of substance P-containing nerves to the rat small intestine.

The concentrations of substance P-like immunoreactivity (SPLI) were measured in mesenteric nerves and vessels of control rats and rats treated with capsaicin during the second day of life. The results showed that there was a significantly higher concentration of SPLI in the jejunum than in the terminal ileum, and that there was a loss of SPLI from these tissues following neonatal treatment with capsaicin. The results suggest that the substance P-containing afferent innervation of the upper small intestine is greater than in the terminal ileum. The significance of these results to the Intensity or Summation hypothesis of visceral pain, and to the thresholds of visceral reflexes from different areas of the small intestine is discussed.

Animals↗